Blimp1 associates with Prmt5 and directs histone arginine methylation in mouse germ cells.
ABSTRACT Blimp1, a transcriptional repressor, has a crucial role in the specification of primordial germ cells (PGCs) in mice at embryonic day 7.5 (E7.5). This SET-PR domain protein can form complexes with various chromatin modifiers in a context-dependent manner. Here, we show that Blimp1 has a novel interaction with Prmt5, an arginine-specific histone methyltransferase, which mediates symmetrical dimethylation of arginine 3 on histone H2A and/or H4 tails (H2A/H4R3me2s). Prmt5 has been shown to associate with Tudor, a component of germ plasm in Drosophila melanogaster. Blimp1-Prmt5 colocalization results in high levels of H2A/H4 R3 methylation in PGCs at E8.5. However, at E11.5, Blimp1-Prmt5 translocates from the nucleus to the cytoplasm, resulting in the loss of H2A/H4 R3 methylation at the time of extensive epigenetic reprogramming of germ cells. Subsequently, Dhx38, a putative target of the Blimp1-Prmt5 complex, is upregulated. Interestingly, expression of Dhx38 is also seen in pluripotent embryonic germ cells that are derived from PGCs when Blimp1 expression is lost. Our study demonstrates that Blimp1 is involved in a novel transcriptional regulatory complex in the mouse germ-cell lineage.
SourceAvailable from: Natalya Nikitina[Show abstract] [Hide abstract]
ABSTRACT: Prdm1 is a global repressor of transcription that plays multiple important roles during embryonic development, including neural crest specification. Prdm1 acts by repressing large sets of genes via sequence-specific recruitment of co-repressors, many of which are epigenetic modifiers. It is not known whether Prdm1 is expressed during neural crest development in chick embryo. Moreover, the mechanism of Prdm1 action or the nature of possible binding partners that mediate its effects in the neural crest had not yet been addressed. Prdm1 binding partners are known to play important roles during embryonic development, yet in many cases no spatiotemporal expression analysis during early vertebrate development has been performed. In this paper we report the expression patterns of Prdm1 and seven of its known or putative binding partners (Hdac1 and 2, Tle1 and 3, G9a, Prmt5, Lsd1) during early stages of chicken embryogenesis. Prdm1 is expressed in the neural plate border and premigratory neural crest during chick development. Six Prdm1 binding partners (except Tle1) are co-expressed with Prdm1 in the prospective neural plate border at HH4-HH6, and all seven show strong and specific expression in the neural plate border at HH7-HH8, suggesting all of them may cooperate with Prdm1 during neural crest development in chick embryos. Copyright © 2014. Published by Elsevier B.V.Gene Expression Patterns 02/2015; DOI:10.1016/j.gep.2014.12.003 · 1.36 Impact Factor
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ABSTRACT: The protein arginine methyltransferase PRMT5 is complexed with the WD-repeat protein MEP50 (also known as Wdr77 or androgen coactivator p44) in vertebrates in a tetramer of heterodimers. MEP50 is hypothesized to be required for protein substrate recruitment to the catalytic domain of PRMT5. Here we demonstrate that the cross-dimer MEP50 is paired with its cognate PRMT5 molecule to promote histone methylation. We employed qualitative methylation assays and a novel ultrasensitive continuous assay to measure enzyme kinetics. We demonstrate that neither full-length human PRMT5 nor the Xenopus laevis PRMT5 catalytic domain have appreciable protein methyltransferase activity. We show that histones H4 and H3 bind PRMT5-MEP50 more efficiently compared to histone H2A(1-20) and H4(1-20) peptides. Histone binding is mediated through histone fold interactions as determined by competition experiments and by high density histone peptide array interaction studies. Nucleosomes are not a substrate for PRMT5-MEP50, consistent with the primary mode of interaction via the histone fold of H3/H4, obscured by DNA in the nucleosome. Mutation of a conserved arginine (R42) on the MEP50 insertion loop impaired the PRMT5-MEP50 enzymatic efficiency by increasing its histone substrate Km, comparable to that of Caenorhabditis elegans PRMT5. We show that PRMT5-MEP50 prefers unmethylated substrates, consistent with a distributive model for dimethylation and suggesting discrete biological roles for mono- and dimethylarginine modified proteins. We propose a model in which MEP50 and PRMT5 simultaneously engage the protein substrate, orienting its targeted arginine to the catalytic site. Copyright © 2015, The American Society for Biochemistry and Molecular Biology.
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ABSTRACT: Class switch DNA recombination (CSR) is central to the maturation of the Ab response because it diversifies Ab effector functions. Like somatic hypermutation, CSR requires activation-induced cytidine deaminase (AID), whose expression is restricted to B cells, as induced by CD40 engagement or dual TLR-BCR engagement (primary CSR-inducing stimuli). By constructing conditional knockout Igh(+/C)γ(1-cre)Rab7(fl/fl) mice, we identified a B cell-intrinsic role for Rab7, a small GTPase involved in intracellular membrane functions, in mediating AID induction and CSR. Igh(+/C)γ(1-cre)Rab7(fl/fl) mice displayed normal B and T cell development and were deficient in Rab7 only in B cells undergoing Igh(C)γ(1-cre) Iγ1-Sγ1-Cγ1-cre transcription, as induced-like Igh germline Iγ1-Sγ1-Cγ1 and Iε-Sε-Cε transcription-by IL-4 in conjunction with a primary CSR-inducing stimulus. These mice could not mount T-independent or T-dependent class-switched IgG1 or IgE responses while maintaining normal IgM levels. Igh(+/C)γ(1-cre)Rab7(fl/fl) B cells showed, in vivo and in vitro, normal proliferation and survival, normal Blimp-1 expression and plasma cell differentiation, as well as intact activation of the noncanonical NF-κB, p38 kinase, and ERK1/2 kinase pathways. They, however, were defective in AID expression and CSR in vivo and in vitro, as induced by CD40 engagement or dual TLR1/2-, TLR4-, TLR7-, or TLR9-BCR engagement. In Igh(+/C)γ(1-cre)Rab7(fl/fl) B cells, CSR was rescued by enforced AID expression. These findings, together with our demonstration that Rab7-mediated canonical NF-κB activation, as critical to AID induction, outline a novel role of Rab7 in signaling pathways that lead to AID expression and CSR, likely by promoting assembly of signaling complexes along intracellular membranes. Copyright © 2015 by The American Association of Immunologists, Inc.