Abstract
The protein arginine methyltransferaseas (PRMTs) family is conserved from yeast to human, and regulates stability, localization and activity of proteins. We have characterized deletion strains corresponding to genes encoding for PRMT1/3/5 (designated amt-1, amt-3 and skb-1, respectively) in Neurospora crassa. Deletion of PRMT-encoding genes conferred altered Arg-methylated protein profiles, as determined immunologically. Δamt-1 exhibited reduced hyphal elongation rates (70% of wild type) and increased susceptibility to the ergosterol biosynthesis inhibitor voriconazole. In ▵amt-3, distances between branches were significantly longer than the wild type, suggesting this gene is required for proper regulation of hyphal branching. Deletion of skb-1 resulted in hyper conidiation (2-fold of the wild type) and increased tolerance to the chitin synthase inhibitor polyoxin D. Inactivation of two Type I PRMTs (amt-1 and amt-3) conferred changes in both asymmetric as well as symmetric protein methylation profiles, suggesting either common substrates and/or cross-regulation of different PRMTs. The PRMTs in N. crassa apparently share cellular pathways which were previously reported to be regulated by the NDR (Nuclear DBF2-related) kinase COT1. Using co-immunprecipitation experiments (with MYC-tagged proteins), we have shown that SKB1 and COT1 physically interacted and the abundance of the 75 kDa MYC::COT1 isoform was increased in a Δskb-1 background. On the basis of immunological detection, we propose the possible involvement of PRMTs in Arg-methylation of COT1.
Highlights
Methylated derivatives of arginine were identified 45 years ago [1]
In the present study we report on the involvement of protein arginine methyltransferases (PRMTs) as mediators of post translational modifications that affect growth and development in N. crassa and focus on the possible cross-talk between arginine methylation and phosphorylation, specific to the interactions between PRMTs and the NDR kinase COT1
All the PRMT-encoding deletion strains produced in this study were viable, radial growth was slower than the wild type.The amt-1;skb-1 double mutant, exhibited abnormal hyphal morphology (Fig S1)
Summary
Methylated derivatives of arginine were identified 45 years ago [1]. Since protein methylation has been recognized as a post translational modification that plays regulatory roles in signal transduction, nucleic transport, activation and repression of genes and mRNA splicing [2]. Arginine methylation is catalyzed by protein arginine methyltransferases (PRMTs) that since being identified [3,4] have been found in the genomes of many eukaryotic organisms [5]. Type I and II (conserved from yeast to human) are responsible for monomethylation. They catalyze asymmetric (aDMA) and symmetric (sDMA) dimethylarginine, respectively. Type III and IV PRMTs catalyze the monomethylation of arginine residues and the formation of monomethylarginine on guanidinium nitrogen, respectively. In human cells there are nine genes considered to encode for PRMTs. Among them, only PRMT1 and PRMT3 (Type I), and PRMT5 (Type II) are conserved through eukaryotic evolution, including members of the fungal kingdom [6]
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