Serine phosphorylation of protein arginine methyltransferase Hmt1 is critical for controlling its protein levels

In eukaryotes, protein arginine methylation is a prevalent post-translational modification found in a multitude of proteins responsible for key biological processes, ranging from transcription to signaling. One model suggests that phosphorylation of serine 9 (S9) in the Saccharomyces cerevisiae major protein arginine methyltransferase Hmt1 is critical for its oligomerization and activity. In this study, we used classic biochemical approaches to demonstrate that neither the S9 phosphomimetic nor the non-phosphorylatable substitution mutants of Hmt1 affect its oligomerization. These mutants remain active in vivo, retaining their ability to methylate the SR-/hnRNP-like protein Npl3 and displaying a monomethylarginine and asymmetric dimethylarginine banding profile similar to that of the wild-type. In cells lacking Dbf2, the proposed kinase responsible for phosphorylating Hmt1 at S9, Npl3 remains methylated. Additionally, monomethylarginine and asymmetric dimethylarginine banding profiles in cells lacking Dbf2 mostly resemble those observed in the wild-type rather than in hmt1Δ cells. Synchronized yeast cells expressing either S9 substitution exhibit entry into the M phase of the cell cycle at a rate similar to that of both wild-type and hmt1Δ cells. Our results suggest that the C-terminal epitope tagging of Hmt1 is responsible for the previously observed loss of enzymatic activities, rather than the S9 phosphorylation status of Hmt1. Finally, we demonstrate that S9 phosphorylation plays a role in maintaining Hmt1 protein levels in vivo. Overall, our finding demonstrates a novel role for Hmt1 S9 phosphorylation in tuning its in vivo protein levels.