International Journal of Molecular Sciences

12 pages, 1798 KiB

Open AccessArticle

Interleukin-10 Facilitates Glutamatergic Synaptic Transmission and Homeostatic Plasticity in Cultured Hippocampal Neurons

by Miroslav N. Nenov, Maxim V. Konakov, Ilia Y. Teplov and Sergey G. Levin

Int. J. Mol. Sci. 2019, 20(13), 3375; https://doi.org/10.3390/ijms20133375 - 9 Jul 2019

Cited by 19 | Viewed by 4695

Abstract

Anti-inflammatory cytokines are known to exert neuroprotective action ameliorating aberrant neuronal network activity associated with inflammatory responses. Yet, it is still not fully understood if anti-inflammatory cytokines play a significant role in the regulation of synaptic activity under normal conditions. Thus, the aim [...] Read more.

Anti-inflammatory cytokines are known to exert neuroprotective action ameliorating aberrant neuronal network activity associated with inflammatory responses. Yet, it is still not fully understood if anti-inflammatory cytokines play a significant role in the regulation of synaptic activity under normal conditions. Thus, the aim of our study was to investigate the effect of Interleukin-10 (IL-10) on neuronal synaptic transmission and plasticity. For this we tested the effect of IL-10 on miniature excitatory postsynaptic currents (mEPSC) and intracellular Ca²⁺ responses using whole-cell patch clamp and fluorescence microscopy in 13–15 DIV primary hippocampal neuroglial culture. We found that IL-10 significantly potentiated basal glutamatergic excitatory synaptic transmission within 15 min after application. Obtained results revealed a presynaptic nature of the effect, as IL-10 in a dose-dependent manner significantly increased the frequency but not the amplitude of mEPSC. Further, we tested the effect of IL-10 on mEPSC in a model of homeostatic synaptic plasticity (HSP) induced by treatment of primary hippocampal culture with 1 µM of tetrodotoxin (TTX) for a 24 h. It was found that 15 min application of IL-10 at established HSP resulted in enhanced mEPSC frequency, thus partially compensating for a decrease in the mEPSC frequency associated with TTX-induced HSP. Next, we studied if IL-10 can influence induction of HSP. We found that co-incubation of IL-10 with 1 µM of TTX for 24 h induced synaptic scaling, significantly increasing the amplitude of mEPSC and Ca²⁺ responses to application of the AMPA agonist, 5-Fluorowillardiine, thus facilitating a compensatory postsynaptic mechanism at HSP condition. Our results indicate that IL-10 potentiates synaptic activity in a dose- and time-dependent manner exerting both presynaptic (short-term exposure) and postsynaptic (long-term exposure) action. Obtained results demonstrate involvement of IL-10 in the regulation of basal glutamatergic synaptic transmission and plasticity at normal conditions. Full article

(This article belongs to the Special Issue Glutamate Receptors in Health and Disease)

► Show Figures

Figure 1

20 pages, 493 KiB

Open AccessReview

Glut 1 in Cancer Cells and the Inhibitory Action of Resveratrol as A Potential Therapeutic Strategy

by Angara Zambrano, Matías Molt, Elena Uribe and Mónica Salas

Int. J. Mol. Sci. 2019, 20(13), 3374; https://doi.org/10.3390/ijms20133374 - 9 Jul 2019

Cited by 151 | Viewed by 14436

Abstract

An important hallmark in cancer cells is the increase in glucose uptake. GLUT1 is an important target in cancer treatment because cancer cells upregulate GLUT1, a membrane protein that facilitates the basal uptake of glucose in most cell types, to ensure the flux [...] Read more.

An important hallmark in cancer cells is the increase in glucose uptake. GLUT1 is an important target in cancer treatment because cancer cells upregulate GLUT1, a membrane protein that facilitates the basal uptake of glucose in most cell types, to ensure the flux of sugar into metabolic pathways. The dysregulation of GLUT1 is associated with numerous disorders, including cancer and metabolic diseases. There are natural products emerging as a source for inhibitors of glucose uptake, and resveratrol is a molecule of natural origin with many properties that acts as antioxidant and antiproliferative in malignant cells. In the present review, we discuss how GLUT1 is involved in the general scheme of cancer cell metabolism, the mechanism of glucose transport, and the importance of GLUT1 structure to understand the inhibition process. Then, we review the current state-of-the-art of resveratrol and other natural products as GLUT1 inhibitors, focusing on those directed at treating different types of cancer. Targeting GLUT1 activity is a promising strategy for the development of drugs aimed at treating neoplastic growth. Full article

(This article belongs to the Special Issue Health Benefits of Resveratrol)

► Show Figures

Graphical abstract

20 pages, 1410 KiB

Open AccessReview

Beyond N-Cadherin, Relevance of Cadherins 5, 6 and 17 in Cancer Progression and Metastasis

by J. Ignacio Casal and Rubén A. Bartolomé

Int. J. Mol. Sci. 2019, 20(13), 3373; https://doi.org/10.3390/ijms20133373 - 9 Jul 2019

Cited by 43 | Viewed by 11553

Abstract

Cell-cell adhesion molecules (cadherins) and cell-extracellular matrix adhesion proteins (integrins) play a critical role in the regulation of cancer invasion and metastasis. Although significant progress has been made in the characterization of multiple members of the cadherin superfamily, most of the published work [...] Read more.

Cell-cell adhesion molecules (cadherins) and cell-extracellular matrix adhesion proteins (integrins) play a critical role in the regulation of cancer invasion and metastasis. Although significant progress has been made in the characterization of multiple members of the cadherin superfamily, most of the published work continues to focus in the switch E-/N-cadherin and its role in the epithelial–mesenchymal transition. Here, we will discuss the structural and functional properties of a subset of cadherins (cadherin 17, cadherin 5 and cadherin 6) that have an RGD motif in the extracellular domains. This RGD motif is critical for the interaction with α2β1 integrin and posterior integrin pathway activation in cancer metastatic cells. However, other signaling pathways seem to be affected by RGD cadherin interactions, as will be discussed. The range of solid tumors with overexpression or “de novo” expression of one or more of these three cadherins is very wide (gastrointestinal, gynaecological and melanoma, among others), underscoring the relevance of these cadherins in cancer metastasis. Finally, we will discuss different evidences that support the therapeutic use of these cadherins by blocking their capacity to work as integrin ligands in order to develop new cures for metastatic patients. Full article

(This article belongs to the Special Issue Activations of Cadherin Signaling in Cancer)

► Show Figures

Graphical abstract

13 pages, 2203 KiB

Open AccessArticle

Role of B-Cell Translocation Gene 1 in the Pathogenesis of Endometriosis

by Jeong Sook Kim, Young Sik Choi, Ji Hyun Park, Jisun Yun, Soohyun Kim, Jae Hoon Lee, Bo Hyon Yun, Joo Hyun Park, Seok Kyo Seo, SiHyun Cho, Hyun-Soo Kim and Byung Seok Lee

Int. J. Mol. Sci. 2019, 20(13), 3372; https://doi.org/10.3390/ijms20133372 - 9 Jul 2019

Cited by 4 | Viewed by 3586

Abstract

Estrogen affects endometrial cellular proliferation by regulating the expression of the c-myc gene. B-cell translocation gene 1 (BTG1), a translocation partner of the c-myc, is a tumor suppressor gene that promotes apoptosis and negatively regulates cellular proliferation and cell-to-cell adhesion. The aim [...] Read more.

Estrogen affects endometrial cellular proliferation by regulating the expression of the c-myc gene. B-cell translocation gene 1 (BTG1), a translocation partner of the c-myc, is a tumor suppressor gene that promotes apoptosis and negatively regulates cellular proliferation and cell-to-cell adhesion. The aim of this study was to determine the role of BTG1 in the pathogenesis of endometriosis. BTG1 mRNA and protein expression was evaluated in eutopic and ectopic endometrium of 30 patients with endometriosis (endometriosis group), and in eutopic endometrium of 22 patients without endometriosis (control group). The effect of BTG1 downregulation on cellular migration, proliferation, and apoptosis was evaluated using transfection of primarily cultured human endometrial stromal cells (HESCs) with BTG1 siRNA. BTG1 mRNA expression level of eutopic and ectopic endometrium of endometriosis group were significantly lower than that of the eutopic endometrium of the control group. Migration and wound healing assays revealed that BTG1 downregulation resulted in a significant increase in migration potential of HESCs, characterized by increased expression of matrix metalloproteinase 2 (MMP2) and MMP9. Downregulation of BTG1 in HESCs significantly reduced Caspase 3 expression, indicating a decrease in apoptotic potential. In conclusion, our data suggest that downregulation of BTG1 plays an important role in the pathogenesis of endometriosis. Full article

(This article belongs to the Special Issue Endometriosis Research: From Bench to Bedside)

► Show Figures

Figure 1

12 pages, 523 KiB

Open AccessReview

The Problem of the Low Rates of CRISPR/Cas9-Mediated Knock-ins in Plants: Approaches and Solutions

by Serge M. Rozov, Natalya V. Permyakova and Elena V. Deineko

Int. J. Mol. Sci. 2019, 20(13), 3371; https://doi.org/10.3390/ijms20133371 - 9 Jul 2019

Cited by 34 | Viewed by 10361

Abstract

The main number of genome editing events in plant objects obtained during the last decade with the help of specific nucleases zinc finger (ZFN), transcription activator-like effector nucleases (TALEN), and clustered regularly interspaced short palindromic repeats (CRISPR)/Cas are the microindels causing frameshift and [...] Read more.

The main number of genome editing events in plant objects obtained during the last decade with the help of specific nucleases zinc finger (ZFN), transcription activator-like effector nucleases (TALEN), and clustered regularly interspaced short palindromic repeats (CRISPR)/Cas are the microindels causing frameshift and subsequent gene knock-out. The knock-ins of genes or their parts, i.e., the insertion of them into a target genome region, are between one and two orders of magnitude less frequent. First and foremost, this is associated with the specific features of the repair systems of higher eukaryotes and the availability of the donor template in accessible proximity during double-strand break (DSB) repair. This review briefs the main repair pathways in plants according to the aspect of their involvement in genome editing. The main methods for increasing the frequency of knock-ins are summarized both along the homologous recombination pathway and non-homologous end joining, which can be used for plant objects. Full article

(This article belongs to the Section Molecular Plant Sciences)

► Show Figures

Graphical abstract

22 pages, 3255 KiB

Open AccessArticle

EGF Receptor Stalls upon Activation as Evidenced by Complementary Fluorescence Correlation Spectroscopy and Fluorescence Recovery after Photobleaching Measurements

by György Vámosi, Elza Friedländer-Brock, Shehu M. Ibrahim, Roland Brock, János Szöllősi and György Vereb

Int. J. Mol. Sci. 2019, 20(13), 3370; https://doi.org/10.3390/ijms20133370 - 9 Jul 2019

Cited by 9 | Viewed by 3768

Abstract

To elucidate the molecular details of the activation-associated clustering of epidermal growth factor receptors (EGFRs), the time course of the mobility and aggregation states of eGFP tagged EGFR in the membranes of Chinese hamster ovary (CHO) cells was assessed by in situ mobility [...] Read more.

To elucidate the molecular details of the activation-associated clustering of epidermal growth factor receptors (EGFRs), the time course of the mobility and aggregation states of eGFP tagged EGFR in the membranes of Chinese hamster ovary (CHO) cells was assessed by in situ mobility assays. Fluorescence correlation spectroscopy (FCS) was used to probe molecular movements of small ensembles of molecules over short distances and time scales, and to report on the state of aggregation. The diffusion of larger ensembles of molecules over longer distances (and time scales) was investigated by fluorescence recovery after photobleaching (FRAP). Autocorrelation functions could be best fitted by a two-component diffusion model corrected for triplet formation and blinking. The slow, 100–1000 ms component was attributed to membrane localized receptors moving with free Brownian diffusion, whereas the fast, ms component was assigned to cytosolic receptors or their fragments. Upon stimulation with 50 nM EGF, a significant decrease from 0.11 to 0.07 μm²/s in the diffusion coefficient of membrane-localized receptors was observed, followed by recovery to the original value in ~20 min. In contrast, the apparent brightness of diffusing species remained the same. Stripe FRAP experiments yielded a decrease in long-range molecular mobility directly after stimulation, evidenced by an increase in the recovery time of the slow component from 13 to 21.9 s. Our observations are best explained by the transient attachment of ligand-bound EGFRs to immobile or slowly moving structures such as the cytoskeleton or large, previously photobleached receptor aggregates. Full article

(This article belongs to the Special Issue Imaging with Fluorescent Proteins)

► Show Figures

Figure 1

26 pages, 6223 KiB

Open AccessArticle

Development of Polymer Blend Electrolyte Membranes Based on Chitosan: Dextran with High Ion Transport Properties for EDLC Application

by Shujahadeen B. Aziz, Muhamad H. Hamsan, Mohd F. Z. Kadir, Wrya O. Karim and Ranjdar M. Abdullah

Int. J. Mol. Sci. 2019, 20(13), 3369; https://doi.org/10.3390/ijms20133369 - 9 Jul 2019

Cited by 102 | Viewed by 5865

Abstract

Solid polymer blend electrolyte membranes (SPBEM) composed of chitosan and dextran with the incorporation of various amounts of lithium perchlorate (LiClO₄) were synthesized. The complexation of the polymer blend electrolytes with the salt was examined using FTIR spectroscopy and X-ray diffraction [...] Read more.

Solid polymer blend electrolyte membranes (SPBEM) composed of chitosan and dextran with the incorporation of various amounts of lithium perchlorate (LiClO₄) were synthesized. The complexation of the polymer blend electrolytes with the salt was examined using FTIR spectroscopy and X-ray diffraction (XRD). The morphology of the SPBEs was also investigated using field emission scanning electron microscopy (FESEM). The ion transport behavior of the membrane films was measured using impedance spectroscopy. The membrane with highest LiClO₄ content was found to exhibit the highest conductivity of 5.16 × 10⁻³ S/cm. Ionic (t_i) and electronic (t_e) transference numbers for the highest conducting electrolyte were found to be 0.98 and 0.02, respectively. Electrochemical stability was estimated from linear sweep voltammetry and found to be up to ~2.3V for the Li⁺ ion conducting electrolyte. The only existence of electrical double charging at the surface of electrodes was evidenced from the absence of peaks in cyclic voltammetry (CV) plot. The discharge slope was observed to be almost linear, confirming the capacitive behavior of the EDLC. The performance of synthesized EDLC was studied using CV and charge–discharge techniques. The highest specific capacitance was achieved to be 8.7 F·g⁻¹ at 20th cycle. The efficiency (η) was observed to be at 92.8% and remained constant at 92.0% up to 100 cycles. The EDLC was considered to have a reasonable electrode-electrolyte contact, in which η exceeds 90.0%. It was determined that equivalent series resistance (R_esr) is quite low and varies from 150 to 180 Ω over the 100 cycles. Energy density (E_d) was found to be 1.21 Wh·kg⁻¹ at the 1st cycle and then remained stable at 0.86 Wh·kg⁻¹ up to 100 cycles. The interesting observation is that the value of P_d increases back to 685 W·kg⁻¹ up to 80 cycles. Full article

(This article belongs to the Special Issue Ion and Molecule Transport in Membrane Systems)

► Show Figures

Graphical abstract

13 pages, 3941 KiB

Open AccessArticle

Synthesis of Graphene Oxide-Fe₃O₄ Based Nanocomposites Using the Mechanochemical Method and in Vitro Magnetic Hyperthermia

by Venkatesha Narayanaswamy, Ihab M. Obaidat, Aleksandr S. Kamzin, Sachin Latiyan, Shilpee Jain, Hemant Kumar, Chandan Srivastava, Sulaiman Alaabed and Bashar Issa

Int. J. Mol. Sci. 2019, 20(13), 3368; https://doi.org/10.3390/ijms20133368 - 9 Jul 2019

Cited by 45 | Viewed by 6191

Abstract

The study presented in this work consists of two parts: The first part is the synthesis of Graphene oxide-Fe₃O₄ nanocomposites by a mechanochemical method which, is a mechanical process that is likely to yield extremely heterogeneous particles. The second part [...] Read more.

The study presented in this work consists of two parts: The first part is the synthesis of Graphene oxide-Fe₃O₄ nanocomposites by a mechanochemical method which, is a mechanical process that is likely to yield extremely heterogeneous particles. The second part includes a study on the efficacy of these Graphene oxide-Fe₃O₄ nanocomposites to kill cancerous cells. Iron powder, ball milled along with graphene oxide in a toluene medium, underwent a controlled oxidation process. Different phases of GO-Fe₃O₄ nanocomposites were obtained based on the composition used for milling. As synthesized nanocomposites were characterized by x-ray diffraction (XRD), alternating magnetic field (AFM), Raman spectroscopy, and a vibrating sample magnetometer (VSM). Additionally, the magnetic properties required to obtain high SAR values (Specific Absorption Rate-Power absorbed per unit mass of the magnetic nanocomposite in the presence of an applied magnetic field) for the composite were optimized by varying the milling time. Nanocomposites milled for different extents of time have shown differential behavior for magneto thermic heating. The magnetic composites synthesized by the ball milled method were able to retain the functional groups of graphene oxide. The efficacy of the magnetic nanocomposites for killing of cancerous cells is studied in vitro using HeLa cells in the presence of an AC (Alternating Current) magnetic field. The morphology of the HeLa cells subjected to 10 min of AC magnetic field changed considerably, indicating the death of the cells. Full article

(This article belongs to the Special Issue Magnetic Nanoparticles 2019)

► Show Figures

Figure 1

18 pages, 5550 KiB

Open AccessArticle

Protective Effects of Curcumin Ester Prodrug, Curcumin Diethyl Disuccinate against H₂O₂-Induced Oxidative Stress in Human Retinal Pigment Epithelial Cells: Potential Therapeutic Avenues for Age-Related Macular Degeneration

by Chawanphat Muangnoi, Umar Sharif, Pahweenvaj Ratnatilaka Na Bhuket, Pornchai Rojsitthisak and Luminita Paraoan

Int. J. Mol. Sci. 2019, 20(13), 3367; https://doi.org/10.3390/ijms20133367 - 9 Jul 2019

Cited by 37 | Viewed by 5014

Abstract

Oxidative stress-induced damage to the retinal pigmented epithelium (RPE), a specialised post-mitotic monolayer that maintains retinal homeostasis, contributes to the development of age-related macular degeneration (AMD). Curcumin (Cur), a naturally occurring antioxidant, was previously shown to have the ability to protect RPE cells [...] Read more.

Oxidative stress-induced damage to the retinal pigmented epithelium (RPE), a specialised post-mitotic monolayer that maintains retinal homeostasis, contributes to the development of age-related macular degeneration (AMD). Curcumin (Cur), a naturally occurring antioxidant, was previously shown to have the ability to protect RPE cells from oxidative stress. However, poor solubility and bioavailability makes Cur a poor therapeutic agent. As prodrug approaches can mitigate these limitations, we compared the protective properties of the Cur prodrug curcumin diethyl disuccinate (CurDD) against Cur in relation to oxidative stress induced in human ARPE-19 cells. Both CurDD and Cur significantly decreased H₂O₂-induced reactive oxygen species (ROS) production and protected RPE cells from oxidative stress-induced death. Both drugs exerted their protective effects through the modulation of p44/42 (ERK) and the involvement of downstream molecules Bax and Bcl-2. Additionally, the expression of antioxidant enzymes HO-1 and NQO1 was also enhanced in cells treated with CurDD and Cur. In all cases, CurDD was more effective than its parent drug against oxidative stress-induced damage to ARPE-19 cells. These findings highlight CurDD as a more potent drug compared to Cur against oxidative stress and indicate that its protective effects are exerted through modulation of key apoptotic and antioxidant molecular pathways. Full article

(This article belongs to the Special Issue Molecular Biology of Age-Related Macular Degeneration (AMD))

► Show Figures

Graphical abstract

14 pages, 2675 KiB

Open AccessArticle

Blockade of STAT3 Signaling Contributes to Anticancer Effect of 5-Acetyloxy-6,7,8,4′-Tetra-Methoxyflavone, a Tangeretin Derivative, on Human Glioblastoma Multiforme Cells

by Yen-Po Cheng, Shiming Li, Wan-Ling Chuang, Chia-Hsuan Li, Guan-Jun Chen, Ching-Chin Chang, Chi-Hung R. Or, Ping-Yi Lin and Chia-Che Chang

Int. J. Mol. Sci. 2019, 20(13), 3366; https://doi.org/10.3390/ijms20133366 - 9 Jul 2019

Cited by 22 | Viewed by 4863

Abstract

Glioblastoma multiforme (GBM) is the most aggressive primary brain tumor with poor prognosis, largely due to resistance to current radiotherapy and Temozolomide chemotherapy. The constitutive activation of Signal Transducer and Activator of Transcription 3 (STAT3) is evidenced as a pivotal driver of GBM [...] Read more.

Glioblastoma multiforme (GBM) is the most aggressive primary brain tumor with poor prognosis, largely due to resistance to current radiotherapy and Temozolomide chemotherapy. The constitutive activation of Signal Transducer and Activator of Transcription 3 (STAT3) is evidenced as a pivotal driver of GBM pathogenesis and therapy resistance, and hence, is a promising GBM drug target. 5-acetyloxy-6,7,8,4′-tetramethoxyflavone (5-AcTMF) is an acetylated derivative of Tangeretin which is known to exert anticancer effects on breast, colon, lung, and multiple myeloma; however, its effect on GBM remains elusive. Herein, we reported that 5-AcTMF suppressed the viability and clonogenicity along with inducing apoptosis in multiple human GBM cell lines. Mechanistic analyses further revealed that 5-AcTMF lowered the levels of Tyrosine 705-phosphorylated STAT3 (p-STAT3), a canonical marker of STAT3 activation, but also dampened p-STAT3 upregulation elicited by Interleukin-6. Notably, ectopic expression of dominant-active STAT3 impeded 5-AcTMF-induced suppression of viability and clonogenicity plus apoptosis induction in GBM cells, confirming the prerequisite of STAT3 blockage for the inhibitory action of 5-AcTMF on GBM cell survival and growth. Additionally, 5-AcTMF impaired the activation of STAT3 upstream kinase JAK2 but also downregulated antiapoptotic BCL-2 and BCL-xL in a STAT3-dependent manner. Moreover, the overexpression of either BCL-2 or BCL-xL abrogated 5-AcTMF-mediated viability reduction and apoptosis induction in GBM cells. Collectively, we, for the first time, revealed the anticancer effect of 5-AcTMF on GBM cells, which was executed via thwarting the JAK2-STAT3-BCL-2/BCL-xL signaling axis. Our findings further implicate the therapeutic potential of 5-AcTMF for GBM treatment. Full article

(This article belongs to the Special Issue Bioactive Phytochemicals for Cancer Prevention and Treatment)

► Show Figures

Graphical abstract

20 pages, 3168 KiB

Open AccessReview

Short Tandem Repeat Expansions and RNA-Mediated Pathogenesis in Myotonic Dystrophy

by Łukasz J. Sznajder and Maurice S. Swanson

Int. J. Mol. Sci. 2019, 20(13), 3365; https://doi.org/10.3390/ijms20133365 - 9 Jul 2019

Cited by 57 | Viewed by 11174

Abstract

Short tandem repeat (STR) or microsatellite, expansions underlie more than 50 hereditary neurological, neuromuscular and other diseases, including myotonic dystrophy types 1 (DM1) and 2 (DM2). Current disease models for DM1 and DM2 propose a common pathomechanism, whereby the transcription of mutant DMPK [...] Read more.

Short tandem repeat (STR) or microsatellite, expansions underlie more than 50 hereditary neurological, neuromuscular and other diseases, including myotonic dystrophy types 1 (DM1) and 2 (DM2). Current disease models for DM1 and DM2 propose a common pathomechanism, whereby the transcription of mutant DMPK (DM1) and CNBP (DM2) genes results in the synthesis of CUG and CCUG repeat expansion (CUG^exp, CCUG^exp) RNAs, respectively. These CUG^exp and CCUG^exp RNAs are toxic since they promote the assembly of ribonucleoprotein (RNP) complexes or RNA foci, leading to sequestration of Muscleblind-like (MBNL) proteins in the nucleus and global dysregulation of the processing, localization and stability of MBNL target RNAs. STR expansion RNAs also form phase-separated gel-like droplets both in vitro and in transiently transfected cells, implicating RNA-RNA multivalent interactions as drivers of RNA foci formation. Importantly, the nucleation and growth of these nuclear foci and transcript misprocessing are reversible processes and thus amenable to therapeutic intervention. In this review, we provide an overview of potential DM1 and DM2 pathomechanisms, followed by a discussion of MBNL functions in RNA processing and how multivalent interactions between expanded STR RNAs and RNA-binding proteins (RBPs) promote RNA foci assembly. Full article

(This article belongs to the Special Issue Myotonic Dystrophy: From Molecular Pathogenesis to Therapeutics)

► Show Figures

Figure 1

20 pages, 2651 KiB

Open AccessArticle

Integrated Approach Reveals Role of Mitochondrial Germ-Line Mutation F18L in Respiratory Chain, Oxidative Alterations, Drug Sensitivity, and Patient Prognosis in Glioblastoma

by Kathleen Keatley, Samuel Stromei-Cleroux, Tammy Wiltshire, Nina Rajala, Gary Burton, William V. Holt, D. Timothy J. Littlewood, Andrew G. Briscoe, Josephine Jung, Keyoumars Ashkan, Simon J. Heales, Geoffrey J. Pilkington, Brigitte Meunier, John E. McGeehan, Iain P. Hargreaves and Rhiannon E. McGeehan

Int. J. Mol. Sci. 2019, 20(13), 3364; https://doi.org/10.3390/ijms20133364 - 9 Jul 2019

Cited by 11 | Viewed by 4649

Abstract

Glioblastoma is the most common and malignant primary brain tumour in adults, with a dismal prognosis. This is partly due to considerable inter- and intra-tumour heterogeneity. Changes in the cellular energy-producing mitochondrial respiratory chain complex (MRC) activities are a hallmark of glioblastoma relative [...] Read more.

Glioblastoma is the most common and malignant primary brain tumour in adults, with a dismal prognosis. This is partly due to considerable inter- and intra-tumour heterogeneity. Changes in the cellular energy-producing mitochondrial respiratory chain complex (MRC) activities are a hallmark of glioblastoma relative to the normal brain, and associate with differential survival outcomes. Targeting MRC complexes with drugs can also facilitate anti-glioblastoma activity. Whether mutations in the mitochondrial DNA (mtDNA) that encode several components of the MRC contribute to these phenomena remains underexplored. We identified a germ-line mtDNA mutation (m. 14798T > C), enriched in glioblastoma relative to healthy controls, that causes an amino acid substitution F18L within the core mtDNA-encoded cytochrome b subunit of MRC complex III. F18L is predicted to alter corresponding complex III activity, and sensitivity to complex III-targeting drugs. This could in turn alter reactive oxygen species (ROS) production, cell behaviour and, consequently, patient outcomes. Here we show that, despite a heterogeneous mitochondrial background in adult glioblastoma patient biopsy-derived cell cultures, the F18L substitution associates with alterations in individual MRC complex activities, in particular a 75% increase in MRC complex II_III activity, and a 34% reduction in CoQ10, the natural substrate for MRC complex III, levels. Downstream characterisation of an F18L-carrier revealed an 87% increase in intra-cellular ROS, an altered cellular distribution of mitochondrial-specific ROS, and a 64% increased sensitivity to clomipramine, a repurposed MRC complex III-targeting drug. In patients, F18L-carriers that received the current standard of care treatment had a poorer prognosis than non-carriers (373 days vs. 415 days, respectively). Single germ-line mitochondrial mutations could predispose individuals to differential prognoses, and sensitivity to mitochondrial targeted drugs. Thus, F18L, which is present in blood could serve as a useful non-invasive biomarker for the stratification of patients into prognostically relevant groups, one of which requires a lower dose of clomipramine to achieve clinical effect, thus minimising side-effects. Full article

(This article belongs to the Special Issue Mitochondrial Dysfunction and Oxidative Damage)

► Show Figures

Graphical abstract

16 pages, 1391 KiB

Open AccessArticle

Network-Based Integrative Analysis of Genomics, Epigenomics and Transcriptomics in Autism Spectrum Disorders

by Noemi Di Nanni, Matteo Bersanelli, Francesca Anna Cupaioli, Luciano Milanesi, Alessandra Mezzelani and Ettore Mosca

Int. J. Mol. Sci. 2019, 20(13), 3363; https://doi.org/10.3390/ijms20133363 - 9 Jul 2019

Cited by 7 | Viewed by 5238

Abstract

Current studies suggest that autism spectrum disorders (ASDs) may be caused by many genetic factors. In fact, collectively considering multiple studies aimed at characterizing the basic pathophysiology of ASDs, a large number of genes has been proposed. Addressing the problem of molecular data [...] Read more.

Current studies suggest that autism spectrum disorders (ASDs) may be caused by many genetic factors. In fact, collectively considering multiple studies aimed at characterizing the basic pathophysiology of ASDs, a large number of genes has been proposed. Addressing the problem of molecular data interpretation using gene networks helps to explain genetic heterogeneity in terms of shared pathways. Besides, the integrative analysis of multiple omics has emerged as an approach to provide a more comprehensive view of a disease. In this work, we carry out a network-based meta-analysis of the genes reported as associated with ASDs by studies that involved genomics, epigenomics, and transcriptomics. Collectively, our analysis provides a prioritization of the large number of genes proposed to be associated with ASDs, based on genes’ relevance within the intracellular circuits, the strength of the supporting evidence of association with ASDs, and the number of different molecular alterations affecting genes. We discuss the presence of the prioritized genes in the SFARI (Simons Foundation Autism Research Initiative) database and in gene networks associated with ASDs by other investigations. Lastly, we provide the full results of our analyses to encourage further studies on common targets amenable to therapy. Full article

(This article belongs to the Special Issue Identification and Characterization of Genetic Components in Autism Spectrum Disorders 2019)

► Show Figures

Graphical abstract

14 pages, 899 KiB

Open AccessArticle

Genetic Structure Analysis of a Collection of Tunisian Durum Wheat Germplasm

by Amine Slim, Luciana Piarulli, Houda Chennaoui Kourda, Mustapha Rouaissi, Cyrine Robbana, Ramzi Chaabane, Domenico Pignone, Cinzia Montemurro and Giacomo Mangini

Int. J. Mol. Sci. 2019, 20(13), 3362; https://doi.org/10.3390/ijms20133362 - 9 Jul 2019

Cited by 25 | Viewed by 4696

Abstract

The Tunisian durum wheat germplasm includes modern cultivars and traditional varieties that are still cultivated in areas where elite cultivars or intensive cultivation systems are not suitable. Within the frame of a collection program of the National Gene Bank of Tunisia (NGBT), durum [...] Read more.

The Tunisian durum wheat germplasm includes modern cultivars and traditional varieties that are still cultivated in areas where elite cultivars or intensive cultivation systems are not suitable. Within the frame of a collection program of the National Gene Bank of Tunisia (NGBT), durum wheat germplasm was collected from different Tunisian agro-ecological zones. The collected samples were studied using simple sequence repeats (SSRs) markers to explore the genetic diversity and evaluate the genetic structure in Tunisian germplasm. The results demonstrated significant diversity in the Tunisian durum wheat germplasm, with clear differentiation between traditional varieties and modern cultivars. The population structure analysis allowed the identification of five subpopulations, two of which appear to be more strongly represented in germplasm collected in central and southern Tunisia, where environmental conditions at critical development phases of the plant are harsher. Moreover these subpopulations are underrepresented in modern varieties, suggesting that traits of adaptation useful for breeding more resilient varieties might be present in central and southern germplasm. Moreover, our results will support, the activity of in situ on farm conservation of Tunisian durum wheat germplasm started by the National Gene Bank of Tunisia along with the ex situ approach. Full article

(This article belongs to the Special Issue Genome Editing in Plants)

► Show Figures

Graphical abstract

13 pages, 2685 KiB

Open AccessArticle

Calcium Homeostasis Is Modified in Skeletal Muscle Fibers of Small Ankyrin1 Knockout Mice

by Enrico Pierantozzi, Péter Szentesi, Dána Al-Gaadi, Tamás Oláh, Beatrix Dienes, Mónika Sztretye, Daniela Rossi, Vincenzo Sorrentino and László Csernoch

Int. J. Mol. Sci. 2019, 20(13), 3361; https://doi.org/10.3390/ijms20133361 - 9 Jul 2019

Cited by 7 | Viewed by 4323

Abstract

Small Ankyrins (sAnk1) are muscle-specific isoforms generated by the Ank1 gene that participate in the organization of the sarcoplasmic reticulum (SR) of striated muscles. Accordingly, the volume of SR tubules localized around the myofibrils is strongly reduced in skeletal muscle fibers of 4- [...] Read more.

Small Ankyrins (sAnk1) are muscle-specific isoforms generated by the Ank1 gene that participate in the organization of the sarcoplasmic reticulum (SR) of striated muscles. Accordingly, the volume of SR tubules localized around the myofibrils is strongly reduced in skeletal muscle fibers of 4- and 10-month-old sAnk1 knockout (KO) mice, while additional structural alterations only develop with aging. To verify whether the lack of sAnk1 also alters intracellular Ca²⁺ handling, cytosolic Ca²⁺ levels were analyzed in stimulated skeletal muscle fibers from 4- and 10-month-old sAnk1 KO mice. The SR Ca²⁺ content was reduced in sAnk1 KO mice regardless of age. The amplitude of the Ca²⁺ transients induced by depolarizing pulses was decreased in myofibers of sAnk1 KO with respect to wild type (WT) fibers, while their voltage dependence was not affected. Furthermore, analysis of spontaneous Ca²⁺ release events (sparks) on saponin-permeabilized muscle fibers indicated that the frequency of sparks was significantly lower in fibers from 4-month-old KO mice compared to WT. Furthermore, both the amplitude and spatial spread of sparks were significantly smaller in muscle fibers from both 4- and 10-month-old KO mice compared to WT. These data suggest that the absence of sAnk1 results in an impairment of SR Ca²⁺ release, likely as a consequence of a decreased Ca²⁺ store due to the reduction of the SR volume in sAnk1 KO muscle fibers. Full article

(This article belongs to the Collection Feature Papers in “Molecular Biology”)

► Show Figures

Figure 1

12 pages, 3457 KiB

Open AccessArticle

Metabolomic Studies of Tissue Injury in Nonhuman Primates Exposed to Gamma-Radiation

by Amrita K. Cheema, Khyati Y. Mehta, Meena U. Rajagopal, Stephen Y. Wise, Oluseyi O. Fatanmi and Vijay K. Singh

Int. J. Mol. Sci. 2019, 20(13), 3360; https://doi.org/10.3390/ijms20133360 - 9 Jul 2019

Cited by 31 | Viewed by 3259

Abstract

Exposure to ionizing radiation induces a complex cascade of systemic and tissue-specific responses that lead to functional impairment over time in the surviving population. However, due to the lack of predictive biomarkers of tissue injury, current methods for the management of survivors of [...] Read more.

Exposure to ionizing radiation induces a complex cascade of systemic and tissue-specific responses that lead to functional impairment over time in the surviving population. However, due to the lack of predictive biomarkers of tissue injury, current methods for the management of survivors of radiation exposure episodes involve monitoring of individuals over time for the development of adverse clinical symptoms and death. Herein, we report on changes in metabolomic and lipidomic profiles in multiple tissues of nonhuman primates (NHPs) that were exposed to a single dose of 7.2 Gy whole-body ⁶⁰Co γ-radiation that either survived or succumbed to radiation toxicities over a 60-day period. This study involved the delineation of the radiation effects in the liver, kidney, jejunum, heart, lung, and spleen. We found robust metabolic changes in the kidney and liver and modest changes in other tissue types at the 60-day time point in a cohort of NHPs. Remarkably, we found significant elevation of long-chain acylcarnitines in animals that were exposed to radiation across multiple tissue types underscoring the role of this class of metabolites as a generic indicator of radiation-induced normal tissue injury. These studies underscore the utility of a metabolomics approach for delineating anticipatory biomarkers of exposure to ionizing radiation. Full article

(This article belongs to the Section Biochemistry)

► Show Figures

Graphical abstract

15 pages, 2506 KiB

Open AccessArticle

CRISPR-Cas9 Mediated TSPO Gene Knockout alters Respiration and Cellular Metabolism in Human Primary Microglia Cells

by Vladimir M. Milenkovic, Dounia Slim, Stefanie Bader, Victoria Koch, Elena-Sofia Heinl, David Alvarez-Carbonell, Caroline Nothdurfter, Rainer Rupprecht and Christian H. Wetzel

Int. J. Mol. Sci. 2019, 20(13), 3359; https://doi.org/10.3390/ijms20133359 - 9 Jul 2019

Cited by 41 | Viewed by 7239

Abstract

The 18 kDa translocator protein (TSPO) is an evolutionary conserved cholesterol binding protein localized in the outer mitochondrial membrane. It has been implicated in the regulation of various cellular processes including oxidative stress, proliferation, apoptosis, and steroid hormone biosynthesis. Since the expression of [...] Read more.

The 18 kDa translocator protein (TSPO) is an evolutionary conserved cholesterol binding protein localized in the outer mitochondrial membrane. It has been implicated in the regulation of various cellular processes including oxidative stress, proliferation, apoptosis, and steroid hormone biosynthesis. Since the expression of TSPO in activated microglia is upregulated in various neuroinflammatory and neurodegenerative disorders, we set out to examine the role of TSPO in an immortalized human microglia C20 cell line. To this end, we performed a dual approach and used (i) lentiviral shRNA silencing to reduce TSPO expression, and (ii) the CRISPR/Cas9 technology to generate complete TSPO knockout microglia cell lines. Functional characterization of control and TSPO knockdown as well as knockout cells, revealed only low de novo steroidogenesis in C20 cells, which was not dependent on the level of TSPO expression or influenced by the treatment with TSPO-specific ligands. In contrast to TSPO knockdown C20 cells, which did not show altered mitochondrial function, the TSPO deficient knockout cells displayed a significantly decreased mitochondrial membrane potential and cytosolic Ca²⁺ levels, as well as reduced respiratory function. Performing the rescue experiment by lentiviral overexpression of TSPO in knockout cells, increased oxygen consumption and restored respiratory function. Our study provides further evidence for a significant role of TSPO in cellular and mitochondrial metabolism and demonstrates that different phenotypes of mitochondrial function are dependent on the level of TSPO expression. Full article

(This article belongs to the Special Issue TSPO and Brain Disorders)

► Show Figures

Graphical abstract

31 pages, 3342 KiB

Open AccessArticle

High Functioning Autism with Missense Mutations in Synaptotagmin-Like Protein 4 (SYTL4) and Transmembrane Protein 187 (TMEM187) Genes: SYTL4- Protein Modeling, Protein-Protein Interaction, Expression Profiling and MicroRNA Studies

by Syed K. Rafi, Alberto Fernández-Jaén, Sara Álvarez, Owen W. Nadeau and Merlin G. Butler

Int. J. Mol. Sci. 2019, 20(13), 3358; https://doi.org/10.3390/ijms20133358 - 9 Jul 2019

Cited by 14 | Viewed by 5687

Abstract

We describe a 7-year-old male with high functioning autism spectrum disorder (ASD) and maternally-inherited rare missense variant of Synaptotagmin-like protein 4 (SYTL4) gene (Xq22.1; c.835C>T; p.Arg279Cys) and an unknown missense variant of Transmembrane protein 187 (TMEM187) gene (Xq28; c.708G>T; p. [...] Read more.

We describe a 7-year-old male with high functioning autism spectrum disorder (ASD) and maternally-inherited rare missense variant of Synaptotagmin-like protein 4 (SYTL4) gene (Xq22.1; c.835C>T; p.Arg279Cys) and an unknown missense variant of Transmembrane protein 187 (TMEM187) gene (Xq28; c.708G>T; p. Gln236His). Multiple in-silico predictions described in our study indicate a potentially damaging status for both X-linked genes. Analysis of predicted atomic threading models of the mutant and the native SYTL4 proteins suggest a potential structural change induced by the R279C variant which eliminates the stabilizing Arg279-Asp60 salt bridge in the N-terminal half of the SYTL4, affecting the functionality of the protein’s critical RAB-Binding Domain. In the European (Non-Finnish) population, the allele frequency for this variant is 0.00042. The SYTL4 gene is known to directly interact with several members of the RAB family of genes, such as, RAB27A, RAB27B, RAB8A, and RAB3A which are known autism spectrum disorder genes. The SYTL4 gene also directly interacts with three known autism genes: STX1A, SNAP25 and STXBP1. Through a literature-based analytical approach, we identified three of five (60%) autism-associated serum microRNAs (miRs) with high predictive power among the total of 298 mouse Sytl4 associated/predicted microRNA interactions. Five of 13 (38%) miRs were differentially expressed in serum from ASD individuals which were predicted to interact with the mouse equivalent Sytl4 gene. TMEM187 gene, like SYTL4, is a protein-coding gene that belongs to a group of genes which host microRNA genes in their introns or exons. The novel Q236H amino acid variant in the TMEM187 in our patient is near the terminal end region of the protein which is represented by multiple sequence alignments and hidden Markov models, preventing comparative structural analysis of the variant harboring region. Like SYTL4, the TMEM187 gene is expressed in the brain and interacts with four known ASD genes, namely, HCFC1; TMLHE; MECP2; and GPHN. TMM187 is in linkage with MECP2, which is a well-known determinant of brain structure and size and is a well-known autism gene. Other members of the TMEM gene family, TMEM132E and TMEM132D genes are associated with bipolar and panic disorders, respectively, while TMEM231 is a known syndromic autism gene. Together, TMEM187 and SYTL4 genes directly interact with recognized important ASD genes, and their mRNAs are found in extracellular vesicles in the nervous system and stimulate target cells to translate into active protein. Our evidence shows that both these genes should be considered as candidate genes for autism. Additional biological testing is warranted to further determine the pathogenicity of these gene variants in the causation of autism. Full article

(This article belongs to the Special Issue Identification and Characterization of Genetic Components in Autism Spectrum Disorders 2019)

► Show Figures

Figure 1

13 pages, 2690 KiB

Open AccessCommunication

Determination of the Role and Active Sites of PKC-Delta-Like from Lamprey in Innate Immunity

by Yang Xu, Huan Zhao, Yang Tian, Kaixia Ren, Nan Zheng and Qingwei Li

Int. J. Mol. Sci. 2019, 20(13), 3357; https://doi.org/10.3390/ijms20133357 - 9 Jul 2019

Cited by 3 | Viewed by 2773

Abstract

Protein kinase C-δ (PKC-δ) is an important protein in the immune system of higher vertebrates. Lampreys, as the most primitive vertebrates, have a uniquevariable lymphocyte receptor (VLR) immune system. PKC-δ-like is a crucial functional gene in lampreys and is highly expressed in their [...] Read more.

Protein kinase C-δ (PKC-δ) is an important protein in the immune system of higher vertebrates. Lampreys, as the most primitive vertebrates, have a uniquevariable lymphocyte receptor (VLR) immune system. PKC-δ-like is a crucial functional gene in lampreys and is highly expressed in their immune organs. In this study, lampreys were stimulated with different immunogens, and lipopolysaccharide (LPS) was found to increase the expression of PKC-δ-like. Overexpression of PKC-δ-like could also effectively activate the innate immune response. We further demonstrated that PKC-δ-like-CF, a catalytic fragment of PKC-δ-like, is responsible for activating the innate immune response, and Thr-211, which is Thr-419 of PKC-δ-like, was confirmed to be the key site affecting PKC-δ-like-CF activity. These results indicated that PKC-δ-like from lamprey may have an important role in the innate immune response. Full article

(This article belongs to the Section Molecular Immunology)

► Show Figures

Figure 1

15 pages, 3726 KiB

Open AccessArticle

Novel Salicylic Acid Analogs Induce a Potent Defense Response in Arabidopsis

by Ian Arthur Palmer, Huan Chen, Jian Chen, Ming Chang, Min Li, Fengquan Liu and Zheng Qing Fu

Int. J. Mol. Sci. 2019, 20(13), 3356; https://doi.org/10.3390/ijms20133356 - 8 Jul 2019

Cited by 27 | Viewed by 5240

Abstract

The master regulator of salicylic acid (SA)-mediated plant defense, NPR1 (NONEXPRESSER OF PR GENES 1) and its paralogs NPR3 and NPR4, act as SA receptors. After the perception of a pathogen, plant cells produce SA in the chloroplast. In the presence of SA, [...] Read more.

The master regulator of salicylic acid (SA)-mediated plant defense, NPR1 (NONEXPRESSER OF PR GENES 1) and its paralogs NPR3 and NPR4, act as SA receptors. After the perception of a pathogen, plant cells produce SA in the chloroplast. In the presence of SA, NPR1 protein is reduced from oligomers to monomers, and translocated into the nucleus. There, NPR1 binds to TGA, TCP, and WRKY transcription factors to induce expression of plant defense genes. A list of compounds structurally similar to SA was generated using ChemMine Tools and its Clustering Toolbox. Several of these analogs can induce SA-mediated defense and inhibit growth of Pseudomonas syringae in Arabidopsis. These analogs, when sprayed on Arabidopsis, can induce the accumulation of the master regulator of plant defense NPR1. In a yeast two-hybrid system, these analogs can strengthen the interactions among NPR proteins. We demonstrated that these analogs can induce the expression of the defense marker gene PR1. Furthermore, we hypothesized that these SA analogs could be potent tools against the citrus greening pathogen Candidatus liberibacter spp. In fact, our results suggest that the SA analogs we tested using Arabidopsis may also be effective for inducing a defense response in citrus. Several SA analogs consistently strengthened the interactions between citrus NPR1 and NPR3 proteins in a yeast two-hybrid system. In future assays, we plan to test whether these analogs avoid degradation by SA hydroxylases from plant pathogens. In future assays, we plan to test whether these analogs avoid degradation by SA hydroxylases from plant pathogens. Full article

(This article belongs to the Special Issue Salicylic Acid Signalling in Plants)

► Show Figures

Figure 1

12 pages, 216 KiB

Open AccessArticle

Correlations between Genetic Polymorphisms in Long Non-Coding RNA PRNCR1 and Gastric Cancer Risk in a Korean Population

by Jang Hee Hong, Eun-Heui Jin, Hyojin Kang, In Ae Chang, Sang-Il Lee and Jae Kyu Sung

Int. J. Mol. Sci. 2019, 20(13), 3355; https://doi.org/10.3390/ijms20133355 - 8 Jul 2019

Cited by 19 | Viewed by 2690

Abstract

We evaluated the association between prostate cancer non-coding RNA 1 (PRNCR1) polymorphisms and the risk of developing gastric cancer (GC) and GC subgroups in Korea. A case–control study was conducted with 437 GC patients and 357 healthy controls using a TaqMan [...] Read more.

We evaluated the association between prostate cancer non-coding RNA 1 (PRNCR1) polymorphisms and the risk of developing gastric cancer (GC) and GC subgroups in Korea. A case–control study was conducted with 437 GC patients and 357 healthy controls using a TaqMan genotyping assay. A chi-squared test, binary logistic regression, and genetic models were used to explore the association between five PRNCR1 polymorphisms and GC risk. After adjusting for gender and age, overall analyses using the recessive model indicated that the rs13252298 GG genotype was significantly associated with increased risk of intestinal-type gastric cancer (IGC). In the stratification analyses, the recessive model indicated that the rs1016343 TT genotype was significantly associated with decreased GC risk in individuals aged <60 years showing lymph node metastasis (LNM)-negative results. The rs13252298 GG genotype in the recessive model showed increased GC risk in subjects aged ≥60 years showing LNM-positive results and those aged ≥60 years in tumor stage III. In the dominant model, the rs16901946 combined genotype (AG/GG) was significantly associated with increased GC risk in subjects aged <60 years with tumor stage III. In the recessive model, the rs16901946 GG genotype was associated with decreased risk of GC and IGC in males aged ≥60 years. Thus, genetic variations in PRNCR1 may contribute to susceptibility to GC. Full article

(This article belongs to the Special Issue Gastric Cancers: Molecular Pathways and Candidate Biomarkers)

17 pages, 1835 KiB

Open AccessReview

by Ying Wang, Ling Leng, Md Khairul Islam, Fanghua Liu, Carol Sze Ki Lin and Shao-Yuan Leu

Int. J. Mol. Sci. 2019, 20(13), 3354; https://doi.org/10.3390/ijms20133354 - 8 Jul 2019

Cited by 24 | Viewed by 5189

Abstract

Cellulosomes are an extracellular supramolecular multienzyme complex that can efficiently degrade cellulose and hemicelluloses in plant cell walls. The structural and unique subunit arrangement of cellulosomes can promote its adhesion to the insoluble substrates, thus providing individual microbial cells with a direct competence [...] Read more.

Cellulosomes are an extracellular supramolecular multienzyme complex that can efficiently degrade cellulose and hemicelluloses in plant cell walls. The structural and unique subunit arrangement of cellulosomes can promote its adhesion to the insoluble substrates, thus providing individual microbial cells with a direct competence in the utilization of cellulosic biomass. Significant progress has been achieved in revealing the structures and functions of cellulosomes, but a knowledge gap still exists in understanding the interaction between cellulosome and lignocellulosic substrate for those derived from biorefinery pretreatment of agricultural crops. The cellulosomic saccharification of lignocellulose is affected by various substrate-related physical and chemical factors, including native (untreated) wood lignin content, the extent of lignin and xylan removal by pretreatment, lignin structure, substrate size, and of course substrate pore surface area or substrate accessibility to cellulose. Herein, we summarize the cellulosome structure, substrate-related factors, and regulatory mechanisms in the host cells. We discuss the latest advances in specific strategies of cellulosome-induced hydrolysis, which can function in the reaction kinetics and the overall progress of biorefineries based on lignocellulosic feedstocks. Full article

(This article belongs to the Special Issue Free and immobilized enzymes for biofuels, biosensing and bioremediation )

► Show Figures

Graphical abstract

24 pages, 10460 KiB

Open AccessArticle

Directed Evolution of P450 BM3 towards Functionalization of Aromatic O-Heterocycles

by Gustavo de Almeida Santos, Gaurao V. Dhoke, Mehdi D. Davari, Anna Joëlle Ruff and Ulrich Schwaneberg

Int. J. Mol. Sci. 2019, 20(13), 3353; https://doi.org/10.3390/ijms20133353 - 8 Jul 2019

Cited by 14 | Viewed by 5771

Abstract

The O-heterocycles, benzo-1,4-dioxane, phthalan, isochroman, 2,3-dihydrobenzofuran, benzofuran, and dibenzofuran are important building blocks with considerable medical application for the production of pharmaceuticals. Cytochrome P450 monooxygenase (P450) Bacillus megaterium 3 (BM3) wild type (WT) from Bacillus megaterium has low to no conversion of the [...] Read more.

The O-heterocycles, benzo-1,4-dioxane, phthalan, isochroman, 2,3-dihydrobenzofuran, benzofuran, and dibenzofuran are important building blocks with considerable medical application for the production of pharmaceuticals. Cytochrome P450 monooxygenase (P450) Bacillus megaterium 3 (BM3) wild type (WT) from Bacillus megaterium has low to no conversion of the six O-heterocycles. Screening of in-house libraries for active variants yielded P450 BM3 CM1 (R255P/P329H), which was subjected to directed evolution and site saturation mutagenesis of four positions. The latter led to the identification of position R255, which when introduced in the P450 BM3 WT, outperformed all other variants. The initial oxidation rate of nicotinamide adenine dinucleotide phosphate (NADPH) consumption increased ≈140-fold (WT: 8.3 ± 1.3 min⁻¹; R255L: 1168 ± 163 min⁻¹), total turnover number (TTN) increased ≈21-fold (WT: 40 ± 3; R255L: 860 ± 15), and coupling efficiency, ≈2.9-fold (WT: 8.8 ± 0.1%; R255L: 25.7 ± 1.0%). Computational analysis showed that substitution R255L (distant from the heme-cofactor) does not have the salt bridge formed with D217 in WT, which introduces flexibility into the I-helix and leads to a heme rearrangement allowing for efficient hydroxylation. Full article

(This article belongs to the Special Issue A Commemorative Issue to Celebrate the Nobel Success of Prof. Frances H. Arnold: Directed Evolution)

► Show Figures

Figure 1

16 pages, 972 KiB

Open AccessArticle

Quantifying the Potential for Future Gene Therapy to Lower Lifetime Risk of Polygenic Late-Onset Diseases

by Roman Teo Oliynyk

Int. J. Mol. Sci. 2019, 20(13), 3352; https://doi.org/10.3390/ijms20133352 - 8 Jul 2019

Cited by 2 | Viewed by 3961

Abstract

Gene therapy techniques and genetic knowledge may sufficiently advance, within the next few decades, to support prophylactic gene therapy for the prevention of polygenic late-onset diseases. The risk of these diseases may, hypothetically, be lowered by correcting the effects of a subset of [...] Read more.

Gene therapy techniques and genetic knowledge may sufficiently advance, within the next few decades, to support prophylactic gene therapy for the prevention of polygenic late-onset diseases. The risk of these diseases may, hypothetically, be lowered by correcting the effects of a subset of common low effect gene variants. In this paper, simulations show that if such gene therapy were to become technically possible; and if the incidences of the treated diseases follow the proportional hazards model with a multiplicative genetic architecture composed of a sufficient number of common small effect gene variants, then: (a) late-onset diseases with the highest familial heritability will have the largest number of variants available for editing; (b) diseases that currently have the highest lifetime risk, particularly those with the highest incidence rate continuing into older ages, will prove the most challenging cases in lowering lifetime risk and delaying the age of onset at a population-wide level; (c) diseases that are characterized by the lowest lifetime risk will show the strongest and longest-lasting response to such therapies; and (d) longer life expectancy is associated with a higher lifetime risk of these diseases, and this tendency, while delayed, will continue after therapy. Full article

(This article belongs to the Special Issue Genome Editing Therapies)

► Show Figures

Figure 1

14 pages, 2933 KiB

Open AccessReview

Diagnostic Roles of Postmortem cTn I and cTn T in Cardiac Death with Special Regard to Myocardial Infarction: A Systematic Literature Review and Meta-Analysis

by Zhipeng Cao, Mengyang Zhao, Chengyang Xu, Tianyi Zhang, Yuqing Jia, Tianqi Wang and Baoli Zhu

Int. J. Mol. Sci. 2019, 20(13), 3351; https://doi.org/10.3390/ijms20133351 - 8 Jul 2019

Cited by 30 | Viewed by 7892

Abstract

Background: Cardiac troponin I (cTn I) and cardiac troponin T (cTn T) are currently widely used as diagnostic biomarkers for myocardial injury caused by ischemic heart diseases in clinical and forensic medicine. However, no previous meta-analysis has summarized the diagnostic roles of postmortem [...] Read more.

Background: Cardiac troponin I (cTn I) and cardiac troponin T (cTn T) are currently widely used as diagnostic biomarkers for myocardial injury caused by ischemic heart diseases in clinical and forensic medicine. However, no previous meta-analysis has summarized the diagnostic roles of postmortem cTn I and cTn T. The aim of the present study was to meta-analyze the diagnostic roles of postmortem cTn I and cTn T for cardiac death in forensic medicine, present a systematic review of the previous literature, and determine the postmortem cut-off values of cTn I and cTn T. Methods: We searched multiple databases for the related literature, performed a meta-analysis to investigate the diagnostic roles of postmortem cardiac troponins, and analyzed the receiver operating characteristic (ROC) curve to determine their postmortem cut-off values. Results and Conclusions: The present meta-analysis demonstrated that postmortem cTn I and cTn T levels were increased in pericardial fluid and serum in cardiac death, especially in patients with acute myocardial infarction (AMI). We determined the postmortem cut-off value of cTn I in the pericardial fluid at 86.2 ng/mL, cTn I in serum at 9.5 ng/mL, and cTn T in serum at 8.025 ng/mL. Full article

(This article belongs to the Special Issue Myocardial Infarction and Myocardial Protection)

► Show Figures

Figure 1

18 pages, 259 KiB

Open AccessReview

Development of Drought-Tolerant Transgenic Wheat: Achievements and Limitations

by Shahbaz Khan, Sumera Anwar, Shaobo Yu, Min Sun, Zhenping Yang and Zhi-qiang Gao

Int. J. Mol. Sci. 2019, 20(13), 3350; https://doi.org/10.3390/ijms20133350 - 8 Jul 2019

Cited by 68 | Viewed by 10528

Abstract

Crop yield improvement is necessary to keep pace with increasing demand for food. Due to climatic variability, the incidence of drought stress at crop growth stages is becoming a major hindering factor to yield improvement. New techniques are required to increase drought tolerance [...] Read more.

Crop yield improvement is necessary to keep pace with increasing demand for food. Due to climatic variability, the incidence of drought stress at crop growth stages is becoming a major hindering factor to yield improvement. New techniques are required to increase drought tolerance along with improved yield. Genetic modification for increasing drought tolerance is highly desirable, and genetic engineering for drought tolerance requires the expression of certain stress-related genes. Genes have been identified which confer drought tolerance and improve plant growth and survival in transgenic wheat. However, less research has been conducted for the development of transgenic wheat as compared to rice, maize, and other staple food. Furthermore, enhanced tolerance to drought without any yield penalty is a major task of genetic engineering. In this review, we have focused on the progress in the development of transgenic wheat cultivars for improving drought tolerance and discussed the physiological mechanisms and testing of their tolerance in response to inserted genes under control or field conditions. Full article

(This article belongs to the Section Molecular Plant Sciences)

22 pages, 3214 KiB

Open AccessReview

DMSO Reductase Family: Phylogenetics and Applications of Extremophiles

by Jose María Miralles-Robledillo, Javier Torregrosa-Crespo, Rosa María Martínez-Espinosa and Carmen Pire

Int. J. Mol. Sci. 2019, 20(13), 3349; https://doi.org/10.3390/ijms20133349 - 8 Jul 2019

Cited by 32 | Viewed by 7270

Abstract

Dimethyl sulfoxide reductases (DMSO) are molybdoenzymes widespread in all domains of life. They catalyse not only redox reactions, but also hydroxylation/hydration and oxygen transfer processes. Although literature on DMSO is abundant, the biological significance of these enzymes in anaerobic respiration and the molecular [...] Read more.

Dimethyl sulfoxide reductases (DMSO) are molybdoenzymes widespread in all domains of life. They catalyse not only redox reactions, but also hydroxylation/hydration and oxygen transfer processes. Although literature on DMSO is abundant, the biological significance of these enzymes in anaerobic respiration and the molecular mechanisms beyond the expression of genes coding for them are still scarce. In this review, a deep revision of the literature reported on DMSO as well as the use of bioinformatics tools and free software has been developed in order to highlight the relevance of DMSO reductases on anaerobic processes connected to different biogeochemical cycles. Special emphasis has been addressed to DMSO from extremophilic organisms and their role in nitrogen cycle. Besides, an updated overview of phylogeny of DMSOs as well as potential applications of some DMSO reductases on bioremediation approaches are also described. Full article

(This article belongs to the Special Issue Extreme Microorganisms in the Nitrogen Cycle)

► Show Figures

Figure 1

17 pages, 7191 KiB

Open AccessReview

VDAC1 and the TSPO: Expression, Interactions, and Associated Functions in Health and Disease States

by Varda Shoshan-Barmatz, Srinivas Pittala and Dario Mizrachi

Int. J. Mol. Sci. 2019, 20(13), 3348; https://doi.org/10.3390/ijms20133348 - 8 Jul 2019

Cited by 70 | Viewed by 8628

Abstract

The translocator protein (TSPO), located at the outer mitochondrial membrane (OMM), serves multiple functions and contributes to numerous processes, including cholesterol import, mitochondrial metabolism, apoptosis, cell proliferation, Ca²⁺ signaling, oxidative stress, and inflammation. TSPO forms a complex with the voltage-dependent anion channel [...] Read more.

The translocator protein (TSPO), located at the outer mitochondrial membrane (OMM), serves multiple functions and contributes to numerous processes, including cholesterol import, mitochondrial metabolism, apoptosis, cell proliferation, Ca²⁺ signaling, oxidative stress, and inflammation. TSPO forms a complex with the voltage-dependent anion channel (VDAC), a protein that mediates the flux of ions, including Ca²⁺, nucleotides, and metabolites across the OMM, controls metabolism and apoptosis and interacts with many proteins. This review focuses on the two OMM proteins TSPO and VDAC1, addressing their structural interaction and associated functions. TSPO appears to be involved in the generation of reactive oxygen species, proposed to represent the link between TSPO activation and VDAC, thus playing a role in apoptotic cell death. In addition, expression of the two proteins in healthy brains and diseased states is considered, as is the relationship between TSPO and VDAC1 expression. Both proteins are over-expressed in in brains from Alzheimer’s disease patients. Finally, TSPO expression levels were proposed as a biomarker of some neuropathological settings, while TSPO-interacting ligands have been considered as a potential basis for drug development. Full article

(This article belongs to the Special Issue TSPO and Brain Disorders)

► Show Figures

Figure 1

14 pages, 10173 KiB

Open AccessArticle

NIRF-Molecular Imaging with Synovial Macrophages-Targeting Vsig4 Nanobody for Disease Monitoring in a Mouse Model of Arthritis

by Fang Zheng, Siyu Luo, Zhenlin Ouyang, Jinhong Zhou, Huanye Mo, Steve Schoonooghe, Serge Muyldermans, Patrick De Baetselier, Geert Raes and Yurong Wen

Int. J. Mol. Sci. 2019, 20(13), 3347; https://doi.org/10.3390/ijms20133347 - 8 Jul 2019

Cited by 13 | Viewed by 5099

Abstract

Nanobody against V-set and Ig domain-containing 4 (Vsig4) on tissue macrophages, such as synovial macrophages, could visualize joint inflammation in multiple experimental arthritis models via single-photon emission computed tomography imaging. Here, we further addressed the specificity and assessed the potential for arthritis monitoring [...] Read more.

Nanobody against V-set and Ig domain-containing 4 (Vsig4) on tissue macrophages, such as synovial macrophages, could visualize joint inflammation in multiple experimental arthritis models via single-photon emission computed tomography imaging. Here, we further addressed the specificity and assessed the potential for arthritis monitoring using near-infrared fluorescence (NIRF) Cy7-labeled Vsig4 nanobody (Cy7-Nb119). In vivo NIRF-imaging of collagen-induced arthritis (CIA) was performed using Cy7-Nb119. Signals obtained with Cy7-Nb119 or isotope control Cy7-NbBCII10 were compared in joints of naive mice versus CIA mice. In addition, pathological microscopy and fluorescence microscopy were used to validate the arthritis development in CIA. Cy7-Nb119 accumulated in inflamed joints of CIA mice, but not the naive mice. Development of symptoms in CIA was reflected in increased joint accumulation of Cy7-Nb119, which correlated with the conventional measurements of disease. Vsig4 is co-expressed with F4/80, indicating targeting of the increasing number of synovial macrophages associated with the severity of inflammation by the Vsig4 nanobody. NIRF imaging with Cy7-Nb119 allows specific assessment of inflammation in experimental arthritis and provides complementary information to clinical scoring for quantitative, non-invasive and economical monitoring of the pathological process. Nanobody labelled with fluorescence can also be used for ex vivo validation experiments using flow cytometry and fluorescence microscopy. Full article

(This article belongs to the Special Issue Macrophages in Inflammation 2019)

► Show Figures

Figure 1

15 pages, 4642 KiB

Open AccessArticle

Comparison between Slow Freezing and Vitrification for Human Ovarian Tissue Cryopreservation and Xenotransplantation

by Sanghoon Lee, Ki-Jin Ryu, Boram Kim, Dahyeon Kang, Yoon Young Kim and Tak Kim

Int. J. Mol. Sci. 2019, 20(13), 3346; https://doi.org/10.3390/ijms20133346 - 8 Jul 2019

Cited by 66 | Viewed by 8186

Abstract

Two methods for the cryopreservation of human ovarian tissue were compared using a xenotransplantation model to establish a safe and effective cryopreservation method. Ovarian tissues were obtained from women who underwent benign ovarian surgery in the gynecology research unit of a university hospital. [...] Read more.

Two methods for the cryopreservation of human ovarian tissue were compared using a xenotransplantation model to establish a safe and effective cryopreservation method. Ovarian tissues were obtained from women who underwent benign ovarian surgery in the gynecology research unit of a university hospital. The tissues were transplanted into 112 ovariectomized female severe combined immunodeficient mice 4 weeks after slow freezing or vitrification cryopreservation. Tissues were retrieved 4 weeks later. Primordial follicular counts decreased after cryopreservation and xenotransplantation, and were significantly higher in the slow freezing group than in the vitrification group (p < 0.001). Immunohistochemistry and TUNEL assay showed that the Ki-67 and CD31 markers of follicular proliferation and angiogenesis were higher in the slow freezing group (p < 0.001 and p = 0.006, respectively) and DNA damage was greater in the vitrification group (p < 0.001). Western blotting showed that vitrification increased cellular apoptosis. Anti-Müllerian hormone expression was low in transplanted samples subjected to both cryopreservation techniques. Electron microscopy revealed primordial follicle deformation in the vitrification group. Slow freezing for ovarian tissue cryopreservation is superior to vitrification in terms of follicle survival and growth after xenotransplantation. These results will be useful for fertility preservation in female cancer patients. Full article

(This article belongs to the Special Issue Molecular Pathophysiology of Uterine and Ovarian Diseases and Dysfunction)

► Show Figures

Figure 1

Journal Menu

Journal Browser

Int. J. Mol. Sci., Volume 20, Issue 13 (July-1 2019) – 263 articles

Further Information

Guidelines

MDPI Initiatives

Follow MDPI