Structure and expression of Rhodnius prolixus GH18 chitinases and chitinase-like proteins: Characterization of the physiological role of RpCht7, a gene from subgroup VIII, in vector fitness and reproduction

Maiara do Valle Faria Gama; Caroline Silva Moraes; Bruno Gomes; Hector Manuel Diaz-Albiter; Rafael Dias Mesquita; Eloy Seabra-Junior; Patrícia Azambuja; Eloi de Souza Garcia; Fernando Ariel Genta

doi:10.3389/fphys.2022.861620

Structure and expression of Rhodnius prolixus GH18 chitinases and chitinase-like proteins: Characterization of the physiological role of RpCht7, a gene from subgroup VIII, in vector fitness and reproduction

Front Physiol. 2022 Oct 3:13:861620. doi: 10.3389/fphys.2022.861620. eCollection 2022.

Authors

Affiliations

¹ Instituto Oswaldo Cruz, FIOCRUZ, Rio de Janeiro, Brazil.
² El Colegio de la Frontera Sur, ECOSUR, Campeche, Mexico.
³ Departamento de Bioquímica, Instituto de Química, Universidade Federal do Rio de Janeiro, Rio de Janeiro, Brazil.
⁴ Instituto Nacional de Ciência e Tecnologia em Entomologia Molecular, Rio de Janeiro, Brazil.
⁵ Universidade Federal Fluminense, UFF, Rio de Janeiro, Brazil.

Abstract

Chitinases are enzymes responsible for the hydrolysis of glycosidic linkages within chitin chains. In insects, chitinases are typically members of the multigenic glycoside hydrolase family 18 (GH18). They participate in the relocation of chitin during development and molt, and in digestion in detritivores and predatory insects, and they control the peritrophic membrane thickness. Chitin metabolism is a promising target for developing vector control strategies, and knowledge of the roles of chitinases may reveal new targets and illuminate unique aspects of their physiology and interaction with microorganisms. Rhodnius prolixus is an important vector of Chagas disease, which is caused by the parasite Trypanosoma cruzi. In this study, we performed annotation and structural characterization of nine chitinase and chitinase-like protein genes in the R. prolixus genome. The roles of their corresponding transcripts were studied in more depth; their physiological roles were studied through RNAi silencing. Phylogenetic analysis of coding sequences showed that these genes belong to different subfamilies of GH18 chitinases already described in other insects. The expression patterns of these genes in different tissues and developmental stages were initially characterized using RT-PCR. RNAi screening showed silencing of the gene family members with very different efficiencies. Based on the knockdown results and the general lack of information about subgroup VIII of GH18, the RpCht7 gene was chosen for phenotype analysis. RpCht7 knockdown doubled the mortality in starving fifth-instar nymphs compared to dsGFP-injected controls. However, it did not alter blood intake, diuresis, digestion, molting rate, molting defects, sexual ratio, percentage of hatching, or average hatching time. Nevertheless, female oviposition was reduced by 53% in RpCht7-silenced insects, and differences in oviposition occurred within 14-20 days after a saturating blood meal. These results suggest that RpCht7 may be involved in the reproductive physiology and vector fitness of R. prolixus.

Keywords: RNAi; Rhodnius prolixus; chitinase; chitinase-like protein; gene expression; glycoside hydrolase family 18; oviposition.