Next Article in Journal
Phylogenetic, Microbiome, and Diet Characterisation of Wall Lizards in the Columbretes Archipelago (Spain): Clues for Their Conservation
Previous Article in Journal
Detection and Quantification of Forest-Agriculture Ecotones Caused by Returning Farmland to Forest Program Using Unmanned Aircraft Imagery
 
 
Search for Articles:
Title / Keyword
Author / Affiliation / Email
Journal
Article Type
 
 
Advanced Search
 
Section
Special Issue
Volume
Issue
Number
Page
 
Journals
Diversity
Volume 14
Issue 5
10.3390/d14050407
diversity-logo
Submit to this Journal Review for this Journal Propose a Special Issue
► Article Menu

Article Menu

share Share announcement Help format_quote Cite question_answer Discuss in SciProfiles thumb_up
...
Endorse textsms
...
Comment
first_page
settings
Order Article Reprints
Font Type:
Arial Georgia Verdana
Font Size:
Aa Aa Aa
Line Spacing:
Column Width:
Background:
Article

The UDP-Glycosyltransferase Gene Family in Achelura yunnanensis (Lepidoptera: Zygaenidae): Identification, Phylogeny, and Diverse Expression Patterns

by
Hai-Yan Xiao
,
Dan-Lu Chen
,
Ting-Ting Lu
,
Yu-Juan Yao
and
Nai-Yong Liu
*
Key Laboratory of Forest Disaster Warning and Control of Yunnan Province, Southwest Forestry University, Kunming 650224, China
*
Author to whom correspondence should be addressed.
Diversity 2022, 14(5), 407; https://doi.org/10.3390/d14050407
Submission received: 29 April 2022 / Revised: 18 May 2022 / Accepted: 20 May 2022 / Published: 21 May 2022
(This article belongs to the Section Animal Diversity)
Browse Figures
Versions Notes

Abstract

:
The caterpillars of the Lepidoptera are important herbivores as most of them belong to serious agricultural and forestry pests. To adapt to their habitats and feeding host plants, the larvae utilize uridine diphosphate (UDP)-glycosyltransferases (UGTs) to metabolize plant defensive compounds and insecticides. However, information on the UGT gene family in Achelura yunnanensis remains scarce. Here, we characterized the UGT genes through gene identification, phylogenic analyses, and comprehensive expression profiles regarding sexes, tissues, and stages. Transcriptome analyses led to the yields of 50 transcripts encoding UGTs in A. yunnanensis, representing a comparable gene number compared to those in other lepidopteran species. Sequence and phylogenetic analyses revealed a low amino acid identity of 28.23% among 31 full-length AyunUGTs, but some members shared relatively high conservation (>50% identities) with a phylogenetically clustered distribution. In addition, the majority of AyunUGTs possessed conserved residues involved in the catalysis and sugar-donor binding. Combining RNA sequencing and PCR approaches, a number of AyunUGTs were found to have the expression in chemosensory or detoxification tissues, possibly associated with the sensing of odorant molecules and the metabolism of toxic chemicals. More importantly, at least 27 AyunUGTs displayed detectable expression in reproductive tissues of both sexes. This study identifies candidate A. yunnanensis UGTs responsible for detoxification, olfaction, and reproduction, allowing us to address putative roles of UGTs in the adaptation of larvae to the habitats and feeding hosts.

Graphical Abstract

1. Introduction

The evolutionary adaptation of insects to habitats is associated with various biotransformation processes, especially for the detoxification pathway that is indispensable for herbivorous insects to detoxify the toxic chemicals derived from their food host plants or the external environment. Usually, these detoxification-related enzymes mainly refer to the phase I (cytochrome P450 monooxygenases, P450s, and carboxylesterases, COEs) and II (glutathione S-transferases, GSTs, and uridine diphosphate (UDP)-glycosyltransferases, UGTs), which are of utmost importance for the survival and reproduction of insects. The phytophagous insects utilize these enzymes to metabolize xenobiotics and endobiotics, which are often toxic to herbivores [1,2]. In comparison to the three superfamilies of P450s, GSTs, and COEs, UGTs in insects have received relatively little attention. As one of the most important detoxification enzymes, insect UGTs have been demonstrated to be mainly responsible for the metabolism of plant allelochemicals and insecticides [3,4,5].
Moths constitute the majority of the Lepidoptera, and their larvae are all herbivores [6,7]. To adapt to the constantly changing habitats and feeding host plants, they must cope with plant-produced or man-made chemicals via a xenobiotic-metabolizing enzyme system. Moth UGTs are capable of catalyzing the conjugation of small lipophilic compounds with sugars, including plant defensive compounds, pesticides, hormones, odorants, and other xenobiotics [4,8,9,10]. The biotransformation process makes the harmful substrates become more water-soluble and readily excretable, helping the caterpillars avoid plant toxins and insecticides. In Bombyx mori, a genome-wide analysis resulted in the yields of 42 UGT genes, of which they were mainly transcribed in reproductive and/or detoxification-related tissues [11]. Furthermore, some BmorUGTs responded to flavonoids, coumarins, and terpenoids, serving as key enzymes in the detoxification of plant toxic compounds and the degradation of odorant molecules [12,13]. In addition, the polyphagous noctuid species also employ the UGT enzymes to metabolize plant-derived defensive substrates. For example, a cotton-produced defensive compound of gossypol was glycosylated by HarmUGT40D1 and HarmUGT41B3 in Helicoverpa armigera, as an adaptation of the cotton bollworm larvae to a major feeding host plant [14]. In Spodoptera frugiperda, SfruUGT33F28 was responsible for the glycosylation of benzoxazinoids, the main defensive compounds derived from maize plants [15]. Similarly, three Helicoverpa species could overcome the defense of capsaicin, especially for the specialist H. assulta [16]. Beyond the detoxification of plant secondary metabolites, moth UGTs are involved in olfaction and insecticide resistance [8,9,17,18,19].
The zygaenid moth, Achelura yunnanensis, is an oligophagous insect herbivore with the larvae feeding solely on the Rosaceae family, particularly its favorable hosts of Cerasus yunnanensis and Photinia glomerata. Therefore, it is considered to be a serious pest of garden plants that generally eats the leaves of host plants and then moves to other hosts for continuing damage [20,21,22]. Like other moths, A. yunnanensis caterpillars are able to metabolize a variety of toxic exogenous and endogenous compounds by detoxification-related tissues, including midguts, fat bodies, and Malpighian tubules. However, no information is available about the detoxification mechanism in this species. Here, we identified and characterized the UGT gene family from A. yunnanensis, one of the most crucial detoxification enzymes. Through sequence and phylogenetic analyses, and expression profiling characteristics, this work identifies molecular candidates involved in detoxification, olfaction, or reproduction, and provides insights into the adaptation of the A. yunnanensis larvae to the feeding host plants.

2. Materials and Methods

2.1. Insects and Tissues

A. yunnanensis larvae rearing was described in detail by Li et al. (2021). Briefly, newly emerged adults were separated by sex, and males and females were maintained in individual cages, supplying with 10% sugar solution. To examine the expression profiles of UGT genes, the following tissues were collected: the 3rd day eggs after virgin females laid eggs; antennae, heads (removing antennae), maxillary palps, midguts, fat bodies, and Malpighian tubules from 4th instar larvae; the 3rd day pupae of both sexes after pupation; adult tissues from 3-day-old moths, including antennae, heads (removing antennae), thoraxes, abdomens, legs, and wings of both sexes, and female pheromone glands; and reproductive tissues from 3-day-old adults, comprising accessory glands (MAG), testes (Te), seminal vesicles (SV), and ejaculatory ducts (ED) of males, as well as FAG, bursa copulatrix (BC), ovaries (Ov), and spermathecae with spermathecal glands (SS). The reproductive tissues were isolated and related fat bodies were cleaned in phosphate-buffered saline (PBS, pH 7.4). The collected tissues were immediately frozen in liquid nitrogen and stored at −80 °C.

2.2. RNA Extraction and Synthesis of First-Strand cDNA

TRIzol Regent (Ambion, Life Technologies, Carlsbad, CA, USA) was used to extract total RNA from each tissue of A. yunnanensis, along with the protocols. RNA concentration and quality were checked on an agarose gel (1%, w/v) and a NanoDrop 1000 Spectrophotometer (Thermo Fisher Scientific, San Jose, CA, USA). Genomic DNA from purified RNA was removed using gDNA Eraser at 42 °C for 2 min. Then, reverse transcription was carried out using a PrimeScript RT Reagent Kit (TaKaRa, Dalian, China) at 37 °C for 15 min, and stopped at 85 °C for 5 s. For reverse transcription (RT)–PCR analyses, 5-fold dilutions of cDNA templates were used, and 15-fold for quantitative real-time PCR (qPCR).

2.3. Identification of Candidate UGTs in A. yunnanensis

Initially, UGT protein sequences of four lepidopteran species, including B. mori, H. armigera, Manduca sexta, and Athetis lepigone, were pooled as a local sequence database. Using these sequences as queries, we identified the UGT genes in A. yunnanensis by TBLASTN against the transcriptomes previously published [20,21]. To confirm whether newly identified AyunUGTs were members of the Glycosyltransferase_GTB-type superfamily, BLAST searches were repeated with each new AyunUGT against the National Center for Biotechnology Information (NCBI) Non-redundant (Nr) protein sequences database.

2.4. Sequence and Phylogenetic Analysis of UGTs

Open reading frames (ORFs) of genes were predicted using an online tool ORF Finder in the NCBI website (https://www.ncbi.nlm.nih.gov/orffinder/ (accessed on 4 March 2022)). Isoelectric point (pI) and molecular weight (Mw) of full-length UGTs were computed by the Compute pI/Mw tool (https://web.expasy.org/compute_pi/ (accessed on 4 March 2022)). The prediction of signal peptides and N-glycosylation sites (NPS) were conducted using the SignalP version 6.0 (Technical University of Denmark, Kongens Lyngby, Denmark) [23] and NetNGlyc version 1.0 (Technical University of Denmark, Kongens Lyngby, Denmark) servers (https://services.healthtech.dtu.dk/service.php?NetNGlyc-1.0 (accessed on 4 March 2022)), respectively. A multiple alignment program MAFFT V7.450 was employed to align the protein sequences and to calculate amino acid identities [24]. If necessary, the alignments were manually adjusted and edited using Jalview version 2.8 (University of Dundee, Scotland, UK) [25].
The maximum-likelihood tree was built using FastTree v2.1.12 (Lawrence Berkeley National Lab, Berkeley, CA, USA), under the Whelan-And-Goldman (WAG) 2001 model with the Gamma20 likelihood and 1000 replications [26]. To determine the distribution of UGT genes typically clustering into monophylogenetic clades on chromosomes, the genes in B. mori, M. sexta, and H. armigera were mapped onto their respective reference genomes by TBLASTN, implemented in Geneious R10.1.3 (https://www.geneious.com/ (accessed on 10 March 2022)). The genomic sequences of three species were retrieved from the NCBI Genome database, referring to B. mori (Bmori_2016v1.0, accession number: GCF_014905235.1) [27], M. sexta (JHU_Msex_v1.0, accession number: GCA_000262585.1) [28], and H. armigera (Harm_1.0, accession number: GCA_002156985.1) [29].

2.5. Expression Profiling Analysis of UGT Genes in A. yunnanensis

Based on the sequenced transcriptomes of 13 tissues [20,21], we first mapped clean reads onto the unigene transcriptomes using Bowtie2 [30]. The expression values of genes were calculated by RSEM v1.2.15 with default parameters [31], as normalized by FPKM (fragments per kilobase of transcript per million mapped reads) [32]. Next, RT–PCR was employed to determine the expression profiles of 31 full-length UGTs in different tissues of A. yunnanensis. The quality and quantity of cDNA templates were measured using a reference gene, ribosomal protein L8 (AyunRPL8) [20]. The primers were designed by Primer Premier 5 (PREMIER Biosoft International, Palo Alto, CA, USA), with the expected amplification sizes of 400–500 bp (Table S1). The reaction was performed with a total volume of 25 μL, following a Taq DNA Polymerase Kit’s instruction (TaKaRa, Dalian, China). The conditions were as follows: initial denaturation at 94 °C for 4 min, followed by 35 cycles of 94 °C for 30 s, 58 °C for 30 s, 72 °C for 40 s, and a final extension at 72 °C for 5 min. PCR amplification products were analyzed on 1.5% (w/v) agarose gels.
In addition, we chose six candidate UGT genes that showed high expression in main chemosensory (larval antennae, larval maxillary palps, antennae, and legs of both sexes) or detoxification (larval midguts, fat bodies, and Malpighian tubules) tissues to examine their relative expression levels. Gene-specific primers were designed by Beacon Designer 8.14 (PREMIER Biosoft International, Palo Alto, CA, USA) (Table S1). qPCR was performed with qTOWER 2.2, according to Bestar® SybrGreen qPCR Mastermix’s suggestions (DBI® Bioscience, Ludwigshafen, Germany). In brief, a reaction mixture of 20 μL was prepared and run under the procedures: 95 °C for 2 min and 40 cycles at 95 °C for 10 s, 58 °C for 31 s, and 72 °C for 30 s. Three biological pools were run for each tissue. Two reference genes, AyunRPL8 and RPS4 [20], were used to normalize the relative expression of target genes using a Q-Gene method [33,34]. Statistical analyses in the data were conducted by one-way ANOVA followed by Fisher’s least significant difference (LSD) test. A significant level was set as p value < 0.05.

3. Results

3.1. Identification of A. yunnanensis UGT Genes

Combining Li et al. (2021) and Nuo et al. (2022) transcriptomic data, a BLAST-based homology search led to the yields of 50 transcripts encoding UGTs in A. yunnanensis. Out of the identified AyunUGTs, 31 relatives harbored complete ORFs, ranging from 1512 to 1641 bp. The remaining 19 genes were partial sequences, encoding 117 to 480 amino acids. Among the full-length AyunUGTs, most of them exhibited extremely low identities to each other at the amino acid level (<30%), with an average value of 28.23%. Exceptionally, four pairs of the proteins shared a high degree of conservation (>80% identities), i.e., AyunUGT8 and UGT12 (93.46%), AyunUGT15 and UGT31 (87.96%), AyunUGT9 and UGT28 (82.66%), as well as AyunUGT19 and UGT30 (82.43%). Like other insect UGTs, AyunUGTs had signal peptides (15–24 amino acids) and similar pI (6.37–9.50) and Mw (57.08–62.57 kDa). The identification of N-glycosylation sites revealed that all 31 full-length UGTs contained at least one site, particularly as many as six for AyunUGT6/17/21/29 (Table 1 and Supplementary Material Additional File S1).

3.2. Sequence and Phylogenetic Characteristics of A. yunnanensis UGTs

To identify conserved residues involved in the catalysis and sugar-donor binding, 31 full-length sequences were aligned. As expected, two catalytic residues were highly conserved, of which the majority of AyunUGTs had identical amino acids at two positions (the first: histidine, H, and the second: aspartic acid, D), with the exception of AyunUGT8/12/16/20/24. In human UGT2B7, eight residues interacting with sugars were as follows: serine–tryptophan–glutamine–glutamic acid (S–W–Q–E, nucleotide binding), threonine–H (T–H, phosphate binding), and D–Q (glucuronic acid binding) [35]. For the nucleotide binding, 30 AyunUGTs possessed the conserved pattern at all the four sites, except for AyunUGT16 (asparagine (N) instead of S at the first position). Nearly all the AyunUGTs had the same “D–Q” pattern with human UGT2B7, though not AyunUGT10 (D–H). In comparison, the residues involved in the binding of phosphates were less conserved, with 16 AyunUGTs presenting the “T–H” pattern. In addition, two donor binding regions (DBRs) containing a signature motif in DBR–2 exhibited high conservation. On the contrary, the transmembrane domain and cytoplasmic tail sequences were more divergent (Figure 1).
With the aligned protein sequence of 184 UGTs from four moths, the phylogenetic tree was generated. In the tree, moth UGTs could be classified into 14 different clades, 12 of which possessed at least one member from A. yunnanensis. None of the AyunUGTs were found in the UGT43 and UGT48 subfamilies. Remarkably, two large clusters were formed in A. yunnanensis, representing AyunUGT6/14/22/32/33/38/39/47 in UGT33 and AyunUGT9/11/15/24/28/31/34/35/36/40/43/45/46/48/49 in UGT40. Such species-specific expansions were also observed in the UGT33 subfamily from three other species, as each representative was presented in one species. To determine the distribution of the clustered genes on chromosomes, they were aligned to the corresponding genomes using BLAST. As a result, the UGT genes were situated on one or two scaffolds in close proximity. In B. mori, eight UGT members were located on chromosome 28 and shared a mean identity of 67.69%. H. armigera UGTs presented a major distribution on scaffolds 395 and 562, of which the latter harbored 14 relatives varying from 56.59% to 91.41% identities. In the case of 11 M. sexta UGTs, they were distributed on scaffolds 00405 and 00641. Moreover, it was noticed that MsexUGT33C1–C6 were six alternatively spliced variants that shared a common 3’–terminal region and more than 75% identities with each other. Among four moth species, the high conservation was obtained in UGT44, UGT47, and UGT50, with a single gene copy in each species. In particular, members within each subfamily shared over 60% identities (Figure 2).

3.3. Stage- and Sex-Specific Expression Profile of A. yunnanensis UGT Genes

As 10 transcripts encoding UGTs were identified exclusively from the initially assembled transcripts which were not clustered by Corset v1.05 [36], their FPKM values were unable to be calculated in various tissues. Thus, we constructed an expression profiling map of 40 AyunUGTs in 13 tissues. The majority of the genes were widely transcribed in tissues, such as AyunUGT1–3/5/7/16–22/25. Focusing on the genes specifically or highly expressed in tissues, it was found that six AyunUGTs had an antenna-enriched expression, including AyunUGT8/9/18/25/30/37/42/46 where AyunUGT8 appeared to be a male-biased relative in the antennae with a 5.12-fold difference compared to females. In contrast, AyunUGT22 expression was female-antenna-predominant. In addition, AyunUGT5, UGT41, and UGT48 were abundantly expressed in female abdomens, legs of both sexes, and male abdomens, respectively (Figure 3A).
To reconstruct and investigate the expression of 31 AyunUGTs with full-length sequences, nine additional tissues from different developmental stages were selected, including three detoxification-related tissues of larvae (midguts, fat bodies, and Malpighian tubules). Consistent with the results by RNA sequencing (RNA-Seq), most of the genes displayed a broad tissue expression. Intriguingly, the transcription of two genes was restricted to larval heads (AyunUGT11) and larval midguts (AyunUGT15). Some genes appeared to have a dominant expression in tissues, for example, AyunUGT4 and UGT31 expressed in larval midguts, AyunUGT6 expressed in larval midguts and fat bodies, and AyunUGT23 expressed in male abdomens. Out of the 31 AyunUGTs, 24 and 22 relatives were detected in antennae and maxillary palps of the larval stage, respectively, while 20 and 18 genes were presented separately in male and female antennae. In three main detoxification tissues, a number of AyunUGTs were found to have the expression in larval midguts, fat bodies, and Malpighian tubules with 26, 22, and 19 relatives, respectively (Figure 3B).
Based on the FPKM and RT–PCR results, we further selected six candidate UGT genes abundantly expressed in chemosensory and/or detoxification tissues to determine their relative levels. qPCR revealed that three genes (AyunUGT9, UGT12, and UGT18) were significantly enriched in chemosensory tissues and slightly presented in some detoxification tissues. AyunUGT9 had a significantly higher expression in larval antennae and maxillary palps in comparison to other tissues. In contrast, the dominant expression of AyunUGT18 was presented in the antennae of the adult stage. AyunUGT12 had the highest expression level in male legs among tested tissues. Notably, AyunUGT8 exhibited particularly high transcription in larval Malpighian tubules, with a 1751.21-fold difference relative to male antennae. The remaining two genes, AyunUGT25 and UGT30, had comparable transcription levels in adult antennae and larval Malpighian tubules (Figure 4).

3.4. Candidate UGT Genes in A. yunnanensis Involving Reproduction

Like most lepidopteran species, A. yunnanensis produces their offspring via sexual reproduction. As indicated in previous studies, reproductive-related genes in insects are rapidly evolving, greatly extending the gene repertoires associated with reproduction, including the UGT gene family [37,38,39]. Here, we surveyed the expression of 31 UGTs with full-length sequences in reproductive tissues. As shown in Figure 5A, the internal reproductive system of female A. yunnanensis was composed of a pair of accessory glands, four pairs of oviducts (ovary), one bursa copulatrix, and one spermatheca with a spermathecal gland. The reproductive system of male moths was made up of four portions, including a pair of accessory glands, one testis, one ejaculatory duct, and a pair of seminal vesicles. Out of the 31 genes, four AyunUGTs had no detectable transcription in eight reproductive tissues, i.e., AyunUGT11/14/29/31. The other 27 AyunUGTs were transcribed in at least one tissue, but eight of them only displayed weak bands such as AyunUGT8/12/15/17/21/23/27/28. Two genes, AyunUGT4 and UGT19, were presented in all tissues. Some genes appeared to be sex-specific transcripts in reproductive tissues, for example, AyunUGT2, UGT6, and UGT24 expressed in males, as well as AyunUGT9 and UGT13 expressed in females (Figure 5B).

4. Discussion

In Lepidoptera, the caterpillars feed on diverse plant species. As a positive feedback, the plants will synthesize and release the secondary metabolites to cope with the herbivores. With the coevolution of moths and plants, the larvae also develop a variety of strategies to overcome the plant toxins, such as metabolic resistance. After encountering plant allelochemicals, the expression of detoxification enzyme genes in larvae is regulated as an adaptation of the larvae to xenobiotics [1,2]. Like the herbivorous insects, the larvae of A. yunnanensis have evolved to recognize and eat specialized host plants. During the process, the UGT gene family associated with the detoxification and insecticide resistance is indispensable for the glycosylation of xenobiotics, including plant secondary metabolites and insecticides [12,14,15,40,41]. Our current study identified and characterized the UGT enzymes in A. yunnanensis. This work has constituted a crucial step underlying the detoxification mechanism and allows the exploration of the specialized feeding adaptation of the A. yunnanensis larvae to the Rosaceae plants.
Insect UGTs have several typical characteristics, including signal peptides and conserved catalytic and sugar-donor binding residues, as implied in gene identification and functions [4]. A. yunnanensis UGTs possessed most, if not all, of these features. Moreover, not only did the majority of key sites have identical residues across A. yunnanensis and other insects, but this conservation also existed in A. yunnanensis and humans [4,17,35,42,43]. Outside the key residues at the conserved positions, most of AyunUGTs shared an average of only 28.23% identities to each other, especially for the diverse N-terminal binding domain. The low identities were generally presented in other insects, such as B. mori (30.44%) [4,11], H. armigera (35.30%) [4,29], M. sexta (31.99%) [28], Tribolium castaneum (35.51%) [4], Anoplophora glabripennis (33.10%) [44], Xylotrechus quadripes (35.03%) [42], and Rhaphuma horsfieldi (34.90%) [43]. Nevertheless, some clustered UGTs possibly derived from gene duplications retained high conservation (>50% identities) as presented in the tree, typically regarding the UGT33 and UGT40 subfamilies as discussed below.
In comparison to phase I metabolizing enzyme genes such as P450s and COEs, moth UGTs harbor a relatively stable gene number, ranging from 42 in Helicoverpa zea, 44 in M. sexta, 45 in B. mori, and 46 in H. armigera to 48 in S. frugiperda [4,11,28,29,45]. Our current study identified a comparable UGT number through the transcriptome analyses (50 relatives), very close to those of the above species with available genomes, and slightly more than 41 conreads in a zygaenid moth Zygaena filipendulae derived from the sequenced transcriptome [46]. In phylogeny, A. yunnanensis UGTs were expanded mainly in the UGT33 and UGT40 subfamilies, accounting for approximately half of all identified genes. Such large-scale expansions of UGTs in the two subfamilies were also reported in S. frugiperda [45], H. armigera [29], B. mori [4], and Epiphyas postvittana [47]. Notably, most members of the UGT33 and UGT40 subfamilies in Plutella xylostella [48] and Spodoptera exigua [19] have been found to have inducible expression by multi-insecticides. Hence, it is reasonably inferred that the lineage-specific expansions in UGT33 and UGT40 of A. yunnanensis may be associated with insecticide resistance, as an adaptation of insects to man-made chemical pesticides.
Concerning these UGT clusters in UGT33, we detected their physical locations in the chromosomes. The tandem arrays of UGTs on scaffolds or chromosomes were probably attributed to gene duplication events, coupled with their high identities. The clusters and high conservation of UGTs were also found in UGT33 and UGT40 of A. yunnanensis, reflecting a common phenomenon that species-specific expansions have occurred in moths. Of the 14 proposed clades, three orthologous groups (UGT44, UGT47, and UGT50) still retained a strict single gene copy in each species, suggesting the genes in each group may bear similar functional roles in moths, together with conserved catalytic and sugar-donor binding residues. Notably, the singleton in UGT50 was also presented in other insects [4,42]. Moreover, insect UGT50 members generally possessed conserved gene structure, for example, UGT50s in B. mori, M. sexta, and H. armigera comprised four introns as characterized in the present study, in agreeable with those of putative orthologs in some coleopteran species and human [42].
In Z. filipendulae, ZfilUGT33A1 could catalyze the synthesis of two cyanogenic glucosides by glycosylation, i.e., linamarin and lotaustralin secreted by Zygaena larvae [49]. Intriguingly, the two toxic cyanogenic glucosides were also isolated and identified from the A. yunnanensis larvae [50]. To identify putative UGTs in A. yunnanensis in response to the two chemicals, we screened putative ortholog(s) of ZfilUGT33A1 by a combination of the identities and phylogeny. As a result, AyunUGT14 and UGT22 showed the highest identities of 62.94% and 63.13% with ZfilUGT33A1, respectively. Further, the three UGT members phylogenetically formed an independent small clade with a high support value. Hence, it is suggested that AyunUGT14 and UGT22 belonging to the UGT33 subfamily probably play a critical role in the glycosylation of the two cyanogenic glucosides.
At the larval stage, detoxification-related tissues, referring to midguts, fat bodies, and Malpighian tubules, are of utmost importance for larvae to metabolize various harmful chemicals. In the expression profiling analyses, a number of AyunUGTs were transcribed in the three tissues. In other lepidopteran species, UGTs enriched in larval midguts, far bodies, or Malpighian tubules were common, and moreover, responded to plant defensive compounds and insecticides [4,11,12,15,19,48]. Given the fact that the larvae of A. yunnanensis feed on the plants of the Rosaceae family [20,21,22], AyunUGTs presented in larval detoxification tissues (especially AyunUGT4/8/15/27/31) are key molecular targets for detoxifying defensive compounds of host plants. Beyond putative roles of AyunUGTs in detoxification, it was found that a comparable number of genes had the expression in main chemosensory tissues, including antennae and maxillary palps. In moths, several studies have suggested olfactory roles of UGTs, such as M. sexta [9], B. mori [4,11], S. littoralis [8], A. lepigone [17], and P. xylostella [18]. In addition, this olfactory association was also indicated in non-lepidopteran species, such as D. melanogaster [51,52,53], Holotrichia parallela [54], X. quadripes [42], and R. horsfieldi [43]. Accordingly, it is inferred that the antenna-enriched AyunUGTs in A. yunnanensis (particularly AyunUGT9, UGT12, and UGT18) may participate in the sensing of odorant molecules, and are regarded as odorant degrading enzymes. Notably, the majority of AyunUGTs had detectable expression in reproductive tissues, suggesting their reproductive roles. In S. litura, transcriptome and proteome analyses revealed the presence of UGTs in reproductive tissues [37]. A similar reproductive expression feature of UGTs was also found in B. mori [4,11], H. armigera [29], and D. melanogaster [4,55], but their functions remain to be identified.

5. Conclusions

Our current study has characterized the UGT gene repertoire in A. yunnanensis, with a combination of transcriptomics, bioinformatics, phylogenetics, and PCR approaches. From the transcriptomes, a total of 50 UGTs were identified, of which some shared high conservation and phylogenetically clustered together. At the key sites, most of AyunUGTs harbored conserved residues with the UGTs from other insects and human. In the expression profiling analyses, a number of UGTs were found to have the expression in chemosensory tissues as supported by PCR and RNA–Seq (FPKM > 1), such as 24 and 22 relatives separately in larval antennae and maxillary palps, 27 in male antennae, 24 in female antennae, 31 in male legs, and 25 in female legs. Moreover, a comparable number of genes were observed in detoxification tissues, including 26, 22, and 19 relatives in larval midguts, fat bodies, and Malpighian tubules, respectively. The expression of at least 27 UGTs was detected in reproductive tissues of female and male moths. Together, this study reports, for the first time, the detoxification-related UGT gene family, and allows for exploring the adaptation of the A. yunnanensis larvae to feeding hosts.

Supplementary Materials

The following supporting information can be downloaded at: https://www.mdpi.com/article/10.3390/d14050407/s1. Table S1: Primers used for the expression profiling analyses of UGT genes in A. yunnanensis; Additional File S1: Amino acid sequences of UGTs in A. yunnanensis.

Author Contributions

Conceptualization, N.-Y.L.; methodology, H.-Y.X.; validation, H.-Y.X. and D.-L.C.; investigation, H.-Y.X., D.-L.C., T.-T.L. and Y.-J.Y.; resources, H.-Y.X.; data curation, H.-Y.X. and N.-Y.L.; writing—original draft preparation, H.-Y.X. and N.-Y.L.; writing—review and editing, H.-Y.X. and N.-Y.L.; supervision, N.-Y.L.; project administration, N.-Y.L.; and funding acquisition, N.-Y.L. All authors have read and agreed to the published version of the manuscript.

Funding

This work was supported by the Yunnan Fundamental Research Project (202001AT070100), the National Natural Science Foundation of China (31960115), and the Yunnan Provincial Support Plan for the Cultivation of High-level Talents Young Top-notch Talents (YNWR_ QNBJ_2019_057).

Institutional Review Board Statement

Not applicable.

Informed Consent Statement

Not applicable.

Data Availability Statement

Not applicable.

Conflicts of Interest

The authors declare no conflict of interest.

References

  1. Heidel-Fischer, H.M.; Vogel, H. Molecular mechanisms of insect adaptation to plant secondary compounds. Curr. Opin. Insect Sci. 2015, 8, 8–14. [Google Scholar] [CrossRef]
  2. Despres, L.; David, J.P.; Gallet, C. The evolutionary ecology of insect resistance to plant chemicals. Trends Ecol. Evol. 2007, 22, 298–307. [Google Scholar] [CrossRef] [PubMed]
  3. Ahn, S.-J.; Chertemps, T.; Maïbèche, M.; Marygold, S.J.; Van Leeuwen, T. Editorial: Invertebrate UDP-Glycosyltransferases: Nomenclature, Diversity and Functions. Front. Physiol. 2021, 12, 748290. [Google Scholar] [CrossRef] [PubMed]
  4. Ahn, S.J.; Vogel, H.; Heckel, D.G. Comparative analysis of the UDP-glycosyltransferase multigene family in insects. Insect Biochem. Mol. Biol. 2012, 42, 133–147. [Google Scholar] [CrossRef] [PubMed]
  5. Nagare, M.; Ayachit, M.; Agnihotri, A.; Schwab, W.; Joshi, R. Glycosyltransferases: The multifaceted enzymatic regulator in insects. Insect Mol. Biol. 2021, 30, 123–137. [Google Scholar] [CrossRef]
  6. Heppner, J.B. Butterflies and Moths (Lepidoptera). In Encyclopedia of Entomology; Capinera, J.L., Ed.; Springer: Dordrecht, The Netherlands, 2008; pp. 626–672. [Google Scholar]
  7. Mitter, C.; Davis, D.R.; Cummings, M.P. Phylogeny and evolution of Lepidoptera. Annu. Rev. Entomol. 2017, 62, 265–283. [Google Scholar] [CrossRef]
  8. Bozzolan, F.; Siaussat, D.; Maria, A.; Durand, N.; Pottier, M.A.; Chertemps, T.; Maibeche-Coisne, M. Antennal uridine diphosphate (UDP)-glycosyltransferases in a pest insect: Diversity and putative function in odorant and xenobiotics clearance. Insect Mol. Biol. 2014, 23, 539–549. [Google Scholar] [CrossRef]
  9. Robertson, H.M.; Martos, R.; Sears, C.R.; Todres, E.Z.; Walden, K.K.; Nardi, J.B. Diversity of odourant binding proteins revealed by an expressed sequence tag project on male Manduca sexta moth antennae. Insect Mol. Biol. 1999, 8, 501–518. [Google Scholar] [CrossRef]
  10. Bock, K.W. Vertebrate UDP-glucuronosyltransferases: Functional and evolutionary aspects. Biochem. Pharmacol. 2003, 66, 691–696. [Google Scholar] [CrossRef]
  11. Huang, F.F.; Chai, C.L.; Zhang, Z.; Liu, Z.H.; Dai, F.Y.; Lu, C.; Xiang, Z.H. The UDP-glucosyltransferase multigene family in Bombyx mori. BMC Genom. 2008, 9, 563. [Google Scholar] [CrossRef] [Green Version]
  12. Luque, T.; Okano, K.; O’Reilly, D.R. Characterization of a novel silkworm (Bombyx mori) phenol UDP-glucosyltransferase. Eur. J. Biochem. 2002, 269, 819–825. [Google Scholar] [CrossRef] [PubMed]
  13. Daimon, T.; Hirayama, C.; Kanai, M.; Ruike, Y.; Meng, Y.; Kosegawa, E.; Nakamura, M.; Tsujimoto, G.; Katsuma, S.; Shimada, T. The silkworm Green b locus encodes a quercetin 5-O-glucosyltransferase that produces green cocoons with UV-shielding properties. Proc. Natl. Acad. Sci. USA 2010, 107, 11471–11476. [Google Scholar] [CrossRef] [PubMed] [Green Version]
  14. Krempl, C.; Sporer, T.; Reichelt, M.; Ahn, S.-J.; Heidel-Fischer, H.; Vogel, H.; Heckel, D.G.; Joußen, N. Potential detoxification of gossypol by UDP-glycosyltransferases in the two Heliothine moth species Helicoverpa armigera and Heliothis virescens. Insect Biochem. Mol. Biol. 2016, 71, 49–57. [Google Scholar] [CrossRef] [PubMed]
  15. Israni, B.; Wouters, F.C.; Luck, K.; Seibel, E.; Ahn, S.-J.; Paetz, C.; Reinert, M.; Vogel, H.; Erb, M.; Heckel, D.G.; et al. The fall armyworm Spodoptera frugiperda utilizes specific UDP-glycosyltransferases to inactivate maize defensive benzoxazinoids. Front. Physiol. 2020, 11, 604754. [Google Scholar] [CrossRef] [PubMed]
  16. Ahn, S.J.; Badenes-Perez, F.R.; Reichelt, M.; Svatos, A.; Schneider, B.; Gershenzon, J.; Heckel, D.G. Metabolic detoxification of capsaicin by UDP-glycosyltransferase in three Helicoverpa species. Arch. Insect. Biochem. Physiol. 2011, 78, 104–118. [Google Scholar] [CrossRef] [PubMed]
  17. Zhang, Y.-N.; Ma, J.-F.; Xu, L.; Dong, Z.-P.; Xu, J.-W.; Li, M.-Y.; Zhu, X.-Y. Identification and expression patterns of UDP-glycosyltransferase (UGT) genes from insect pest Athetis lepigone (Lepidoptera: Noctuidae). J. Asia-Pac. Entomol. 2017, 20, 253–259. [Google Scholar] [CrossRef]
  18. He, P.; Zhang, Y.-F.; Hong, D.-Y.; Wang, J.; Wang, X.-L.; Zuo, L.-H.; Tang, X.-F.; Xu, W.-M.; He, M. A reference gene set for sex pheromone biosynthesis and degradation genes from the diamondback moth, Plutella xylostella, based on genome and transcriptome digital gene expression analyses. BMC Genom. 2017, 18, 219. [Google Scholar] [CrossRef] [Green Version]
  19. Hu, B.; Zhang, S.H.; Ren, M.M.; Tian, X.R.; Wei, Q.; Mburu, D.K.; Su, J.Y. The expression of Spodoptera exigua P450 and UGT genes: Tissue specificity and response to insecticides. Insect Sci. 2019, 26, 199–216. [Google Scholar] [CrossRef] [Green Version]
  20. Li, G.-C.; Nuo, S.-M.; Wang, Z.-Q.; Yang, A.-J.; Liu, N.-Y. Identification and expression profiling of chemosensory membrane protein genes in Achelura yunnanensis (Lepidoptera: Zygaenidae). Comp. Biochem. Physiol. D Genom. Proteom. 2021, 40, 100876. [Google Scholar] [CrossRef]
  21. Nuo, S.M.; Yang, A.J.; Li, G.C.; Xiao, H.Y.; Liu, N.Y. Transcriptome analysis identifies candidate genes in the biosynthetic pathway of sex pheromones from a zygaenid moth, Achelura yunnanensis (Lepidoptera: Zygaenidae). PeerJ 2021, 9, e12641. [Google Scholar] [CrossRef]
  22. Li, G.C.; Zhao, Y.J.; Li, J.L.; Lu, G.Y.; Liu, N.Y. Ultrastructure of sensilla on the antennae, proboscis and tarsi of adult Achelura yunnanensis (Lepidoptera: Zygaenidae). Acta Entomol. Sin. 2020, 63, 1385–1398. [Google Scholar]
  23. Teufel, F.; Almagro Armenteros, J.J.; Johansen, A.R.; Gíslason, M.H.; Pihl, S.I.; Tsirigos, K.D.; Winther, O.; Brunak, S.; von Heijne, G.; Nielsen, H. SignalP 6.0 predicts all five types of signal peptides using protein language models. Nat. Biotechnol. 2022. [Google Scholar] [CrossRef] [PubMed]
  24. Katoh, K.; Standley, D.M. MAFFT multiple sequence alignment software version 7: Improvements in performance and usability. Mol. Biol. Evol. 2013, 30, 772–780. [Google Scholar] [CrossRef] [PubMed] [Green Version]
  25. Waterhouse, A.M.; Procter, J.B.; Martin, D.M.A.; Clamp, M.; Barton, G.J. Jalview Version 2—A multiple sequence alignment editor and analysis workbench. Bioinformatics 2009, 25, 1189–1191. [Google Scholar] [CrossRef] [Green Version]
  26. Price, M.N.; Dehal, P.S.; Arkin, A.P. FastTree 2—Approximately maximum-likelihood trees for large alignments. PLoS ONE 2010, 5, e9490. [Google Scholar] [CrossRef] [PubMed]
  27. Xia, Q.; Zhou, Z.; Lu, C.; Cheng, D.; Dai, F.; Li, B.; Zhao, P.; Zha, X.; Cheng, T.; Chai, C.; et al. A draft sequence for the genome of the domesticated silkworm Bombyx mori. Science 2004, 306, 1937. [Google Scholar]
  28. Kanost, M.R.; Arrese, E.L.; Cao, X.; Chen, Y.R.; Chellapilla, S.; Goldsmith, M.R.; Grosse-Wilde, E.; Heckel, D.G.; Herndon, N.; Jiang, H.; et al. Multifaceted biological insights from a draft genome sequence of the tobacco hornworm moth, Manduca sexta. Insect Biochem. Mol. Biol. 2016, 76, 118–147. [Google Scholar] [CrossRef]
  29. Pearce, S.L.; Clarke, D.F.; East, P.D.; Elfekih, S.; Gordon, K.H.J.; Jermiin, L.S.; McGaughran, A.; Oakeshott, J.G.; Papanikolaou, A.; Perera, O.P.; et al. Genomic innovations, transcriptional plasticity and gene loss underlying the evolution and divergence of two highly polyphagous and invasive Helicoverpa pest species. BMC Biol. 2017, 15, 63. [Google Scholar] [CrossRef] [Green Version]
  30. Langmead, B.; Salzberg, S.L. Fast gapped-read alignment with Bowtie 2. Nat. Methods 2012, 9, 357–359. [Google Scholar] [CrossRef] [Green Version]
  31. Li, B.; Dewey, C.N. RSEM: Accurate transcript quantification from RNA-Seq data with or without a reference genome. BMC Bioinform. 2011, 12, 323. [Google Scholar] [CrossRef] [Green Version]
  32. Trapnell, C.; Williams, B.A.; Pertea, G.; Mortazavi, A.; Kwan, G.; van Baren, M.J.; Salzberg, S.L.; Wold, B.J.; Pachter, L. Transcript assembly and quantification by RNA-Seq reveals unannotated transcripts and isoform switching during cell differentiation. Nat. Biotechnol. 2010, 28, 511–515. [Google Scholar] [CrossRef] [PubMed] [Green Version]
  33. Simon, P. Q-Gene: Processing quantitative real-time RT-PCR data. Bioinformatics 2003, 19, 1439–1440. [Google Scholar] [CrossRef] [PubMed] [Green Version]
  34. Muller, P.Y.; Janovjak, H.; Miserez, A.R.; Dobbie, Z. Processing of gene expression data generated by quantitative real-time RT-PCR. BioTechniques 2002, 32, 1372–1374, 1376, 1378–1379. [Google Scholar] [PubMed]
  35. Miley, M.J.; Zielinska, A.K.; Keenan, J.E.; Bratton, S.M.; Radominska-Pandya, A.; Redinbo, M.R. Crystal structure of the cofactor-binding domain of the human phase II drug-metabolism enzyme UDP-glucuronosyltransferase 2B7. J. Mol. Biol. 2007, 369, 498–511. [Google Scholar] [CrossRef] [PubMed] [Green Version]
  36. Davidson, N.M.; Oshlack, A. Corset: Enabling differential gene expression analysis for de novo assembled transcriptomes. Genome Biol. 2014, 15, 410. [Google Scholar] [PubMed] [Green Version]
  37. Xiao, H.Y.; Li, G.C.; Wang, Z.Q.; Guo, Y.R.; Liu, N.Y. Combined transcriptomic, proteomic and genomic analysis identifies reproductive-related proteins and potential modulators of female behaviors in Spodoptera litura. Genom. 2021, 113, 1876–1894. [Google Scholar] [CrossRef] [PubMed]
  38. Swanson, W.J.; Clark, A.G.; Waldrip-Dail, H.M.; Wolfner, M.F.; Aquadro, C.F. Evolutionary EST analysis identifies rapidly evolving male reproductive proteins in Drosophila. Proc. Natl. Acad. Sci. USA 2001, 98, 7375–7379. [Google Scholar] [CrossRef] [Green Version]
  39. Wilburn, D.B.; Swanson, W.J. From molecules to mating: Rapid evolution and biochemical studies of reproductive proteins. J. Proteom. 2016, 135, 12–25. [Google Scholar] [CrossRef] [Green Version]
  40. Celorio-Mancera, M.d.l.P.; Ahn, S.-J.; Vogel, H.; Heckel, D.G. Transcriptional responses underlying the hormetic and detrimental effects of the plant secondary metabolite gossypol on the generalist herbivore Helicoverpa armigera. BMC Genom. 2011, 12, 575. [Google Scholar] [CrossRef] [Green Version]
  41. Li, X.; Zhu, B.; Gao, X.; Liang, P. Over-expression of UDP–glycosyltransferase gene UGT2B17 is involved in chlorantraniliprole resistance in Plutella xylostella (L.). Pest Manag. Sci. 2017, 73, 1402–1409. [Google Scholar] [CrossRef]
  42. Yin, N.-N.; Zhao, Y.-J.; Zhu, J.-Y.; Liu, N.-Y. Antennal UDP-glycosyltransferase genes in the coffee white stemborer, Xylotrechus quadripes. J. Asia-Pac. Entomol. 2019, 22, 1145–1153. [Google Scholar] [CrossRef]
  43. Wang, Z.Q.; Yin, N.N.; Zhao, N.; Liu, N.Y. Identification and expression characterization of UDP-glucosyltransferase genes in Rhaphuma horsfieldi. Chinese J. Appl. Entomol. 2020, 57, 898–910. [Google Scholar]
  44. McKenna, D.D.; Scully, E.D.; Pauchet, Y.; Hoover, K.; Kirsch, R.; Geib, S.M.; Mitchell, R.F.; Waterhouse, R.M.; Ahn, S.J.; Arsala, D.; et al. Genome of the Asian longhorned beetle (Anoplophora glabripennis), a globally significant invasive species, reveals key functional and evolutionary innovations at the beetle-plant interface. Genome Biol. 2016, 17, 227. [Google Scholar] [CrossRef] [Green Version]
  45. Gouin, A.; Bretaudeau, A.; Nam, K.; Gimenez, S.; Aury, J.-M.; Duvic, B.; Hilliou, F.; Durand, N.; Montagné, N.; Darboux, I.; et al. Two genomes of highly polyphagous lepidopteran pests (Spodoptera frugiperda, Noctuidae) with different host-plant ranges. Sci. Rep. 2017, 7, 11816. [Google Scholar] [CrossRef] [PubMed]
  46. Zagrobelny, M.; Scheibye-Alsing, K.; Jensen, N.B.; Møller, B.L.; Gorodkin, J.; Bak, S. 454 pyrosequencing based transcriptome analysis of Zygaena filipendulae with focus on genes involved in biosynthesis of cyanogenic glucosides. BMC Genom. 2009, 10, 574. [Google Scholar] [CrossRef] [Green Version]
  47. Thrimawithana, A.H.; Wu, C.; Christeller, J.T.; Simpson, R.M.; Hilario, E.; Tooman, L.K.; Begum, D.; Jordan, M.D.; Crowhurst, R.; Newcomb, R.D.; et al. The genomics and population genomics of the light brown apple moth, Epiphyas postvittana, an invasive tortricid pest of horticulture. Insects 2022, 13, 264. [Google Scholar] [CrossRef] [PubMed]
  48. Li, X.; Shi, H.; Gao, X.; Liang, P. Characterization of UDP-glucuronosyltransferase genes and their possible roles in multi-insecticide resistance in Plutella xylostella (L.). Pest Manag. Sci. 2018, 74, 695–704. [Google Scholar] [CrossRef]
  49. Jensen, N.B.; Zagrobelny, M.; Hjernø, K.; Olsen, C.E.; Houghton-Larsen, J.; Borch, J.; Møller, B.L.; Bak, S. Convergent evolution in biosynthesis of cyanogenic defence compounds in plants and insects. Nat. Commun. 2011, 2, 273. [Google Scholar] [CrossRef] [Green Version]
  50. Huang, S.Z.; Gong, W.C.; Ma, Q.Y.; Niu, Y.; Mei, W.L.; Chen, C.; Zhao, Y.X. Isolation and identification of cyanogenic glycosides from larval secretions of Achelura yunnanensis ( Lepidoptera: Zygaenidae) and the bioactivity against Tapinoma melanocephalum (Hymenoptera: Fromicidae). Acta Entomol. Sin. 2013, 56, 207–211. [Google Scholar]
  51. Wang, Q.; Hasan, G.; Pikielny, C.W. Preferential expression of biotransformation enzymes in the olfactory organs of Drosophila melanogaster, the antennae. J. Biol. Chem. 1999, 274, 10309–10315. [Google Scholar] [CrossRef] [Green Version]
  52. Younus, F.; Chertemps, T.; Pearce, S.L.; Pandey, G.; Bozzolan, F.; Coppin, C.W.; Russell, R.J.; Maibeche-Coisne, M.; Oakeshott, J.G. Identification of candidate odorant degrading gene/enzyme systems in the antennal transcriptome of Drosophila melanogaster. Insect Biochem. Mol. Biol. 2014, 53, 30–43. [Google Scholar] [CrossRef] [PubMed] [Green Version]
  53. Fraichard, S.; Legendre, A.; Lucas, P.; Chauvel, I.; Faure, P.; Neiers, F.; Artur, Y.; Briand, L.; Ferveur, J.-F.; Heydel, J.-M. Modulation of sex pheromone discrimination by a UDP-glycosyltransferase in Drosophila melanogaster. Genes 2020, 11, 237. [Google Scholar] [CrossRef] [PubMed] [Green Version]
  54. Wang, S.; Liu, Y.; Zhou, J.J.; Yi, J.K.; Pan, Y.; Wang, J.; Zhang, X.X.; Wang, J.X.; Yang, S.; Xi, J.H. Identification and tissue expression profiling of candidate UDP-glycosyltransferase genes expressed in Holotrichia parallela motschulsky antennae. Bull. Entomol. Res. 2018, 108, 807–816. [Google Scholar] [CrossRef] [PubMed]
  55. Ahn, S.-J.; Marygold, S.J. The UDP-glycosyltransferase family in Drosophila melanogaster: Nomenclature update, gene expression and phylogenetic analysis. Front. Physiol. 2021, 12, 648481. [Google Scholar] [CrossRef] [PubMed]
Diversity 14 00407 g001 550
Figure 1. Multiple alignment of amino acid sequences of 31 full-length UGTs in A. yunnanensis. The signal peptide, N–terminus, C–terminus, donor binding regions (DBR–1 and DBR–2), a transmembrane domain, and a cytoplasmic tail are marked on the top of the alignment. Triangles represent two catalytic residues. The signature motif is indicated by a magenta dotted box. Numbers 1, 2, and 3 represent amino acids interacting with nucleotides, phosphates and glucosides, respectively.
Figure 1. Multiple alignment of amino acid sequences of 31 full-length UGTs in A. yunnanensis. The signal peptide, N–terminus, C–terminus, donor binding regions (DBR–1 and DBR–2), a transmembrane domain, and a cytoplasmic tail are marked on the top of the alignment. Triangles represent two catalytic residues. The signature motif is indicated by a magenta dotted box. Numbers 1, 2, and 3 represent amino acids interacting with nucleotides, phosphates and glucosides, respectively.
Diversity 14 00407 g001
Diversity 14 00407 g002 550
Figure 2. Phylogenetic tree of UGTs in four moths. Based on the classification of UGT subfamilies in moth species, AyunUGTs clustered into 12 subfamilies. Newly identified AyunUGTs are shown in red. In the UGT33 subfamily, three typical UGT clusters in three species with available genomes are presented on scaffolds (Sca.) or chromosomes (Chr.). Arrows indicate the transcriptional orientation of genes. In M. sexta, six alternative splicing variants (UGT33C1–UGT33C6) sharing a common 3’-terminus are indicated by solid lines.
Figure 2. Phylogenetic tree of UGTs in four moths. Based on the classification of UGT subfamilies in moth species, AyunUGTs clustered into 12 subfamilies. Newly identified AyunUGTs are shown in red. In the UGT33 subfamily, three typical UGT clusters in three species with available genomes are presented on scaffolds (Sca.) or chromosomes (Chr.). Arrows indicate the transcriptional orientation of genes. In M. sexta, six alternative splicing variants (UGT33C1–UGT33C6) sharing a common 3’-terminus are indicated by solid lines.
Diversity 14 00407 g002
Diversity 14 00407 g003a 550Diversity 14 00407 g003b 550
Figure 3. Expression profile of 31 full-length UGT genes in different tissues of A. yunnanensis. (A) Expression of UGT genes as determined by FPKM values. (B) Expression of UGT genes as determined by RT–PCR. LA, larval antennae; LH, larval heads without antennae; LMP, larval maxillary palps; LMg, larval midguts; LFB, larval fat bodies; LMT, larval Malpighian tubules; FPu, female pupae; Mpu, male pupae; An, antennae; He, heads without antennae; Th, thoraxes; Ab, abdomens; Le, legs; Wi, wings; and PG, pheromone glands.
Figure 3. Expression profile of 31 full-length UGT genes in different tissues of A. yunnanensis. (A) Expression of UGT genes as determined by FPKM values. (B) Expression of UGT genes as determined by RT–PCR. LA, larval antennae; LH, larval heads without antennae; LMP, larval maxillary palps; LMg, larval midguts; LFB, larval fat bodies; LMT, larval Malpighian tubules; FPu, female pupae; Mpu, male pupae; An, antennae; He, heads without antennae; Th, thoraxes; Ab, abdomens; Le, legs; Wi, wings; and PG, pheromone glands.
Diversity 14 00407 g003aDiversity 14 00407 g003b
Diversity 14 00407 g004 550
Figure 4. qPCR analysis of six candidate UGT genes in A. yunnanensis. Data denote mean ± standard errors. Different lowercases above bars represent significant differences among tissues (p < 0.05, ANOVA, LSD). LA, larval antennae; LFB, larval fat bodies; LMP, larval maxillary palps; LMg, larval midguts; LMT, larval Malpighian tubules; MA, male antennae; ML, male legs; FA, female antennae; FL, female legs.
Figure 4. qPCR analysis of six candidate UGT genes in A. yunnanensis. Data denote mean ± standard errors. Different lowercases above bars represent significant differences among tissues (p < 0.05, ANOVA, LSD). LA, larval antennae; LFB, larval fat bodies; LMP, larval maxillary palps; LMg, larval midguts; LMT, larval Malpighian tubules; MA, male antennae; ML, male legs; FA, female antennae; FL, female legs.
Diversity 14 00407 g004
Diversity 14 00407 g005 550
Figure 5. Expression profile of 31 full-length UGT genes in reproductive tissues of A. yunnanensis. (A) Reproductive systems of female and male moths. (B) Expression of UGT genes in eight reproductive tissues. Asterisks mean that the transcription of the genes was not detected in tissues. MAG, male accessory glands; Te, testes; SV, seminal vesicles; ED, ejaculatory ducts; FAG, female accessory glands; BC, bursa copulatrix; Ov, ovaries; and SS, spermathecae with spermathecal glands.
Figure 5. Expression profile of 31 full-length UGT genes in reproductive tissues of A. yunnanensis. (A) Reproductive systems of female and male moths. (B) Expression of UGT genes in eight reproductive tissues. Asterisks mean that the transcription of the genes was not detected in tissues. MAG, male accessory glands; Te, testes; SV, seminal vesicles; ED, ejaculatory ducts; FAG, female accessory glands; BC, bursa copulatrix; Ov, ovaries; and SS, spermathecae with spermathecal glands.
Diversity 14 00407 g005
Table
Table 1. Information for 31 full-length UGT genes in A. yunnanensis.
Table 1. Information for 31 full-length UGT genes in A. yunnanensis.
GeneORF (bp)SP (aa)pIMw (kDa)NPS
UGT11560187.7359.90114, 273, 419
UGT21557177.3259.5363, 330
UGT31593178.8960.0363, 184, 247, 294, 343
UGT41575187.6560.2867, 427, 474
UGT51542188.7858.2749, 169, 318, 447, 501
UGT61602197.6861.3968, 138, 199, 308, 479, 523
UGT71575216.5261.2952, 244, 326, 452, 516
UGT81563209.1158.33237, 302
UGT91554209.1859.2982, 175, 245, 511
UGT101641218.9762.5752, 239, 333
UGT111545179.2259.2171, 89, 102, 237, 285
UGT121563209.0658.27103, 237, 302
UGT131557208.9559.75128
UGT141569227.7160.6971, 192, 289, 472
UGT151545188.9859.13120, 240
UGT161581206.3760.23147, 236
UGT171566208.9760.8851, 93, 240, 274, 426, 449
UGT181593218.8360.56252, 436
UGT191539198.8459.4160, 65, 102
UGT201563187.2959.64117, 251
UGT211512157.3457.0861, 72, 279, 315, 416, 417
UGT221575247.2560.8373, 291
UGT231584206.5560.8169, 429, 477
UGT241536218.9958.6074, 238, 289
UGT251554178.1059.9666, 147, 463
UGT261578227.5960.2471, 477
UGT271557176.6959.5166, 189, 474, 515
UGT281578209.1360.1082, 175, 245
UGT291623169.5061.2978, 119, 205, 231, 327, 406
UGT301539198.7659.3465
UGT311548188.1658.91120, 240, 415, 507
ORF, open reading frame; SP, signal peptide; aa, amino acid; pI, isoelectric point; Mw, molecular weight; and NPS, N-glycosylation predicted site.
Publisher’s Note: MDPI stays neutral with regard to jurisdictional claims in published maps and institutional affiliations.

Share and Cite

MDPI and ACS Style

Xiao, H.-Y.; Chen, D.-L.; Lu, T.-T.; Yao, Y.-J.; Liu, N.-Y. The UDP-Glycosyltransferase Gene Family in Achelura yunnanensis (Lepidoptera: Zygaenidae): Identification, Phylogeny, and Diverse Expression Patterns. Diversity 2022, 14, 407. https://doi.org/10.3390/d14050407

AMA Style

Xiao H-Y, Chen D-L, Lu T-T, Yao Y-J, Liu N-Y. The UDP-Glycosyltransferase Gene Family in Achelura yunnanensis (Lepidoptera: Zygaenidae): Identification, Phylogeny, and Diverse Expression Patterns. Diversity. 2022; 14(5):407. https://doi.org/10.3390/d14050407

Chicago/Turabian Style

Xiao, Hai-Yan, Dan-Lu Chen, Ting-Ting Lu, Yu-Juan Yao, and Nai-Yong Liu. 2022. "The UDP-Glycosyltransferase Gene Family in Achelura yunnanensis (Lepidoptera: Zygaenidae): Identification, Phylogeny, and Diverse Expression Patterns" Diversity 14, no. 5: 407. https://doi.org/10.3390/d14050407

APA Style

Xiao, H. -Y., Chen, D. -L., Lu, T. -T., Yao, Y. -J., & Liu, N. -Y. (2022). The UDP-Glycosyltransferase Gene Family in Achelura yunnanensis (Lepidoptera: Zygaenidae): Identification, Phylogeny, and Diverse Expression Patterns. Diversity, 14(5), 407. https://doi.org/10.3390/d14050407

Note that from the first issue of 2016, this journal uses article numbers instead of page numbers. See further details here.

Article Metrics

Back to TopTop