Effect of depletion of ZFC3H1, PABPN1 and CNOT4 on gene expression in human cells.

Codon usage bias of an organism has been reported to affect the level of gene expression by influencing this process at multiple steps. Here, we have identified a role of poly-A exosome targeting complex (PAXT), which is a substrate adaptor of nuclear RNA exosome, in regulating codon usage effect on gene expression. PAXT complex subunits, ZFC3H1 and PABPN1, regulate stability and subcellular localization of mRNA in a codon usage dependent manner. This suggests that codon usage dependent mRNA stability and nuclear export are important factors of codon usage effect on gene expression in human cells. Also, a subunit of CCR4-NOT complex, CNOT4, was found to play a significant role in this regulation by affecting the nuclear mRNA levels and their subcellular localization in codon usage dependent manner. Overall design: To investigate a role of nuclear exosome PAXT complex and CCR4-NOT complex subunit, CNOT4, in the regulation of codon-usage effect on gene expression, we depleted ZFC3H1, PABPN1 and CNOT4 in HEK293 cells using siRNA for 72 h. Control (treated with negative control siRNA for 72 h) and knockdown cells were harvested and total, cytosolic and nuclear RNA fractions were isolated. RNA-seq analysis was performed using total, cytosolic and nuclear RNA of control and knockdown cells.