Investigação do neuropeptídeo F curto (sNPF) e de seu receptor (sNPFR) em tecidos de Rhodnius prolixus: implicações no controle do vetor da doença de Chagas

Abstract

Neuropeptides regulate physiological processes in arthropods, such as feeding and behavior. Primarily produced in the central nervous system, they interact with G protein-coupled receptors (GPCRs), triggering intracellular signaling cascades. Among them, the short neuropeptide F (sNPF) and its receptor, sNPFR, stand out for their role in regulating feeding and immunity, exhibiting evolutionary conservation and functional similarity to the vertebrate neuropeptide Y (NPY). In Rhodnius prolixus, a single precursor protein generates an active form of sNPF. However, the coding sequence of its receptor remains incompletely elucidated. In hematophagous insects, understanding feeding behavior is crucial for vector control strategies. Unlike other insects, the R. prolixus genome contains three contiguous partial genes (RPRC002266, RPRC002268, and RPRC002269) associated with the sNPFR receptor. Thus, this study aims to identify the complete sequence of sNPFR, perform an in silico analysis of the sNPF and sNPFR proteins—including three-dimensional modeling of their structures and the prediction of protein-ligand binding energy via molecular docking—as well as investigate their expression profiles in different tissues. Total RNA was extracted from nymph tissues, treated with DNase I, and reverse-transcribed into cDNA. PCR reactions using specific primers for sNPFR genes amplified the target sequence, which was subsequently cloned and sequenced. Amino acid translation was performed using the Expasy tool. Protein topology was predicted using DeepTMHMM, while gene expression in different tissues was evaluated by qPCR, with RproR18S as the reference gene. Sequencing revealed three coding regions: exon 1 in the RPRC002266 gene, exon 2 in RPRC002268, and exon 2 in RPRC002269. The resulting protein contains seven transmembrane domains, characteristic of GPCRs, with conserved positions. The highest sNPFR transcript levels were observed in the brain, followed by the gut, Malpighian tubule, and fat body. Transcripts for sNPF were found in the brain and fat body. Based on the genome annotation showing three genes for sNPFR, this may represent a trans-splicing event, which has been previously described in Hemiptera. The broad expression of sNPFR reinforces its diverse physiological role, while sNPF appears to have more specific functions in the brain and fat body. This study represents an initial step toward elucidating the sNPF/sNPFR signaling pathway in R. prolixus, opening new avenues for studying the behavior of this insect.