Structural and functional insights into the Diabrotica virgifera virgifera ATP-binding cassette transporter gene family

Abstract

Background The western corn rootworm, Diabrotica virgifera virgifera , is a pervasive pest of cultivated maize in North America and Europe, which has adapted to survive exposure to multiple insecticidal agents. Due to their role in insecticide transport, we sought to identify members of the ATP-binding cassette (ABC) transporter family in D. v. virgifera using a transcriptomics approach.Results In this study, 65 putative D. v. virgifera ABC ( Dvv ABC) transporters were identified within a combined transcriptome assembly generated from embryonic, larval, adult male, and adult female RNA-sequence libraries. Phylogenetic analysis placed the deduced amino-acid sequences of the Dvv ABC transporters into eight subfamilies (A to H). Of these, eight shared structural and functional conservation with Tribolium castaneum ABC transporter orthologs known to exhibit overt RNA interference (RNAi) knockdown phenotypes. Interestingly, depletion of DvvABCB_19147 and DvvABCG_3712 transcripts in adult females produced detrimental reproductive and developmental phenotypes (egg-laying or -hatching defects), demonstrating the potential of these genes as targets for RNAi-mediated insect control tactics.Conclusions By combining sequence data from four libraries covering three distinct life stages, we have produced a relatively comprehensive de novo transcriptome assembly for D. v. virgifera . Moreover, we have identified 65 members of the ABC transporter family, and provided the first insights into the developmental and physiological roles of ABC transporters in this pest species.

A draft Diabrotica virgifera virgifera genome: insights into control and host plant adaption by a major maize pest insect

“…ATP-binding cassette transporters comprise one of the largest gene families wherein subfamilies A-H in arthropods function in a variety of transmembrane transport activities [114], including involvement in development [115] and detoxification [116], and potentially in Bt resistance mechanisms. There are 188 complete and partial RefSeq annotated ABC transporters in Dvir_2.0, which is greater than a transcriptome-based estimate of 65 from a prior D. v. virgifera transcriptome-based estimate [117]. Phylogenetic reconstruction using 93 Dvir_2.0 models containing a full-length NDB placed 10 members into an ABCB clade, suggesting the number of orthologs are nearly double that predicted in T. castaneum and equal to that in A. viridicyanea.…”

“…The unordered genomic contigs generated by SOAPdenovo were joined based on sequential gapped alignments with two transcript assemblies. The first round of transcript-guided scaffolding used 87,996 transcripts assembled across different growth stages (NCBI TSA accession GHNJ00000000.1 [117] to query against the D. v. virgifera SOAPdenovo GapCloser assembly with the BLASTlike alignment tool (BLAT) [163]. Searches used default parameters, except that the resulting.psl file header was suppressed (-noHead) and minimum sequence identity (-minIdentity) was set at 0.95.…”

“…TP binding cassette (ABC) transporters are involved in Bt resistance and xenobiotic efflux mechanisms. This gene family was identified by merging evidence from three searches: 1) keyword search of Dvir_2.0 RefSeq protein model fasta headers (gene descriptions) with "ABC transporter" and "multidrug resistance protein"; 2) importation of all D. v. virgifera NCBI Dvir_2.0 RefSeq protein models into a local database that was queried with T. castaneum orthologs retrieved from the UniProt database (UniProt Consortium 2015; orthologs D2A4C6, D6WES5, D6WK76, D2A4N9, D6WDY5, D6WD20, D2A232, and D6X2Z9) using BLASTp [182], with results filtered for an E-value ≤ 10 -20 and high-scoring segment pairs (HSP) length ≥ 25 amino acids; and 3) query of the above local BLAST database of Dvir_2.0 protein models with 65 ABC transporter protein sequences in silico translated from a prior D. v virgifera transcriptome assembly [117] with results filtered at an E-value cutoff ≤ 10 -60 and percent identity ≥ 90%. The unique non-redundant set of putative ABC transporter models with ≤ 80% coverage of corresponding T. castaneum orthologs or < 100% coverage of previously predicted D. v virgifera ABC transporter proteins were considered putative partial coding regions, and reconstructed in instances when a Dvir_2.0 gene model showed nonoverlapping alignment to > 1 query.…”

Comparative genomic analysis of ABC transporter genes in Tenebrio molitor and four other tenebrionid beetles (Coleoptera: Tenebrionidea)

“…We performed a maximum likelihood analysis using the ABC transporters identified in the present study and those from T. castaneum and D. virgifera virgifera (Adedipe et al, 2019; Broehan et al, 2013). Phylogenetic analysis indicated that they are classified into eight subfamilies (ABCA‐H) with high bootstrap values, among which ABCC and ABCG were the largest subfamilies (Figure 2).…”

“…Multiple amino acid sequence alignment of the ABC transporters identified in this study and those from T. castaneum (Broehan et al, 2013) and Diabrotica virgifera virgifera identified from transcriptomic data (Adedipe et al, 2019) were performed using the MUSCLE package with default parameters in MEGA X (Kumar et al, 2018). ABC transporters used for phylogenetic analysis are listed in Supporting Information: Table .…”