Protein Arginine Methyltransferase Prmt5-Mep50 Methylates Histones H2A and H4 and the Histone Chaperone Nucleoplasmin in Xenopus laevis Eggs

Abstract

Background: Histones are specifically modified in the Xenopus egg. Results: A complex of Prmt5 and Mep50 methylates histones H2A and H4 and the histone chaperone nucleoplasmin on an arginine in a conserved motif. Conclusion: Arginine methylation is enriched in the egg and targets chromatin-acting proteins. Significance: Histone arginine methylation probably results in specification of the pluripotent developmental program.

Developmentally Regulated Post-translational Modification of Nucleoplasmin Controls Histone Sequestration and Deposition

“…Glutamyltransferase activity exists within Xenopus oocyte extracts (van Dijk et al, 2007) and egg extracts ( data not shown ), consistent with our observed Npm glutamylation and our in vitro TTLL4 modification of Npm. We previously demonstrated that PRMT5-MEP50 is abundant in eggs and methylates Npm (Wilczek et al, 2011). Phosphorylation of Npm is catalyzed by CK2 (Taylor et al, 1987; Vancurova et al, 1995) and the cyclin-dependent kinase cdc2/cdk1 (Cotten et al, 1986).…”

“…We used the following antibodies in this study: a purified monoclonal Npm antibody (Dilworth et al, 1987), polyclonal rabbit antibody generated against full length recombinant Npm, anti-glutamylation TTbIII, SG I polyclonal antibodies (gift of Dr. Frankfurter, Spano and Frankfurter, 2010), and histone and methylarginine polyclonal antibodies (Wilczek et al, 2011). …”

“…Npm with Ser to Asp phosphomimetic mutations on predicted, but not known, phosphorylation sites showed an increase in affinity for histones H2A-H2B (Taneva et al, 2009). We previously showed that PRMT5 methylates Npm on its C-terminus (Wilczek et al, 2011). Glutamylation, an isopeptide addition of a glutamic acid to the γ-carboxyl of a primary chain glutamate residue occurs on the Npm-family member Nucleophosmin (Npm1) (van Dijk et al, 2008).…”

Histone H2A and H4 N-terminal Tails Are Positioned by the MEP50 WD Repeat Protein for Efficient Methylation by the PRMT5 Arginine Methyltransferase

“…laevis PRMT5-MEP50 complex was produced and purified as described (5). XlMEP50 mutagenesis was performed by mutating Arg-42 (codon: CGC) to glutamine (codon: CAG) or glutamic acid (codon: GAG).…”

Methods Applied to the Study of Protein Arginine Methylation

“…A major interest currently is developing methyl-specific antibodies against motifs that are found on histone tails and that form part of the histone code (Di Lorenzo & Bedford, 2011). There are high-quality methyl-specific antibodies, which can be used for ChIP and ChIP-seq experiments, now available for many of the methylarginine motifs that are found on histone tails (Migliori et al, 2012;Waldmann et al, 2011;Wilczek et al, 2011).…”