Abstract
Arginine methylation is a common posttranslational modification that has far-reaching cellular effects. Trypanosoma brucei is an early-branching eukaryote with four characterized protein arginine methyltransferases (PRMTs), one additional putative PRMT, and over 800 arginine methylated proteins, suggesting that arginine methylation has widespread impacts in this organism. While much is known about the activities of individual T. brucei PRMTs (TbPRMTs), little is known regarding how TbPRMTs function together in vivo. In this study, we analyzed single and selected double TbPRMT knockdowns for the impact on expression of TbPRMTs and global methylation status. Repression of TbPRMT1 caused a decrease in asymmetric dimethylarginine and a marked increase in monomethylarginine that was catalyzed by TbPRMT7, suggesting that TbPRMT1 and TbPRMT7 can compete for the same substrate. We also observed an unexpected and strong interdependence between TbPRMT1 and TbPRMT3 protein levels. This finding, together with the observation of similar methyl landscape profiles in TbPRMT1 and TbPRMT3 repressed cells, strongly suggests that these two enzymes form a functional complex. We show that corepression of TbPRMT6/7 synergistically impacts growth of procyclic-form T. brucei. Our findings also implicate the actions of noncanonical, and as yet unidentified, PRMTs in T. brucei. Together, our studies indicate that TbPRMTs display a functional interplay at multiple levels.
Highlights
Arginine methylation is a common posttranslational modification entailing the transfer of a methyl group from the methyl donor S-adenosylmethionine (AdoMet) to the terminal nitrogen of a peptidyl arginine
TbPRMT6 RNA was increased to 127% of wild-type levels protein levels were unchanged, suggesting that the modest decrease in TbPRMT3 RNA is within the range of experimental error
We report the impacts of repressing specific T. brucei PRMTs (TbPRMTs) both on the levels of other TbPRMTs and the global cellular methyl landscape
Summary
Arginine methylation is a common posttranslational modification entailing the transfer of a methyl group from the methyl donor S-adenosylmethionine (AdoMet) to the terminal nitrogen of a peptidyl arginine. Arginine methylation affects myriad cellular functions including transcriptional regulation, mRNA splicing, DNA repair, and signal transduction (Bedford and Clarke 2009). The addition of a methyl group does not affect the charge of the arginine residue, it does increase its mass which can have both positive and negative effects on its hydrogen bonding capabilities (Horowitz and Trievel 2012). RNA-binding proteins are key regulators of parasite function. RNA-binding proteins constitute a large number of arginine methylated proteins in higher organisms (Pahlich et al 2006; Bedford and Clarke 2009), and this prompted us to examine the arginine methylome of T. brucei to ask if the same is true in this early-branching a 2014 The Authors.
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