Utilizing a chromosomal-length genome assembly to annotate the Wnt signaling pathway in the Asian citrus psyllid, Diaphorina citri

Abstract

The Asian citrus psyllid, Diaphorina citri, is an insect vector that transmits Candidatus Liberibacter asiaticus, the causal agent of the Huanglongbing (HLB), or citrus greening disease. This disease has devastated Florida’s citrus industry, and threatens California’s industry as well as other citrus producing regions around the world. To find novel solutions to the disease, a better understanding of the vector is needed. The D. citri genome has been used to identify and characterize genes involved in Wnt signaling pathways. Wnt signaling is utilized for many important biological processes in metazoans, such as patterning and tissue generation. Curation based on RNA sequencing data and sequence homology confirms 24 Wnt signaling genes within the D. citri genome, including homologs for beta-catenin, Frizzled receptors, and seven Wnt-ligands. Through phylogenetic analysis, we classify D. citri Wnt ligands as Wg/Wnt1, Wnt5, Wnt6, Wnt7, Wnt10, Wnt11, and WntA. The D. citri version 3.0 genome with chromosomal length scaffolds reveals a conserved Wnt1-Wnt6-Wnt10 gene cluster with a gene configuration like that in Drosophila melanogaster. These findings provide greater insight into the evolutionary history of D. citri and Wnt signaling in this important hemipteran vector. Manual annotation was essential for identifying high quality gene models. These gene models can be used to develop molecular systems, such as CRISPR and RNAi, which target and control psyllid populations to manage the spread of HLB. Manual annotation of Wnt signaling pathways was done as part of a collaborative community annotation project.

Workflows for Rapid Functional Annotation of Diverse Arthropod Genomes

“…Furthermore, functional annotation provides information about pathways and gene families that are poorly annotated or absent from gene sets, providing useful information that can be used to direct targeted manual curation of genes. Manual curation of gene models is a well-established activity in the arthropod research community using Apollo [49] through community databases, such as the i5k Workspace@NAL [21], VectorBase [50], the Hymenoptera Genome Database [51], Citrus Greening Database [52][53][54][55][56][57][58], and others. Functional annotation would support this focus while extending the utility of the genome for the research community.…”

Diaci v3.0: Chromosome-level assembly,de novotranscriptome and manual annotation ofDiaphorina citri,insect vector of Huanglongbing

“…Particular ligand genes have been lost in various insects [74] and D. citri is no exception. We were unable to find orthologs of Wnt8/D or Wnt9 [54], which are also absent in pea aphids. We also did not find Wnt16 , although it is present in several other hemipterans [70, 76].…”

“…One of the most important lessons learned during this project is that our genome annotation and the student experience both benefit when students are recruited earlier in their academic career (first or second year students) and supported through a longer period than traditional undergraduate research experiences where students typically start research as juniors or seniors. These students can be involved from the start of analysis until publication, with a few generating their own first-author publications [50][51][52][53][54][55]. Students can join and contribute to genome annotation after first year introductory biology courses that cover basic molecular biology, which provides the necessary background to understand gene structure and function.…”

“…The community curation team, comprising undergraduate and graduate students and their mentors (Table 4 above), manually annotated several new sets of genes in the Diaci v3.0 official gene set, OGSv3, focusing our efforts on orthogroups, pathways of interest to the D. citri research community, and genes that have been targeted for RNAi-based pest control in other insects [63][64][65]. Detailed reports for these gene sets are included in 11 accompanying articles published as data releases by Gigabyte [50][51][52][53][54][55]61,62,[66][67][68]. Sequences of the manually curated transcripts and predicted proteins are available for download at citrusgreening.org [8].…”

Quantification of new and archived Diaphorina citri transcriptome data using a chromosomal length D. citri genome assembly reveals the vector’s tissue-specific transcriptional response to citrus greening disease

“…Still, a major roadblock is the functional annotation of the gene models. While automated pipelines for annotation exist at NCBI and elsewhere [63], these efforts are supplemented by manual annotation efforts [64–68] for D. citri and other arthropods [57]. Future work on understanding how the improved genome leads to improved quantification at the proteome level is also needed, and we hope such studies are inspired by the findings we present here.…”