Perfil de expressão e análise filogenética dos genes da proteína desacopladora mitocondrial durante o desenvolvimento e estresse em soja [Glycine max (L.) Merr.]

Abstract

Several studies have evidenced that the main function of the mitochondrial uncoupling protein in plants (pUCP) is related to reactive oxygen species (ROS) regulation. In silico analysis suggests the existence of multigenic families to pUCPs codification, however further studies are yet needed to establish the pUCPs genetic expression profile, just like the gene amount in each species and their phylogenetic relations. The current work had as objective to characterize, analyze phylogeneticly and evaluate the expression profile of the pUCP multigenic family in different tissues during the soybean development [Glycine max (L.) MERR.] and in stress conditions. It has been performed an in silico analysis on the soybean genome and on other legumes available in the database WGS, revealing a codifier multigenic family for pUCP, UCP1 and 2 with 9 exons, UCP3 with 2 exons, and UCP4 and 5 with only 1 exon. Amongst the legumes analyzed, the soybean stood out with the greater number of genes, 10 genes in total, giving four GmUCP1 genes, one GmUCP2, one GmUCP3, two GmUCP4 and two GmUCP5, along with the presence of an alternative splicing on GmUCP1b1 gene. Specific primers have been designed for each GmUCP member in order to analize the expression profiles in different tissues (dry and doused seed, flowers, pods, cotyledons, unifoliate and trifoliate leaves, roots, hypocotyl and epicotyl) during the soybean development. For the assays in stress conditions have been used soybean leaves and roots with thirteen days after sowing (DAS) which have been subjected to osmotic stress caused by the application of polyethylene glycol (PEG) and biotic stress caused by salicylic acid (SA). The total RNA from each sample has been extracted in order to perform the RT-qPCR. The ct values have been obtained through the realplex program and analyzed through the GeNorm program. The genetic expression profile has shown that all genes were expressed in every tissue/organ analyzed during the soybean development, with the exception on some genes in dry seeds and epicotyl. The different expression profiles of each gene during the development of each tissue/organ suggest that occurs a spatial/temporal gene regulation among the GmUCP members. The expression profiles of the GmUCP genes in soybean during the stress conditions have varied, once 2 genes have shown steady expression in both tissues/ stress, 7 genes have shown a drop in the expression profile, while only 4 genes have shown an increase of the transcript levels.