mRNAs coding for A1 and A2 Isoforms of translation factor eEF1 demonstrate different half-lives while A1 and A2 proteins are similarly stable in MCF7 cells

Abstract

Eukaryotic translation elongation factor 1A (eEF1A) exists as two 98 % homologous isoforms eEF1A1 and eEF1A2 that are tissue/development specific and differentially linked to apoptosis/cancerogenesis. A2 is overexpressed in a number of tumors while unusual expression of A1 is observed in injured muscles. To approach a possible mechanism underlying induced changes in the relative amounts of the isoforms we examined the intrinsic stability of the proteins and their mRNAs in human cancer cells. Aim. To estimate half-life of the isoforms of eEF1A at mRNA and protein level in human cancer cells. Methods. To measure mRNA stability the transcriptional block technique was applied, with subsequent analysis of the mRNA level by qPCR. To determine the protein decay rate the translation was blocked by cycloheximide and changes in the protein level were detected by Western blot. Results. Calculation of the protein stability revealed half-life of 72 for eEF1A1 and 95 hours for eEF1A2. Half-life of EEF1A1 and EEF1A2 mRNAs were 3 and 60 hours respectively. Conclusions. Despite similar protein stability, the isoforms of eEF1A dramatically differ in the half-lives of their mRNAs, suggesting that the mRNA decay mechanism is one of the main regulators of eEF1A1/A2 amount in MCF7 cancer cells.