Molecular characterization and expression analysis of PPP1R3C in hypoxia-tolerant Indian catfish, Clarias batrachus (Linnaeus, 1758) under hypoxia

Abstract

Hypoxia is an important environmental stressor that leads to rapid adaptive changes in metabolic organization. However, the molecular mechanisms of hypoxia tolerance in fish remain largely unknown. The present work was focused on understanding the molecular mechanisms and signaling pathways that may lead to tolerance of Clarias batrachus to hypoxic stress. Protein phosphatase 1 regulatory subunit 3C (PPP1R3C) is a new hypoxia-inducible factor (HIF) targeted gene and is regulated by HIF-1 under hypoxic conditions. Overexpression of PPP1R3C increases glycogen accumulation through activation of several enzymes and processes. In this study, for the first time, full length cDNA of PPP1R3C from C. batrachus was characterized and its expression pattern in the brain, liver, muscle and spleen under short (progressive hypoxia; PH, 1h, 6h and 12h) and long-term (natural) hypoxic conditions was investigated. The complete cDNA of PPP1R3C was of 1499 bp, encoding 285 amino acid residues. The identified protein had a protein phosphatase 1 binding motif and a carbohydrate binding domain, thought to be involved in the regulation of glycogen metabolism. Short-term hypoxia exposure caused significant increase in PPP1R3C transcripts in the liver (6h; 6.96 fold and 12h; 3.91 fold) and muscle (progressive hypoxia; 3.46 fold), while, after long-term hypoxia exposure, significant up-regulation in the liver (7.77 fold) and spleen (6.59 fold) tissues was observed. No significant differences were observed in the brain for any time periods. Thus PPP1R3C may play an important role in the tolerance of C. batrachus to hypoxia.

Keywords: CBD21; Clarias batrachus; Expression; HIF; Hypoxia; MEGA4; MW; Molecular Evolutionary Genetics Analysis version 4; ORF; PPP1R3C; PTG; UTR; carbohydrate binding domain type 21; hypoxia inducible factor; isoelectric point; kDa; kilodalton; molecular weight; open reading frame; pI; poly adenylational; polyA(+); protein targeting to glycogen; qRT-PCR; quantitative real-time PCR; rRNA; ribosomal RNA; untranslated region.