Identificação e caracterização molecular da família das expansinas no fruto do Coffea arábica L.

Abstract

The mechanism of fruit development in coffee plants is characterized by intense cell division, elongation and relaxation of the cell wall, processes related to the action of certain proteins, including expansins. Expansins (EXPs) represent a gene family and symbol in four subfamilies: α-expansin (EXPA), β-expansin (EXPB), expansin-like A (EXLA) and expansin-like B (EXLB). The aim of this study was to characterize expansins using bioinformatics tools and the molecular characterization of coffee fruits in the perisperm, endosperm and pericarp phases. The characterization of these genes began with the collection of information contained in the NCBI (National Center for Biotechnology Information) and Phytozome databases. Then, the following tools were used: ExPASsy and GRAVY (for physicochemical information); PlantmPLoc (subcellular localization); TAIR (orthologous genes); Gene structure: GSDS (exon/intro); MapGene2chromosomoe (MG2C- physical mapping of chromosomes); sequence alignment using MEGA 11 (phylogenetic analysis); Phyre2 (3D protein structures); MEME (Multiple Em for Motif Elicitation- conserved domains); PlantCARE (gene regulatory regions); and TBtools information on Ka/Ks (non-coincident and symbolic) and collinearity test. For gene expression, C. arabica L. grains were found at four development stages (30 DAF, 90 DAF, 120 DAF and 180 DAF) and subjected to qRT- PCR (quantitative real-time reverse transcription PCR) analysis. Forty-five expansin genes were characterized in silico and their physicochemical patterns contained a variation of 240-300 amino acids (aa), molecular weight 21.18 to 32.27 kDa, isoelectric point (pI) 4.35 to 9.68, and all were located in the cell wall. However, it was observed that their appearances seemed hydrophobic. The gene structure (exon/introns) varied from 2/1 to 6/5. The phylogenetic relationship classified them according to the four expansin subfamilies, forming five groups. The genes were unevenly distributed in the 11 chromosomes of C. arabica L.. The conserved domains varied from two to five. The DPBB and CBM63 domains were described in their 3D structures of each protein. In the

synthesis of the CaEXPA genes, it showed greater similarity with the genomes of Oryza sativa and Arabidopsis thaliana, and the duplication of some genes was positive in the Ka/Ks analysis. The qRT-PCR result showed that the CaEXPA1, CaEXPA2, CaEXPA4, CaEXPA6, CaEXLA10, and CaEXLB11 genes were expressed during the four periods of fruit development. The CaEXPA1 gene showed greater pressure during 30 DAF, 90 DAF, and 120 DAF, while the CaEXPA4 gene was the most expressed among the genes evaluated, having a high expression in all periods of coffee bean development. This information establishes the basis for future research on the functional characteristics and molecular mechanisms of expansion genes in C. arabica L. in different coffee bean development projects.