Characterization of CAPA peptide and its receptor in Bemisia tabaci (Gennadius)

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dc.contributor.advisorJindal, Vikas
dc.contributor.authorSudeshna
dc.date.accessioned2022-07-30T14:55:22Z
dc.date.available2022-07-30T14:55:22Z
dc.date.issued2022
dc.description.abstractBemisia tabaci (Gennadius) is a species complex having economic importance with worldwide distribution. Being an exclusive phloem feeder, diuresis is a key physiological process that helps in excreting out the waste substances. CAPA peptide and its G-protein coupled receptor (GPCR) interaction is key factor responsible for diuresis process in insects. The present investigations aimed to functionally characterize CAPA receptor (CAPAr) gene, its expression profile and characterize the CAPA peptides activating the CAPAr in B. tabaci AsiaII 1. We identified two isoforms of CAPAr gene and named these as BtabCAPAr-1 (1263 bp, 421aa) and BtabCAPAr-2 (1065 bp, 355aa) that showed 98.26% and 98.56% identity with the predicted CAPAr gene sequence available in NCBI GenBank, respectively. The isoform differs by missing of 198 nucleotides and 66 aa residues in the BtabCAPAr-2 as compared to BtabCAPAr-1. The nucleotide sequence alignment of BtabCAPAr-1 and BtabCAPAr-2 isoforms with the reference gene (XM_019044129.1) revealed the nucleotide substitutions at 22 and 16 positions, respectively. Amino acid sequence alignment revealed that ‘Alanine’ in the predicted gene (XM_019044129.1) was replaced by ‘Glutamine’ in BtabCAPAr isoforms. BtabCAPAr-1 and BtabCAPAr-2 isoforms code for six and five exons, respectively, however, in BtabCAPAr-2 isoform, exon 3 was found missing. The BtabCAPAr-1 and BtabCAPAr-2 have seven and five transmembrane domain polypeptide, respectively. Phylogenetic analysis showed that BtabCAPAr gene isoforms and the CAPA receptor gene in the other insect orders forms formed a single clade, which depicts close association among them. The expression of BtabCAPAr gene was significantly high in the adult stage (3.76 fold) in comparison to the egg. The expression in egg, nymphal stages and pupal stage was statistically at par. RNAi of CAPAr gene in B. tabaci Asia II 1 resulted into reduced survival and fecundity of whitefly. Significantly high adult mortality was recorded when whiteflies were fed with dsBtabCAPAr @1.0 µg/µl (30.04%) in comparison to dsgfp @1.0 µg/µl (7.29%) after 96 hr of dsRNA feeding. After 48 hr of dsRNA feeding, mortality was significantly higher in dsBtabCAPAr @1.0 µg/µl (30.74%) in comparison to dsgfp @1 µg/µl (1.14%). The minimum fecundity was recorded in dsBtabCAPAr @1.0 µg/µl (53.51 eggs/ female) which was statistically on a par with dsBtabCAPAr @0.5 µg/µl (59.30 eggs/ female) and dsBtabCAPAr @0.1 µg/µl (61.76 eggs/ female) but significantly lower than dsgfp @1 µg/µl (85.79 eggs/ female) and sucrose (92.98 eggs/ female). The effect of dsBtabCAPAr feeding on the development duration of egg, nymphs, pupa and total development period and egg and nymphal mortality was found to be non-significant. The q-RT-PCR studies further confirmed the downregulation of CAPAr gene expression by 44.00 and 41.00 per cent in whitefly adults fed with dsBtabCAPAr @1.0 µg/µl and dsBtabCAPAr @0.5 µg/µl, respectively. The dsRNA in the artificial diet was found to be stable in concentrations viz. dsBtabCAPAr @1.0 µg/µl, dsBtabCAPAr @0.5 µg/µl and dsgfp 1.0 µg/µl upto 96 hr of feeding. Feeding and internalization of artificial diet was confirmed by observing blue stained whitefly adults fed with synthetic food dye (1%). The pharmacological characterization of BtabCAPAr-1 isoform was carried out through heterologous expression assay in Chinese Hamster Ovary cells. The BtabCAPAr-1 isoform responded strongly to CAPA-2 (EC50 = 0.053 nM) followed by CAPA-1 peptide (EC50 = 0.067 nM). However, BtabCAPAr-1 did not respond strongly to CAPA-3 peptide (EC50 = 1147 nM). The study showed high mortality and low fecundity when BtabCAPAr gene was silenced and CAPA-2 and CAPA-1 were identified as authentic peptides, which activate BtabCAPAr gene. These results proved that CAPAr gene may be potential target for development of novel pesticides through RNAi and peptidomimetics approach for effective management of whitefly.en_US
dc.identifier.citationSudeshna (2022) Characterization of CAPA peptide and its receptor in Bemisia tabaci (Gennadius) (Unpublished Ph.D. Dissertation). Punjab Agricultural University, Ludhiana, Punjab, India.en_US
dc.identifier.urihttps://krishikosh.egranth.ac.in/handle/1/5810185959
dc.keywordsBemisia tabaci, G-protein coupled receptor, gene isoforms, CAPA peptide, gene expression, RNA interference, heterologous expression assayen_US
dc.language.isoEnglishen_US
dc.pages108en_US
dc.publisherPunjab Agricultural University, Ludhianaen_US
dc.research.problemCharacterization of CAPA peptide and its receptor in Bemisia tabaci (Gennadius)en_US
dc.subEntomologyen_US
dc.themeCharacterization of CAPA peptide and its receptor in Bemisia tabaci (Gennadius)en_US
dc.these.typePh.Den_US
dc.titleCharacterization of CAPA peptide and its receptor in Bemisia tabaci (Gennadius)en_US
dc.typeThesisen_US
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