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A single gene, csuF, has been previously shown to encode the protein for the outer membrane of Bacteroides thetaiotaomicron. Identification and characterization showed that the CsuF protein does not have sequences similar to that of Bacteroides chondroitin lyase II despite having the same predicted molecular weight and phenotype. Amino acid sequence analysis of CsuF revealed features suggestive of an outer membrane protein and a suggested a role in utilization of chondroitin sulfate and hyaluronic acid.

Abstract Background Protein arginine methylation is a post-translational modification involved in important biological processes such as transcription and RNA processing. This modification is catalyzed by both type I and II protein arginine methyltransferases (PRMTs). One of the most conserved type I PRMTs is PRMT1, the homolog of which is Hmt1 in Saccharomyces cerevisiae. Hmt1 has been shown to play a role in various gene expression steps, such as promoting the dynamics of messenger ribonucleoprotein particle (mRNP) biogenesis, pre-mRNA splicing, and silencing of chromatin. To determine the full extent of Hmt1’s involvement during gene expression, we carried out a genome-wide location analysis for Hmt1. Results A comprehensive genome-wide binding profile for Hmt1 was obtained by ChIP-chip using NimbleGen high-resolution tiling microarrays. Of the approximately 1000 Hmt1-binding sites found, the majority fall within or proximal to an ORF. Different occupancy patterns of Hmt1 across genes with different transcriptional rates were found. Interestingly, Hmt1 occupancy is found at a number of other genomic features such as tRNA and snoRNA genes, thereby implicating a regulatory role in the biogenesis of these non-coding RNAs. RNA hybridization analysis shows that Hmt1 loss-of-function mutants display higher steady-state tRNA abundance relative to the wild-type. Co-immunoprecipitation studies demonstrate that Hmt1 interacts with the TFIIIB component Bdp1, suggesting a mechanism for Hmt1 in modulating RNA Pol III transcription to regulate tRNA production. Conclusions The genome-wide binding profile of Hmt1 reveals multiple potential new roles for Hmt1 in the control of eukaryotic gene expression, especially in the realm of non-coding RNAs. The data obtained here will provide an important blueprint for future mechanistic studies on the described occupancy relationship for genomic features bound by Hmt1.

The role played by the protein arginine methyltransferase Hmt1 in the preservation and establishment of silent chromatin domains is examined. The results have shown a novel function for arginine methylation in the maintenance of genome stability and the establishment of silent chromatin.

The discovery of four new factors required for mRNA export through a screen of all annotated nonessential Saccharomyces cerevisiae genes is reported. Results demonstrate coupling of mRNA surveillance to mRNA export and suggest specificity of the RNA surveillance machinery for different transcript populations.

Hmt1 is recruited during the beginning of the transcriptional elongation process. The research data suggest a model whereby Hmt1 affects transcriptional elongation and influences recruitment of RNA-processing factors.

Signaling levels within sensory neurons must be tightly regulated to allow cells to integrate information from multiple signaling inputs and to respond to new stimuli. Herein we report a new role for the cGMP-dependent protein kinase EGL-4 in the negative regulation of G protein-coupled nociceptive chemosensory signaling. C. elegans lacking EGL-4 function are hypersensitive in their behavioral response to low concentrations of the bitter tastant quinine and exhibit an elevated calcium flux in the ASH sensory neurons in response to quinine. We provide the first direct evidence for cGMP/PKG function in ASH and propose that ODR-1, GCY-27, GCY-33 and GCY-34 act in a non-cell-autonomous manner to provide cGMP for EGL-4 function in ASH. Our data suggest that activated EGL-4 dampens quinine sensitivity via phosphorylation and activation of the regulator of G protein signaling (RGS) proteins RGS-2 and RGS-3, which in turn downregulate Gα signaling and behavioral sensitivity. [ABSTRACT FROM AUTHOR]

G protein-coupled receptors are 7-pass transmembrane receptors that couple to heterotrimeric G proteins to mediate cellular responses to a diverse array of stimuli. Understanding the mechanisms that regulate G protein-coupled receptors is crucial to manipulating their signaling for therapeutic benefit. One key regulatory mechanism that contributes to the functional diversity of many signaling proteins is post-translational modification. Whereas phosphorylation remains the best studied of such modifications, arginine methylation by protein arginine methyltransferases is emerging as a key regulator of protein function. We previously published the first functional evidence that arginine methylation of G protein-coupled receptors modulates their signaling. We report here a third receptor that is regulated by arginine methylation, the Caenorhabditis elegans SER-2 tyramine receptor. We show that arginines within a putative methylation motif in the third intracellular loop of SER-2 are methylated by PRMT5 in vitro. Our data also suggest that this modification enhances SER-2 signaling in vivo to modulate animal behavior. The identification of a third G protein-coupled receptor to be functionally regulated by arginine methylation suggests that this post-translational modification may be utilized to regulate signaling through a broad array of G protein-coupled receptors. [ABSTRACT FROM AUTHOR]

Cotranscriptional recruitment of pre-mRNA splicing factors to their genomic targets facilitates efficient and ordered assembly of a mature messenger ribonucleoprotein particle (mRNP). However, how the cotranscriptional recruitment of splicing factors is regulated remains largely unknown. Here, we demonstrate that protein arginine methylation plays a novel role in regulating this process in Saccharomyces cerevisiae. Our data show that Hmt1, the major type I arginine methyltransferase, methylates Snp1, a U1 small nuclear RNP (snRNP)-specific protein, and that the mammalian Snp1 homolog, U1-70K, is likewise arginine methylated. Genome-wide localization analysis reveals that the deletion of the HMT1 gene deregulates the recruitment of U1 snRNP and its associated components to intron-containing genes (ICGs). In the same context, splicing factors acting downstream of U1 snRNP addition bind to a reduced number of ICGs. Quantitative measurement of the abundance of spliced target transcripts shows that these changes in recruitment result in an increase in the splicing efficiency of developmentally regulated mRNAs. We also show that in the absence of either Hmt1 or of its catalytic activity, an association between Snp1 and the SR-like protein Npl3 is substantially increased. Together, these data support a model whereby arginine methylation modulates dynamic associations between SR-like protein and pre-mRNA splicing factor to promote target specificity in splicing. [ABSTRACT FROM AUTHOR]

Establishment and maintenance of silent chromatin in the Saccharomyces cerevisiae involves a step-wise assembly of the SIR complex. Here we demonstrate a role for the protein arginine methyltransferase Hmt1 in this process. In the absence of catalytically active Hmt1, yeast cells display increased transcription from silent chromatin regions and increased mitotic recombination within tandem repeats of rDNA. At the molecular level, loss of Hmt1's catalytic activity results in decreased Sir2 and dimethylated Arg-3 histone H4 occupancy across silent chromatin regions. These data suggest a model whereby protein arginine methylation affects the establishment and maintenance of silent chromatin. [ABSTRACT FROM AUTHOR]

Protein arginine methylation has emerged as a key post-translational modification responsible for many facets of eukaryotic gene expression. To better understand the extent of this modification in cellular pathways, we carried out bioorthogonal methylation profiling in Saccharomyces cerevisiae to comprehensively identify the in vivo substrates of the major yeast protein arginine methyltransferase Hmt1. Gene ontology analysis of candidate substrates revealed an enrichment of proteins involved in the process of translation. We verified one such factor, eIF1A, by in vitro methylation. Three sites on eIF1A were found to be responsible for its methylation: R13, R14, and R62, with varied capacity by which each site contributed to the overall methylation capacity in vitro . To determine the role of methylation in eIF1A function, we used a battery of arginine-to-alanine substitution mutants to evaluate translation fidelity in these mutants. Our data show that substitution mutants at R13 and R14 in the N-terminal tail improved the fidelity of start codon recognition in an initiation fidelity assay. Overall, our data suggest that Hmt1-mediated methylation of eIF1A fine-tunes the fidelity of start codon recognition for proper translation initiation.

Protein arginine methylation has emerged to be an important regulator of cellular protein functions. Techniques that uncover the presence of methylarginines on a protein provide an important step towards understanding the functional role of arginine methylation. Here, we describe several common methods used to detect the presence of protein arginine methylation in Saccharomyces cerevisiae.