RNA-Seq analysis of wild-type and ttpA-C231S, ttpA-E236fsx, and ttpA-Y242fsx mutant D. discoideum amebae

In many eukaryotes, mRNAs containing specific AU-rich motifs are regulated by proteins of the tristetraprolin (TTP) family, which bind to these motifs through a tandem zinc finger (TZF) domain. This binding leads to promotion of subsequent deadenylation and decay, partly through a conserved carboxyl-terminal CNOT1 binding domain. We explored the physiological role of the single TTP family member (TtpA) expressed in Dictyostelium discoideum. This study shows the effect of three distinct putative null mutations in the ttpA gene on transcript expression in amebae in growth medium during surface culture under axenic conditions. The cells exhibited similar external and gene expression phenotypes, suggesting that they are all null mutants, despite two different types of mutations (two frame shifts and one amino acid substitution). Dictyostelium discoideum amebae (strain AX3) were grown in normal growth medium under axenic conditions in plastic flasks until they were approximately 80% confluent. Four cultures of wild-type cells and four cultures of each type of mutant cells were then frozen and used for RNA extraction and RNA-Seq analysis.