Published online : 26 May 2021 Article Outline
Data Release
Utilizing a chromosomal-length genome assembly to annotate the Wnt signaling pathway in the Asian citrus psyllid, Diaphorina citri
Chad Vosburg
Chad Vosburg
Indian River State College, Fort Pierce, FL 34981, USA
Department of Plant Pathology and Environmental Microbiology, Pennsylvania State University, University Park, PA 16802, USA
Department of Plant Pathology and Environmental Microbiology, Pennsylvania State University, University Park, PA 16802, USA
Find this author on Google Scholar
Find this author on PubMedMax Reynolds
Max Reynolds
Indian River State College, Fort Pierce, FL 34981, USA
Rita Noel
Rita Noel
Indian River State College, Fort Pierce, FL 34981, USA
Teresa Shippy
Teresa Shippy
KSU Bioinformatics Center, Division of Biology, Kansas State University, Manhattan, KS 66506, USA
Prashant S. Hosmani
Prashant S. Hosmani
Boyce Thompson Institute, Ithaca, NY 14853, USA
Mirella Flores-Gonzalez
Mirella Flores-Gonzalez
Boyce Thompson Institute, Ithaca, NY 14853, USA
Lukas A. Mueller
Lukas A. Mueller
Boyce Thompson Institute, Ithaca, NY 14853, USA
Wayne B. Hunter
Wayne B. Hunter
USDA-ARS, US Horticultural Research Laboratory, Fort Pierce, FL 34945, USA
Susan J. Brown
Susan J. Brown
KSU Bioinformatics Center, Division of Biology, Kansas State University, Manhattan, KS 66506, USA
Tom D’Elia
Tom D’Elia
Indian River State College, Fort Pierce, FL 34981, USA
Surya Saha
* Surya Saha
Boyce Thompson Institute, Ithaca, NY 14853, USA
Animal and Comparative Biomedical Sciences, University of Arizona, Tucson, AZ 85721, USA
Animal and Comparative Biomedical Sciences, University of Arizona, Tucson, AZ 85721, USA
Corresponding author. E-mail: suryasaha@cornell.edu
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Cite this article as...
Chad Vosburg, Max Reynolds, Rita Noel, Teresa Shippy, Prashant S. Hosmani, Mirella Flores-Gonzalez, Lukas A. Mueller, Wayne B. Hunter, Susan J. Brown, Tom D’Elia, Surya Saha, Utilizing a chromosomal-length genome assembly to annotate the Wnt signaling pathway in the Asian citrus psyllid,
Article Information
Journal title: Gigabyte
Publisher name: GigaScience Press
Publisher location: Sha Tin, New Territories, Hong Kong SAR
Article series: Asian citrus psyllid community annotation
Received: 18 December 2020
Accepted: 18 May 2021
Published: 26 May 2021
Preprint submitted to: https://doi.org/10.1101/2020.09.21.306100
Copyright © The Author(s) 2021.
http://creativecommons.org/licenses/by/4.0/
Data Description
Introduction
Diaphorina citri (NCBI:txid121845) is the insect vector of Huanglongbing (HLB), or citrus greening disease, which has devastated global citrus production [1, 2]. HLB management is heavily based on controlling the spread of D. citri. To better understand the insect’s biology, the D. citri genome has been manually annotated to curate accurate gene model predictions. Accurate gene models can be used to develop novel insect control systems that utilize molecular therapeutics such as CRISPR (clustered regularly interspaced short palindromic repeats) and RNA interference (RNAi) to control the spread of D. citri [3, 4]. These molecular therapeutics would be gene-specific, thus would reduce reliance on broad-spectrum insecticides that have given rise to resistant D. citri populations [5–7].
Context
Here, we report D. citri genes involved in both canonical and noncanonical Wnt signaling. Wnt signaling is important for many biological processes in metazoans, such as patterning, cell polarity, tissue generation, and stem cell maintenance [8–10]. In the model insects Drosophila melanogaster and Tribolium castaneum, knockout and knockdown of Wnt ligands and other Wnt signaling components have detrimental effects on embryo development and adult homeostasis [11–16]. Wnt signaling components could therefore be effective knockout targets to limit the spread of D. citri, thus reducing HLB incidence. We curated a comprehensive repertoire of Wnt signaling genes in D. citri. Twenty-four gene models corresponding to canonical and noncanonical Wnt signaling genes have been annotated, including seven Wnt ligands, three frizzled homologs, arrow, armadillo/beta-catenin, and receptor tyrosine kinases ROR and doughnut. We were unable to find Wnt8/D, Wnt9, and Wnt16 as well as Wnt2-4, which have been lost in insects. The mechanisms of Wnt signaling appear to be mostly conserved and comparable to those found in D. melanogaster (Table 1). A model for canonical Wnt signaling in D. citri based on curated genes is shown (Figure 1). This is an important first step towards understanding critical biological processes that might be targeted to control the spread of D. citri, and may provide broader insights into the mechanisms of Wnt signaling in this important hemipteran vector.
Methods
Figure 1.
Theoretical model of canonical Wnt signaling cascade in D. citri based on curated genes. (1) Wnt is secreted. (2) Wnt concentration gradient forms. (3) Wnt binds to Frizzled and releases Armadillo. (4) Armadillo migrates into the nucleus, associates with transcription factor Pangolin, and regulates gene expression. (5) Armadillo is degraded in the absence of Wnt.
Table 1
Summary of gene copy numbers in various model insect species, including Diaphorina citri. Wnt pathway ortholog numbers in five different insect species. Drosophila melanogaster, Apis mellifera, Tribolium castaneum, and Acyrthosiphon pisum copy numbers were determined using Flybase, OrthoDB, NCBI Genbank, Uniprot, and several other publications [15, 20–22]. Diaphorina citri numbers represent the number of manually annotated genes in the D. citri v3.0 genome.
| Gene | Drosophila melanogaster | Apis mellifera | Tribolium castaneum | Acyrthosiphon pisum | Diaphorina citri v3 |
|---|---|---|---|---|---|
| Wnt1 | 1 | 1 | 1 | 1 | 1 |
| Wnt5 | 1 | 1 | 1 | 1 | 1 |
| Wnt6 | 1 | 1 | 1 | 0 | 1 |
| Wnt7 | 1 | 1 | 1 | 1 | 1 |
| Wnt8/D | 1 | 0 | 1 | 0 | 0 |
| Wnt9 | 1 | 0 | 1 | 0 | 0 |
| Wnt10 | 1 | 1 | 1 | 0 | 1 |
| Wnt11 | 0 | 1 | 1 | 1 | 1 |
| Wnt16 | 0 | 0 | 0 | 1 | 0 |
| WntA | 0 | 1 | 1 | 1 | 1 |
| Pangolin | 1 | 1 | 1 | 1 | 1 |
| Armadillo | 1 | 1 | 2 | 2 | 1 |
| Wntless | 1 | 1 | 1 | 1 | 1 |
| Porcupine | 1 | 1 | 1 | 1 | 1 |
| Derailed | 2 | 1 | 0 | 1 | 1 |
| Doughnut | 1 | 1 | 1 | 1 | 1 |
| Arrow | 1 | 1 | 1 | 1 | 1 |
| Frizzled | 4 | 2 | 3 | 2 | 3 |
| ROR | 2 | 2 | 3 | 2 | 2 |
| Dishevelled | 1 | 1 | 1 | 1 | 1 |
| Shaggy | 1 | 1 | 1 | 2 | 1 |
| Axin | 1 | 1 | 1 | 1 | 1 |
| ck1-gamma | 1 | 1 | 1 | 1 | 1 |
| Apc | 2 | 1 | 1 | 1 | 1 |
Figure 2.
Protocol for psyllid genome curation [17]. https://www.protocols.io/widgets/doi?uri=dx.doi.org/10.17504/protocols.io.bniimcce
To summarize, orthologous protein sequences for Wnt pathway genes were collected from the National Center for Biotechnology Information (NCBI) (RRID:SCR_006472) protein database [18] and used to BLAST (RRID:SCR_004870) search the D. citri MCOT transcriptome database [19]. The MCOT transcriptome is a transcriptome assembly utilizing Maker (RRID:SCR_005309), Cufflinks (RRID:SCR_014597), Oases (RRID:SCR_011896), and Trinity (RRID:SCR_013048) pipelines to provide a comprehensive set of predicted gene models. High-scoring MCOT models (accessions available in Table 2) were then searched on the NCBI protein database using NCBI BLAST to confirm the viability of the predicted MCOT models. The high-scoring MCOT models that had promising NCBI search results were used to search the D. citri genome. Genome regions containing computationally predicted gene models with high sequence identity to the query sequence from the MCOT transcriptome were investigated within JBrowse (RRID:SCR_001004). Gene models were modified using the Apollo (RRID:SCR_001936) gene annotation platform, based on mapped DNA-Seq, RNA-Seq, Iso-Seq, orthologous proteins, and other lines of evidence to edit and confirm manual annotations and gene structure. The gene models were analyzed with NCBI BLAST to assess their completeness. MUSCLE (RRID:SCR_011812) multiple sequence alignments of the D. citri gene model sequences and orthologous sequences were created through MEGA7 (RRID:SCR_000667) [23]. Neighbor-joining trees were constructed using MEGA7 with p-distance for determining branch length and 1000 bootstrapping replications to measure the precision of branch placement. In special cases, phylogenetic analysis in conjunction with NCBI BLAST scores was used to properly name and characterize the manually annotated gene models.
RNA-seq data from whole body adults and nymphs raised on C. medica and C. sinensis are available from NCBI BioProject PRJNA609978. We used proteins from Drosophila melanogaster (Dm) [24], Tribolium castaneum [25], Bombyx mori [26], Apis mellifera [27], Nasonia vitripennis
[28], Acyrthosiphon pisum [29], Nilaparvata lugens
[30, 31], Sipha flava [32], Halyomorpha halys [33], Cimex lectularius [34], Aedes aegypti [35], Anopheles gambiae [36], Branchiostoma floridae [37], Penaeus vannamei
[38], Folsomia candida [39], Spodoptera litura [40], Homo sapiens [32] and Oncopeltus fasciatus [34, 41].
Figure 3.
Neighbor-joining tree of Wnt protein sequences. Phylogenetic analysis was performed to categorize the seven D. citri Wnt genes (signified by dots). Wnt families are distinguished by clades and are color coded. Bootstrap values are based on 1000 replicates and values under 25 are removed. Ortholog sequences were collected from NCBI protein database [18]. Analysis was performed using MEGA7 [23].
Figure 4.
Wnt genes in six insects. A colored box indicates the presence of a Wnt subfamily (1–11, 16, and A) in that insect, while a white box indicates the loss of a subfamily. For example, all six species have Wnt1 and Wnt5, none have Wnt2-4, and only A. pisum has Wnt16. Homologs of Wnt8 in T. castaneum and D. melanogaster are also referred to as WntD.
Figure 5.
Wnt1-6-10 cluster comparison. Organization of the Wnt1-6-10 cluster in D. citri is similar to that of D. melanogaster and differs from what may be a basal arthropod gene arrangement seen in A. gambiae, T. castaneum, A. mellifera, and D. pulex. Gene lengths are not to scale.
Data validation and quality control
The loss of Wnt ligand genes is more common in insects than in other metazoans [20], which leads to a highly variable array of Wnt genes and Wnt signaling components from species to species [15, 21, 22, 42]. We performed a phylogenetic analysis to characterize the D. citri Wnt repertoire (Figure 3). The ortholog sequences used for this analysis were collected from the NCBI protein database [18]; see the ‘Availability of Supporting Data and Materials’ section for accession numbers. Seven D. citri Wnts were identified and classified as Wnt1 (also known as wingless), Wnt5, Wnt6, Wnt7, Wnt10, Wnt11, and WntA (Figures 3 and 4). In comparison, seven Wnt genes have been identified in D. melanogaster, nine in T. castaneum, and six in Acyrthosiphon pisum
[22, 42]. The collection of Wnt genes found in D. citri is like that found in other insects, and no Wnt subfamilies have been identified as being unique to D. citri. Contrary to previous reports [43], D. citri does appear to possess a Wnt6 gene.
Wnt1, Wnt6, and Wnt10 typically occur close together in a highly conserved gene cluster [44, 45]. The chromosomal length genome assembly in v3.0 suggests that this cluster is also conserved in D. citri, located at a position between 26.4 Mb (megabases) and 26.6 Mb on scaffold 4 (i.e. chromosome 4) [46]. In comparison, the Wnt1-6-10 cluster is located at a position between 7.30 Mb and 7.38 Mb on chromosome 2L of D. melanogaster, and between 5.50 Mb and 5.53 Mb on linkage group 5 in T. castaneum. The only gene from this cluster present in A. pisum is Wnt1, which is located on the X chromosome. The close phylogenetic relationship of Wnt1, Wnt6, and Wnt10 in D. citri (Figure 3) supports the hypothesis that this cluster is the result of an ancient duplication event, one that may predate the divergence of cnidarians and bilaterians [45]. The orientation of these clustered D. citri Wnt genes is like that found in D. melanogaster and differs from what may be a basal arthropodal organization of Wnts found in species of Coleoptera, Hymenoptera, and Cladocera (Figure 5). Wnt9 is also associated with this gene cluster when present in the genome. However, as with A. pisum, Wnt9 was not found in the D. citri genome and appears to have been lost during evolution. A second Wnt cluster, Wnt5 and Wnt7, is also common among non-insect metazoans. This cluster is not seen in D. citri; however, D. citri Wnt5 and 7 are located relatively close to one another (within 220 Kb [kilobase pairs]) on scaffold 13 (i.e. chromosome 13).
The mechanisms that act to conserve these Wnt gene clusters are not well understood. In the basal metazoan Nematostella vectensis, clustered Wnt genes do not exhibit similar expression patterns or Hox-like collinearity [44], and may not share regulatory elements. Whole body transcript expression data from egg, nymph, and adult stages [47] obtained from the Citrus Greening Expression Network (CGEN) [48] shows varying levels of expression among the clustered genes in different life stages of D. citri (Figure 6). However, it appears that Wnt1 and 10 are similarly upregulated during embryonic psyllid development and downregulated during the adult stage. Similar transcript levels of Wnt1 and 6 are seen in the nymphal stage. This suggests there may be shared regulation dependent upon life stage. Furthermore, ordering within the clusters is subject to rearrangement (Figure 5) [42, 44]. This may indicate that gene directionality is not a factor in conserving this cluster. Our annotation findings support the hypothesis that the Wnt1-6-10 cluster is being preserved through either natural selection or an unknown mechanism. A better understanding of the regulatory hierarchy controlling Wnt expression might shed light on the significance of Wnt gene associations in the genome. Future characterization of the coding and noncoding regions surrounding these D. citri Wnts (e.g., tandem repeat analysis) could also provide more insight into the mechanisms causing Wnt duplication events.
Figure 6.
Transcript levels of clustered Wnt transcripts during different D. citri life stages. Whole body transcript expression data from egg, nymph, and adult stages [47] were collected from CGEN [48]. The psyllids were raised on Citrus macrophylla and were not infected with Candidatus Liberibacter asiaticus. Expression values shown in transcripts per million (TPM).
The organization of the genomic reference sequence into chromosomal length scaffolds was essential for revealing D. citri gene clustering. Because of their shorter scaffold lengths, previous genome assemblies were often unsupportive in confirming the proximity of genes. Genome v2.0 assembly errors had likely misrepresented the location of Wnt10, making it appear to be separated from Wnt1 and Wnt6. A complete Wnt1-6-10 cluster was found in the improved chromosome length assembly v3.0. Thus, the quality of the reference genome should be considered when performing phylogenetic studies.
Orthologs for Wnt2, Wnt3, Wnt4, Wnt8/D, Wnt9, and Wnt16 were not located in the D. citri genome. The close identity of certain Wnt subfamilies makes it difficult to distinguish between them; however, the loss of Wnt2–4 is expected because they are absent in all insects [20]. Apis mellifera and the hemipteran A. pisum have been reported to lack Wnt8/D. Perhaps this Wnt subfamily has been lost in the divergence from other insect groups [22]. Additionally, Wnt16 was not found in D. citri v3.0. This finding contrasts with the gene predictions of other hemipteran genomes available at NCBI, namely A. pisum, Sipha flava, and Nilaparvata lugens (Figure 3). Based on whole body RNA expression data collected from CGEN, Wnt6 has the highest average transcript levels of all the Wnt genes in both nymph and adult psyllids (Figure 7). The relatively high number of transcripts suggests that Wnt6 is important during both metamorphosis and adult stage homeostasis, and may be a good knockout target for molecular therapeutics. Transcript expression of Wnt6 in adults is mainly concentrated in the legs and thorax, averaging 102 transcripts per million (TPM) and 272 TPM, respectively. This is considerably higher than all other Wnt genes in these tissues, which only average between 0.26 and 3.00 TPM. It is unclear if other Wnts can be upregulated to compensate for the loss of Wnt6. Perhaps targeting multiple Wnt genes, or the mechanisms by which Wnt is secreted (i.e. Porcupine and Wntless), would be more disruptive to D. citri physiology.
Figure 7.
Transcript levels of D. citri Wnt repertoire in both nymph and adult psyllids from whole body RNA extractions. Green bars indicate the average transcript levels for Wnt in nymph samples [47], and gray bars represent the average transcript levels for Wnt in adult samples. Averages are based on six nymph samples and six adult samples. Expression levels shown in transcripts per million (TPM). Standard deviation of samples is shown by error bars. RNA-seq data was collected from CGEN [48].
Table 2
Evidence supporting gene annotation. Manually annotated Wnt pathway genes in Diaphorina citri. There are 24 gene models in total. Each gene model has been assigned an identifier, and the evidence used to validate or modify the structure of the gene model has been listed. MCOT transcriptome identifiers that best support the manual annotation are also listed. The table is marked with an ‘X’ when supporting evidence of de novo transcriptome, Iso-Seq, RNA-Seq and ortholog support is present. MCOT: comprehensive transcriptome based on genome MAKER, Cufflinks, Oases, and Trinity transcript predictions; MAKER: gene predictions; De novo transcriptome: an independent transcriptome using Iso-Seq long-reads and RNA-Seq data; Iso-Seq transcripts: full-length transcripts generated with Pacific Biosciences technology; RNA-Seq: reads mapped to genome are also used as supporting evidence for splice junctions; Ortholog evidence: proteins from related hemipteran species and Drosophila melanogaster.
| Gene | OGS Identifier | MCOT | de novo transcriptome | Iso-Seq | RNA-Seq | Ortholog |
|---|---|---|---|---|---|---|
| Wnt1 | Dcitr04g11660.1.1 | MCOT05703.0.CO | X | X | X | |
| Wnt5 | Dcitr13g03650.1.1 | MCOT16538.0.CO | X | X | ||
| Wnt6 | Dcitr04g11650.1.1 | MCOT21516.1.CO | X | X | X | |
| Wnt7 † | Dcitr13g03730.1.1 | MCOT12704.0.CO | X | X | X | |
| Wnt10 | Dcitr04g11640.1.1 | MCOT09136.0.MO | X | X | X | |
| Wnt11 | Dcitr09g05250.1.1 | MCOT15024.0.CT | X | X | ||
| WntA | Dcitr13g02920.1.1 | MCOT02236.1.CT | X | X | X | |
| Pangolin † | Dcitr06g15680.1.1 | MCOT15454.2.CC | X | X | ||
| Armadillo | Dcitr10g09220.1.1 | MCOT18153.0.CT | X | X | X | |
| Wntless | Dcitr01g07340.1.1 | MCOT02320.0.CC | X | X | X | X |
| Porcupine | Dcitr13g04750.1.1 | MCOT19771.0.CO | X | X | X | |
| Derailed | Dcitr01g12220.1.1 | MCOT04433.1.CO | X | X | X | |
| Doughnut | Dcitr01g07650.1.1 | MCOT18207.0.CT | X | X | X | X |
| Arrow | Dcitr11g02670.1.1 | MCOT01906.1.CO | X | X | X | X |
| Frizzled | Dcitr04g04630.1.1 | MCOT11925.0.MO | X | X | ||
| Frizzled 2 | Dcitr10g03570.1.1 | MCOT07682.0.MO | X | X | X | |
| Frizzled 3 | Dcitr01g12100.1.1 | MCOT03353.0.CC | X | X | ||
| ROR1 | Dcitr05g14430.1.1 | MCOT18375.0.CT | X | X | X | X |
| Dcitr05g14430.1.2 | MCOT01992.1.CT | |||||
| ROR2 | Dcitr08g10450.1.1 | MCOT22482.0.CC | X | X | X | X |
| Dishevelled | Dcitr01g03830.1.1 | MCOT11762.0.MO | X | X | X | |
| Shaggy | Dcitr03g15060.1.1 | MCOT06728.0.CT | X | X | X | X |
| Axin | Dcitr07g09620.1.1 | MCOT05716.1.CT | X | X | ||
| ck1-gamma | Dcitr11g04200.1.1 | MCOT05782.2.CO | X | X | X | X |
| Apc-like | Dcitr07g12790.1.1 | MCOT14853.2.CO | X | X |
†Gene is manually annotated as a partial model in Genome v3.0. A complete representation of the gene and protein sequence can be determined with MCOT transcriptome data.
Several receptors and co-receptors associated with canonical and noncanonical signaling have been identified (Table 2). Three paralogs for the Wnt receptor encoding frizzled have been found in D. citri. We classified and numerically designated D. citri’s three frizzled genes based on how their encoded protein sequences form clades with D. melanogaster orthologs (Figure 8). Our analysis showed that D. citri, and other hemipterans such as Halymorpha halys and N. lugens, possess a Frizzled protein like that of D. melanogaster’s Frizzled 3. Some hemipteran Frizzled orthologs form a distinct clade separate from the Dipteran sequences (Figure 8). The hemipteran clade suggests that these genes might belong to a different subfamily of Frizzled, maybe one specific to Hemiptera. However, this ortholog has not been reported in the A. pisum genome [22].
Orthologs for both ROR1 and ROR2 have been identified. Interestingly, ROR1 has two isoforms, the first of which contains an immunoglobulin (IG) domain that is lacking from isoform 2 (Figure 9). ROR1 isoform 2 (Dcitr05g14430.1.2) appears to average higher transcript levels in D. citri egg, nymph, and adult tissues than ROR1 isoform 1 (Dcitr05g14430.1.1) based on CGEN data (Figure 10). Many transcripts for isoform 2 were detected in the psyllid egg (Figure 10). This suggests that expression of isoform 2 may be important in the early developmental stages of D. citri.
Figure 8.
Neighbor-joining tree of insect Frizzled protein sequences. Proteins grouped in the Frizzled 1 subfamily are highlighted in green, Frizzled 2 in orange, Frizzled 3 in blue, and Frizzled 4 in magenta. Circles indicate the D. citri sequences. Ortholog sequences were collected from the NCBI protein database [18]. Some NCBI sequences (such as XP_006568530.1, XP_008188372.2, and XP_022194032.1) may have numeric labels derived from computational predictions that do not reflect sequence or functional similarity. Analysis was performed using MEGA7 [23].
Figure 9.
Domain comparison of ROR1 isoforms. The immunoglobulin domain (IG_like) is present in isoform 1. Other shared domains include a cysteine-rich frizzled domain (CRD_FZ), a Kringle domain (KR), and a protein kinase catalytic domain (PKc_like). Domains were calculated and visualized using the NCBI Conserved Domain Architecture Retrieval Tool (CDART).
Figure 10.
Expression of ROR1 isoforms in egg, nymph and adult D. citri. Pink bars indicate the average transcript levels for isoform 1 (Dcitr05g14430.1.1), and orange bars indicate the average transcript levels for isoform 2 (Dcitr05g14430.1.2). Note: only one egg sample was used for comparison. Egg transcripts from the whole egg (one sample), nymph transcripts from the whole body (six samples), and adult transcripts from the whole body, abdomen, and thorax (14 samples) are shown. Expression values shown in transcripts per million (TPM). Data labels note the average TPM. Standard deviation of samples, when available, is shown by error bars. RNA-seq data was collected from CGEN [48].
Conclusion
Controlling the spread of D. citri is an important strategy for reducing the spread of HLB. With this study, we hope to provide a greater insight into D. citri biology, as well as accurate gene models that can be used in future research and applications. We have curated a comprehensive repertoire of Wnt signaling genes in D. citri. In total, 24 gene models corresponding to canonical and noncanonical Wnt signaling have been annotated. The mechanisms of Wnt signaling appear to be mostly conserved and comparable to those found in D. melanogaster and other insects. These findings provide a greater insight into the evolutionary history of D. citri and Wnt signaling in this important hemipteran vector. Manual annotation and an improved genome assembly with chromosomal length scaffold were essential for identifying high quality gene models.
Reuse potential
The manually curated genes will be included in the Citrus Greening Expression Network (CGEN) [48] as a part of the Official Gene Set version 3. This visualization tool is useful for understanding psyllid biology and comparative analysis because it contains public transcriptomics data for Diaphorina citri from various tissues, life stages, CLas infection levels and citrus hosts. Future work could utilize these gene models in developing CRISPR and RNAi systems that target and disrupt critical biological processes in D. citri, thus controlling the spread of HLB. This work was done as part of a collaborative community annotation project [49].
Data availability
Our annotation and gene curation workflow is described by Shippy et al.
[17]. The Diaphorina citri genome assembly, official gene sets, and transcriptome data are accessible on the Citrus Greening website [50]. All accessions for genes used for phylogentic analysis are provided within this report (Tables 2, 3, Figure 8). We have included the Newick and Multiple Sequence Alignment files used to construct the Wnt neighbor-joining phylogenetic tree and other data is available in the GigaScience GigaDB repository [51].
Table 3
Accessions for Wnt phylogenetic tree.
| NCBI accession | Species | NCBI protein name | Referred to in Figure 3 as |
|---|---|---|---|
| XP_002609873.1 | Branchiostoma floridae | Hypothetical protein BRAFLDRAFT_60204 | WntA |
| XP_024085687.1 | Cimex lectularius | Wnt-8b-like | Wnt8 |
| XP_014257242.2 | Cimex lectularius | Wnt-7b isoform X1 | Wnt7 |
| NP_476972.2 | Drosophila melanogaster | Wnt oncogene analog 4 isoform A | Wnt9 |
| NP_476924.1 | Drosophila melanogaster | Wnt oncogene analog 5 isoform A | Wnt5 |
| NP_476810.1 | Drosophila melanogaster | Wnt oncogene analog 2 isoform A | Wnt7 |
| NP_609109.3 | Drosophila melanogaster | Wnt oncogene analog 10 | Wnt10 |
| NP_609108.3 | Drosophila melanogaster | Wnt oncogene analog 6 isoform B | Wnt6 |
| NP_523502.1 | Drosophila melanogaster | Wingless | Wnt1 |
| NP_650272.1 | Drosophila melanogaster | wnt inhibitor of dorsal | Wnt8/D |
| ALO81632.1 | Penaeus vannamei | Wnt-16 | Wnt16 |
| OXA45577.1 | Folsomia candida | Wnt-16 | Wnt16 |
| XP_025422997.1 | Sipha flava | Wnt-16-like | Wnt16 |
| XP_022821085.1 | Spodoptera litura | Wnt-4-like | Wnt9 |
| XP_015835609.1 | Tribolium castaneum | Wnt-4 | Wnt9 |
| XP_008196351.1 | Tribolium castaneum | Wnt-7b isoform X1 | Wnt7 |
| XP_008195370.1 | Tribolium castaneum | Wnt-1 | WntA |
| XP_015835988.1 | Tribolium castaneum | Wnt-11b-1 isoform X1 | Wnt11 |
| XP_008193179.1 | Tribolium castaneum | Wnt-10a isoform X1 | Wnt10 |
| NP_001164137.1 | Tribolium castaneum | Wnt6 protein precursor | Wnt6 |
| NP_001107822.1 | Tribolium castaneum | wingless precursor | Wnt1 |
| XP_974684.1 | Tribolium castaneum | Wnt-5b | Wnt5 |
| XP_971439.1 | Tribolium castaneum | Wnt-8a isoform X1 | Wnt8 |
| XP_021702998.1 | Aedes aegypti | Wnt-4 | WntA |
| XP_557821.3 | Anopheles gambiae | AGAP008678-PA | WntA |
| XP_006561993.1 | Apis mellifera | Wnt-5b isoform X1 | Wnt5 |
| XP_006557287.1 | Apis mellifera | Wnt-7b isoform X1 | Wnt7 |
| XP_006567803.2 | Apis mellifera | Wnt-11b | Wnt11 |
| XP_016771882.1 | Apis mellifera | Wnt-6 isoform X1 | Wnt6 |
| XP_026300091.1 | Apis mellifera | Wnt-1 | Wnt1 |
| XP_396944.4 | Apis mellifera | Wnt-10b | Wnt10 |
| XP_001949667.2 | Acyrthosiphon pisum | Wnt-5b | Wnt5 |
| XP_016664156.1 | Acyrthosiphon pisum | Wnt-16 | Wnt16 |
| XP_001948541.2 | Acyrthosiphon pisum | Wnt-2 | Wnt7 |
| XP_001947400.1 | Acyrthosiphon pisum | Wnt-1 | WntA |
| XP_001944637.3 | Acyrthosiphon pisum | Wnt-11b-like isoform X1 | Wnt11 |
| XP_001945295.1 | Acyrthosiphon pisum | Wnt-1 | Wnt1 |
| XP_022184533.1 | Nilaparyata lugens | Wnt-16-like | Wnt16 |
| XP_022188550.1 | Nilaparyata lugens | Wnt-7b | Wnt7 |
| BAB62039.1 | Homo sapiens | WNT5B | Wnt5B |
| NP_003382.1 | Homo sapiens | Wnt-2 precursor | Wnt2 |
| NP_057171.2 | Homo sapiens | Wnt-16 isoform 2 | Wnt16 |
| NP_004616.2 | Homo sapiens | Wnt-7a precursor | Wnt7a |
| NP_478679.1 | Homo sapiens | Wnt-7b precursor | Wnt7b |
| NP_004617.2 | Homo sapiens | Wnt-11 precursor | Wnt11 |
| NP_003386.1 | Homo sapiens | Wnt-9a precursor | Wnt9a |
| NP_003387.1 | Homo sapiens | Wnt-9b isoform 1 precursor | Wnt9b |
| NP_110388.2 | Homo sapiens | Wnt-4 precursor | Wnt4 |
| NP_079492.2 | Homo sapiens | Wnt-10a precursor | Wnt10a |
| NP_003385.2 | Homo sapiens | Wnt-10b precursor | Wnt10b |
| NP_006513.1 | Homo sapiens | Wnt-6 precursor | Wnt6 |
| NP_005421.1 | Homo sapiens | proto-oncogene Wnt-1 precursor | Wnt1 |
| NP_001287867.1 | Homo sapiens | Wnt-8a isoform 1 precursor | Wnt8 |
Editor’s note
This article is one of a series of Data Releases crediting the outputs of a student-focused and community-driven manual annotation project, curating gene models and – if required – correcting assembly anomalies, for the Diaphorina citri genome project [46].
Declarations
List of abbreviations
CGEN: Citrus Greening Expression Network; HLB: Huanglongbing; MCOT: Maker, Cufflinks, Oases, and Trinity; NCBI: National Center for Biotechnology Information
Ethical approval
Not applicable.
Consent for publication
Not applicable.
Competing interests
The authors declare that they have no competing interests.
Authors’ contributions
WBH, SJB, TD and LAM conceptualized the study; TD, SS, TDS and SJB supervised the study; SJB, TD, SS, and LAM contributed to project administration; CV, MR, and RN conducted investigation; PH, MF-G, and SS contributed to software development; SS, TDS, PH, and MF-G developed methodology; SJB, TD, WBH, and LAM acquired funding; CV prepared and wrote the original draft; TD, SS, TDS, and SJB reviewed and edited the draft.
Funding
This work was supported by USDA-NIFA grants 2015-70016-23028, HSI 1300394 and 2020-70029-33199.
Acknowledgements
We thank Alistair McGregor and Joshua Benoit for valuable discussions.
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