Understanding the Role of Phosphorylation in the Nonsense Mediated mRNA Decay Surveillance Mechanism

Abstract

Transcripts containing premature termination codons and frameshift mutations are targeted for rapid turnover by a RNA surveillance mechanism known as nonsense-mediated mRNA decay (NMD). The proteins encoded by the UPF1, UPF2, UPF3 and HRP1 genes have been shown to play a critical role in the activation of this pathway in yeast. Despite many recent observations that emphasize the importance of NMD in controlling the stability of aberrant mRNAs, little is known about how this process is regulated. Previous studies on UPF1 and UPF2 orthologues in C. elegans and humans have demonstrated that these NMD factors are phosphoproteins. Surprisingly, the S. cerevisiae orthologues, Upf1p and Upf2p, while necessary for NMD, have not been demonstrated to be phosphorylated. Here we provide evidence that both yeast Upf1p and Upf2p are phosphoproteins and that binding of Upf2p to Hrp1p is modulated by the phosphorylation status of Upf2p. These data suggest that similar to humans and nematodes, the activity of the NMD pathway in yeast may be regulated by a phosphorylation and dephosphorylation cycle. Furthermore, the results presented here demonstrate that Upf2p is a cytoplasmic protein in yeast cells. Interestingly, Upf2p is localized to discrete cytoplasmic foci. These findings suggest that NMD might be a compartmentalized process in yeast cells and propose an intricate manner to regulate gene expression. This work is supported by grants from the NIH to C.I.G. (GM008102-3052, U54 Award CA96297-03, KO1 HL-04355-05). P.G. is supported by the MARC Program at UPR-RP 2T34GMO7821)