Abstract: Arginine methylation is a common post-translational modification functioning as an epigenetic regulator of transcription and playing key roles in pre-mRNA splicing, DNA damage signaling, mRNA translation, cell signaling, and cell fate decision. Recently, a wealth of studies using transgenic mouse models and selective PRMT inhibitors helped define physiological roles for protein arginine methyltransferases (PRMTs) linking them to diseases such as cancer and metabolic, neurodegenerative, and muscular disorders. This review describes the recent molecular advances that have been uncovered in normal and diseased mammalian cells. Arginine methylation is a common post-translational modification functioning as an epigenetic regulator of transcription and playing key roles in pre-mRNA splicing, DNA damage signaling, mRNA translation, cell signaling, and cell fate decision. Recently, a wealth of studies using transgenic mouse models and selective PRMT inhibitors helped define physiological roles for protein arginine methyltransferases (PRMTs) linking them to diseases such as cancer and metabolic, neurodegenerative, and muscular disorders. This review describes the recent molecular advances that have been uncovered in normal and diseased mammalian cells. Post-translational modifications (PTMs) lie at the heart of the fields of epigenetics and signal transduction. PTMs are involved in nearly all signaling cascades, often initiating or amplifying dynamic signals, which can be finely tuned to accommodate cellular requirements. The post-translation modification of histones as well as DNA methylation are key events of epigenetics influencing gene expression. Therefore, the enzymes that deposit and remove PTMs and DNA methylation are prime drug targets to alter specific pathways or gene expression and to treat certain diseases, including cancer. Arginine methylation is gaining traction as a key PTM, largely due to the generation of antibodies capable of detecting methylarginines, advanced proteomic techniques, small molecule inhibitors of protein arginine methyltransferases (PRMTs), and new transgenic animals to model human disease. With the interesting possibility that arginine demethylases exist, further roles of arginine methylation will soon be uncovered. The goal of this review is to describe the recent molecular advances that are regulated by arginine methylation. The nitrogen atoms of arginine within polypeptides can be modified to contain methyl groups, a process termed arginine methylation. In mammals, arginine methylation is a modification as common as phosphorylation and ubiquitination (Larsen et al., 2016Larsen S.C. Sylvestersen K.B. Mund A. Lyon D. Mullari M. Madsen M.V. Daniel J.A. Jensen L.J. Nielsen M.L. Proteome-wide analysis of arginine monomethylation reveals widespread occurrence in human cells.Sci. Signal. 2016; 9: rs9Crossref PubMed Scopus (157) Google Scholar). It is carried out by the nine members of the PRMT family (Bedford and Clarke, 2009Bedford M.T. Clarke S.G. Protein arginine methylation in mammals: who, what, and why.Mol. Cell. 2009; 33: 1-13Abstract Full Text Full Text PDF PubMed Scopus (716) Google Scholar), although others may exist, such as the putative arginine methyltransferase NDUFAF7 (Zurita Rendón et al., 2014Zurita Rendón O. Silva Neiva L. Sasarman F. Shoubridge E.A. The arginine methyltransferase NDUFAF7 is essential for complex I assembly and early vertebrate embryogenesis.Hum. Mol. Genet. 2014; 23: 5159-5170Crossref PubMed Google Scholar). PRMTs catalyze the transfer of a methyl group from S-adenosylmethionine (SAM) to the guanidino nitrogen atoms of arginine. This reaction results in the formation of methylarginine and S-adenosylhomocysteine. There are three main forms of methylarginines identified in eukaryotes: ω-NG-monomethylarginine (MMA), ω-NG,NG-asymmetric dimethylarginine (aDMA), and ω-NG,N’G-symmetric dimethylarginine (sDMA) (Figure 1). PRMTs fall into three categories according to their catalytic activity; type I (PRMT1, PRMT2, PRMT3, PRMT4, PRMT6, and PRMT8) and type II (PRMT5 and PRMT9) enzymes carry out the formation of MMA as an intermediate before the establishment of aDMA or sDMA, respectively (Yang et al., 2015Yang Y. Hadjikyriacou A. Xia Z. Gayatri S. Kim D. Zurita-Lopez C. Kelly R. Guo A. Li W. Clarke S.G. Bedford M.T. PRMT9 is a type II methyltransferase that methylates the splicing factor SAP145.Nat. Commun. 2015; 6: 6428Crossref PubMed Scopus (53) Google Scholar). PRMT7 is a type III enzyme that catalyzes only the formation of MMA, and thus far, histones are its only known substrates (Feng et al., 2013Feng Y. Maity R. Whitelegge J.P. Hadjikyriacou A. Li Z. Zurita-Lopez C. Al-Hadid Q. Clark A.T. Bedford M.T. Masson J.Y. Clarke S.G. Mammalian protein arginine methyltransferase 7 (PRMT7) specifically targets RXR sites in lysine- and arginine-rich regions.J. Biol. Chem. 2013; 288: 37010-37025Crossref PubMed Scopus (43) Google Scholar). Arginine methylation is known to play a major role in gene regulation because of the ability of the PRMTs to deposit key activating (histone H4R3me2a, H3R2me2s, H3R17me2a, H3R26me2a) or repressive (H3R2me2a, H3R8me2a, H3R8me2s, H4R3me2s) histone marks. In addition, there are many substrates that are non-histones involved in biological processes including transcription, cell signaling, mRNA translation, DNA damage signaling, receptor trafficking, protein stability, and pre-mRNA splicing (Auclair and Richard, 2013Auclair Y. Richard S. The role of arginine methylation in the DNA damage response.DNA Repair (Amst.). 2013; 12: 459-465Crossref PubMed Scopus (15) Google Scholar, Yang and Bedford, 2013Yang Y. Bedford M.T. Protein arginine methyltransferases and cancer.Nat. Rev. Cancer. 2013; 13: 37-50Crossref PubMed Scopus (251) Google Scholar). Arginine- and glycine-rich motifs, termed RGG/RG motifs, are among the most common amino acid sequences favored by PRMTs, and these motifs often function in both nucleic acid binding and mediating protein-protein interactions (Thandapani et al., 2013Thandapani P. O’Connor T.R. Bailey T.L. Richard S. Defining the RGG/RG motif.Mol. Cell. 2013; 50: 613-623Abstract Full Text Full Text PDF PubMed Scopus (116) Google Scholar). The presence of glycine neighboring arginine is predicted to enhance conformational flexibility and facilitate the access of the arginine to the active site of PRMTs. Not all PRMTs methylate RGG/RG motifs. CARM1/PRMT4 prefers arginine neighboring a PGM-rich (proline, glycine, and methionine) motif (Yang and Bedford, 2013Yang Y. Bedford M.T. Protein arginine methyltransferases and cancer.Nat. Rev. Cancer. 2013; 13: 37-50Crossref PubMed Scopus (251) Google Scholar), while PRMT7 prefers RxR motifs surrounded by a lysine-rich environment (Feng et al., 2013Feng Y. Maity R. Whitelegge J.P. Hadjikyriacou A. Li Z. Zurita-Lopez C. Al-Hadid Q. Clark A.T. Bedford M.T. Masson J.Y. Clarke S.G. Mammalian protein arginine methyltransferase 7 (PRMT7) specifically targets RXR sites in lysine- and arginine-rich regions.J. Biol. Chem. 2013; 288: 37010-37025Crossref PubMed Scopus (43) Google Scholar). The main consequence of arginine methylation is alteration of its binding interactions. Given the nature of arginine with its five potential hydrogen-bond donors, the steric effects from adding a methyl group will affect interactions with the hydrogen-bond acceptors of their interacting partners without changing the charge (Fuhrmann et al., 2015Fuhrmann J. Clancy K.W. Thompson P.R. Chemical biology of protein arginine modifications in epigenetic regulation.Chem. Rev. 2015; 115: 5413-5461Crossref PubMed Scopus (0) Google Scholar). The methylation of arginines enhances interactions with Tudor domains (Gayatri and Bedford, 2014Gayatri S. Bedford M.T. Readers of histone methylarginine marks.Biochim. Biophys. Acta. 2014; 1839: 702-710Crossref PubMed Scopus (0) Google Scholar). The Tudor domains of SMN (Survival of motor neuron), SPF30 (Splicing factor 30), and TDRD1/2/3/6/9/11 (Tudor domain-containing protein) are currently the main known methylarginine-interacting domains (Tripsianes et al., 2011Tripsianes K. Madl T. Machyna M. Fessas D. Englbrecht C. Fischer U. Neugebauer K.M. Sattler M. Structural basis for dimethylarginine recognition by the Tudor domains of human SMN and SPF30 proteins.Nat. Struct. Mol. Biol. 2011; 18: 1414-1420Crossref PubMed Scopus (0) Google Scholar). Tudor domains form an aromatic cage such that cation-π contacts occur between their aromatic residues and the cationic carbon of methylarginine. The methyl groups provide increased hydrophobicity within the Tudor domain binding pocket. There are ∼36 proteins that harbor at least one Tudor domain in humans compared to >2,000 proteins containing RGG/RG motifs, suggesting that other methylarginine-interacting domains are likely to be discovered. Tudor domains as methylarginine interactors were recently reviewed (Gayatri and Bedford, 2014Gayatri S. Bedford M.T. Readers of histone methylarginine marks.Biochim. Biophys. Acta. 2014; 1839: 702-710Crossref PubMed Scopus (0) Google Scholar). The existence of enzymes capable of demethylating methylarginines is central to the concept that arginine methylation is a dynamic modification. The existence of arginine demethylases is controversial (Yang and Bedford, 2013Yang Y. Bedford M.T. Protein arginine methyltransferases and cancer.Nat. Rev. Cancer. 2013; 13: 37-50Crossref PubMed Scopus (251) Google Scholar). A putative arginine demethylase, JmjD6, was identified but later shown to be a lysine hydroxylase (Webby et al., 2009Webby C.J. Wolf A. Gromak N. Dreger M. Kramer H. Kessler B. Nielsen M.L. Schmitz C. Butler D.S. Yates 3rd, J.R. et al.Jmjd6 catalyses lysyl-hydroxylation of U2AF65, a protein associated with RNA splicing.Science. 2009; 325: 90-93Crossref PubMed Scopus (192) Google Scholar). Recently, it was shown that certain lysine demethylases (KDM3A, KDM4E, KDM5C) also possess arginine demethylation activity in vitro (Walport et al., 2016Walport L.J. Hopkinson R.J. Chowdhury R. Schiller R. Ge W. Kawamura A. Schofield C.J. Arginine demethylation is catalysed by a subset of JmjC histone lysine demethylases.Nat. Commun. 2016; 7: 11974Crossref PubMed Google Scholar). Since the enzymatic mode of action for arginine demethylation is similar to lysine demethylation, it seems likely that other JmjC proteins will also demethylate methylarginine. Further investigation is required to confirm the function of these dual lysine/arginine demethylases in vivo and to explore how these two different activities interplay. The PRMT structures reveal the presence of seven β strand catalytic domains with variations in their structure that make them unique. Type I and II enzymes have a central cavity and two opposing active sites in their head-to-tail homodimer structure (Zhang and Cheng, 2003Zhang X. Cheng X. Structure of the predominant protein arginine methyltransferase PRMT1 and analysis of its binding to substrate peptides.Structure. 2003; 11: 509-520Abstract Full Text Full Text PDF PubMed Scopus (217) Google Scholar). These active sites contain a highly conserved SAM binding pocket characterized by the presence of an E-loop critical for substrate recognition and methylation. PRMT5 is unique in that it requires the methylosome protein 50 (MEP50/WDR77) to form an active enzymatic complex, and this interaction defines its distributive mode of action (Antonysamy et al., 2012Antonysamy S. Bonday Z. Campbell R.M. Doyle B. Druzina Z. Gheyi T. Han B. Jungheim L.N. Qian Y. Rauch C. et al.Crystal structure of the human PRMT5:MEP50 complex.Proc. Natl. Acad. Sci. USA. 2012; 109: 17960-17965Crossref PubMed Scopus (96) Google Scholar). Type III enzyme PRMT7 is an exception, as it lacks the central cavity and acts as a monomer that acquires a homodimer-like structure with two catalytic domains both essential for its activity. PRMT7 also requires the E-loop, which is critical for preserving its unique type III activity (Debler et al., 2016Debler E.W. Jain K. Warmack R.A. Feng Y. Clarke S.G. Blobel G. Stavropoulos P. A glutamate/aspartate switch controls product specificity in a protein arginine methyltransferase.Proc. Natl. Acad. Sci. USA. 2016; 113: 2068-2073Crossref PubMed Google Scholar, Jain et al., 2016Jain K. Warmack R.A. Debler E.W. Hadjikyriacou A. Stavropoulos P. Clarke S.G. Protein Arginine Methyltransferase Product Specificity Is Mediated by Distinct Active-site Architectures.J. Biol. Chem. 2016; 291: 18299-18308Crossref PubMed Scopus (0) Google Scholar). PRMT1 is the main enzyme responsible for the generation of aDMA in proteins, and it has a preference for RGG/RG motifs (Thandapani et al., 2013Thandapani P. O’Connor T.R. Bailey T.L. Richard S. Defining the RGG/RG motif.Mol. Cell. 2013; 50: 613-623Abstract Full Text Full Text PDF PubMed Scopus (116) Google Scholar); however, there are numerous exceptions. PRMT1 functions as a transcriptional co-activator by depositing dimethylarginines on H4R3 (Bedford and Clarke, 2009Bedford M.T. Clarke S.G. Protein arginine methylation in mammals: who, what, and why.Mol. Cell. 2009; 33: 1-13Abstract Full Text Full Text PDF PubMed Scopus (716) Google Scholar). PRMT1 also methylates RNA binding and DNA damage proteins to modulate RNA metabolism and maintain genome stability, respectively (Auclair and Richard, 2013Auclair Y. Richard S. The role of arginine methylation in the DNA damage response.DNA Repair (Amst.). 2013; 12: 459-465Crossref PubMed Scopus (15) Google Scholar). PRMT1 was shown to homodimerize (Zhang and Cheng, 2003Zhang X. Cheng X. Structure of the predominant protein arginine methyltransferase PRMT1 and analysis of its binding to substrate peptides.Structure. 2003; 11: 509-520Abstract Full Text Full Text PDF PubMed Scopus (217) Google Scholar), and its activity to be highly dependent on, two methionine residues, M48 and M155, which alter substrate specificity and abolish catalytic activity if mutated (Gui et al., 2014Gui S. Gathiaka S. Li J. Qu J. Acevedo O. Hevel J.M. A remodeled protein arginine methyltransferase 1 (PRMT1) generates symmetric dimethylarginine.J. Biol. Chem. 2014; 289: 9320-9327Crossref PubMed Scopus (10) Google Scholar). Interestingly, loss of PRMT1 activity increases MMA- and sDMA-proteins owing to substrate scavenging by other PRMTs (Dhar et al., 2013Dhar S. Vemulapalli V. Patananan A.N. Huang G.L. Di Lorenzo A. Richard S. Comb M.J. Guo A. Clarke S.G. Bedford M.T. Loss of the major Type I arginine methyltransferase PRMT1 causes substrate scavenging by other PRMTs.Sci. Rep. 2013; 3: 1311Crossref PubMed Scopus (0) Google Scholar). PRMT1 is regulated by alternative splicing resulting in seven different isoforms, suggesting that each isoform with its distinctive N terminus has unique substrate preference (Goulet et al., 2007Goulet I. Gauvin G. Boisvenue S. Côté J. Alternative splicing yields protein arginine methyltransferase 1 isoforms with distinct activity, substrate specificity, and subcellular localization.J. Biol. Chem. 2007; 282: 33009-33021Crossref PubMed Scopus (92) Google Scholar). PRMT1 is regulated by oxidation (Morales et al., 2015Morales Y. Nitzel D.V. Price O.M. Gui S. Li J. Qu J. Hevel J.M. Redox Control of Protein Arginine Methyltransferase 1 (PRMT1) Activity.J. Biol. Chem. 2015; 290: 14915-14926Crossref PubMed Scopus (5) Google Scholar) and microRNAs including miR503 (Li et al., 2015aLi B. Liu L. Li X. Wu L. miR-503 suppresses metastasis of hepatocellular carcinoma cell by targeting PRMT1.Biochem. Biophys. Res. Commun. 2015; 464: 982-987Crossref PubMed Scopus (7) Google Scholar). PRMT2 has been shown to function as a transcriptional repressor (Ganesh et al., 2006Ganesh L. Yoshimoto T. Moorthy N.C. Akahata W. Boehm M. Nabel E.G. Nabel G.J. Protein methyltransferase 2 inhibits NF-kappaB function and promotes apoptosis.Mol. Cell. Biol. 2006; 26: 3864-3874Crossref PubMed Scopus (0) Google Scholar). Notably, PRMT2 expression increases in hypoxic conditions (Yildirim et al., 2006Yildirim A.O. Bulau P. Zakrzewicz D. Kitowska K.E. Weissmann N. Grimminger F. Morty R.E. Eickelberg O. Increased protein arginine methylation in chronic hypoxia: role of protein arginine methyltransferases.Am. J. Respir. Cell Mol. Biol. 2006; 35: 436-443Crossref PubMed Scopus (0) Google Scholar) and decreases in elevated glucose conditions. PRMT3 is unique among PRMTs for its zinc-finger at its N terminus, which confers substrate specificity (Frankel and Clarke, 2000Frankel A. Clarke S. PRMT3 is a distinct member of the protein arginine N-methyltransferase family. Conferral of substrate specificity by a zinc-finger domain.J. Biol. Chem. 2000; 275: 32974-32982Crossref PubMed Scopus (0) Google Scholar). It methylates ribosomal proteins (Swiercz et al., 2007Swiercz R. Cheng D. Kim D. Bedford M.T. Ribosomal protein rpS2 is hypomethylated in PRMT3-deficient mice.J. Biol. Chem. 2007; 282: 16917-16923Crossref PubMed Scopus (0) Google Scholar), and its activity is regulated by interacting proteins such as DAL-1/4.1B (Singh et al., 2004Singh V. Miranda T.B. Jiang W. Frankel A. Roemer M.E. Robb V.A. Gutmann D.H. Herschman H.R. Clarke S. Newsham I.F. DAL-1/4.1B tumor suppressor interacts with protein arginine N-methyltransferase 3 (PRMT3) and inhibits its ability to methylate substrates in vitro and in vivo.Oncogene. 2004; 23: 7761-7771Crossref PubMed Scopus (0) Google Scholar). CARM1/PRMT4 is known for its transcription coactivator function. It methylates H3R17 and H3R26 and also potentiates transcription by methylating and recruiting transcription factors directly. CARM1 activity is regulated by association with CBP/p300 acetyltransferase and UPF1. CBP-mediated acetylation of histone H3K18, and subsequently at K23, is sufficient to recruit CARM1 to methylate H3R17 (Bedford and Clarke, 2009Bedford M.T. Clarke S.G. Protein arginine methylation in mammals: who, what, and why.Mol. Cell. 2009; 33: 1-13Abstract Full Text Full Text PDF PubMed Scopus (716) Google Scholar). CARM1 also regulates nonsense-mediated mRNA decay by associating with UPF1 and recruiting it to premature terminating codon-containing transcripts (Sanchez et al., 2016Sanchez G. Bondy-Chorney E. Laframboise J. Paris G. Didillon A. Jasmin B.J. Côté J. A novel role for CARM1 in promoting nonsense-mediated mRNA decay: potential implications for spinal muscular atrophy.Nucleic Acids Res. 2016; 44: 2661-2676Crossref PubMed Scopus (1) Google Scholar). CARM1 is involved in regulating pre-mRNA splicing (Cheng et al., 2007Cheng D. Côté J. Shaaban S. Bedford M.T. The arginine methyltransferase CARM1 regulates the coupling of transcription and mRNA processing.Mol. Cell. 2007; 25: 71-83Abstract Full Text Full Text PDF PubMed Scopus (210) Google Scholar, Kuhn et al., 2011Kuhn P. Chumanov R. Wang Y. Ge Y. Burgess R.R. Xu W. Automethylation of CARM1 allows coupling of transcription and mRNA splicing.Nucleic Acids Res. 2011; 39: 2717-2726Crossref PubMed Scopus (29) Google Scholar). Its expression is regulated by various microRNAs including miR-181c (Xu et al., 2013Xu Z. Jiang J. Xu C. Wang Y. Sun L. Guo X. Liu H. MicroRNA-181 regulates CARM1 and histone arginine methylation to promote differentiation of human embryonic stem cells.PLoS ONE. 2013; 8: e53146Crossref PubMed Scopus (0) Google Scholar), miR-223 (Vu et al., 2013Vu L.P. Perna F. Wang L. Voza F. Figueroa M.E. Tempst P. Erdjument-Bromage H. Gao R. Chen S. Paietta E. et al.PRMT4 blocks myeloid differentiation by assembling a methyl-RUNX1-dependent repressor complex.Cell Rep. 2013; 5: 1625-1638Abstract Full Text Full Text PDF PubMed Scopus (21) Google Scholar), and miR-15 (Liu et al., 2014Liu X. Wang L. Li H. Lu X. Hu Y. Yang X. Huang C. Gu D. Coactivator-associated arginine methyltransferase 1 targeted by miR-15a regulates inflammation in acute coronary syndrome.Atherosclerosis. 2014; 233: 349-356Abstract Full Text Full Text PDF PubMed Scopus (0) Google Scholar). CARM1 automethylation and O-linked-β-N-acetylglucosaminidation determines substrate specificity (Charoensuksai et al., 2015Charoensuksai P. Kuhn P. Wang L. Sherer N. Xu W. O-GlcNAcylation of co-activator-associated arginine methyltransferase 1 regulates its protein substrate specificity.Biochem. J. 2015; 466: 587-599Crossref PubMed Scopus (5) Google Scholar). PRMT6 is predominantly nuclear and methylates RGG/RG motif; however, it also methylates arginines neighboring charged residues as observed with HIV Tat (Boulanger et al., 2005Boulanger M.C. Liang C. Russell R.S. Lin R. Bedford M.T. Wainberg M.A. Richard S. Methylation of Tat by PRMT6 regulates human immunodeficiency virus type 1 gene expression.J. Virol. 2005; 79: 124-131Crossref PubMed Scopus (127) Google Scholar). PRMT6-mediated methylation is generally associated with transcriptional repression by generating H3R2me2a (Neault et al., 2012Neault M. Mallette F.A. Vogel G. Michaud-Levesque J. Richard S. Ablation of PRMT6 reveals a role as a negative transcriptional regulator of the p53 tumor suppressor.Nucleic Acids Res. 2012; 40: 9513-9521Crossref PubMed Scopus (38) Google Scholar). PRMT6 automethylation increases its stability (Singhroy et al., 2013Singhroy D.N. Mesplède T. Sabbah A. Quashie P.K. Falgueyret J.P. Wainberg M.A. Automethylation of protein arginine methyltransferase 6 (PRMT6) regulates its stability and its anti-HIV-1 activity.Retrovirology. 2013; 10: 73Crossref PubMed Scopus (13) Google Scholar). Brain-specific PRMT8 is membrane-bound type I arginine methyltransferase. X-ray crystallography shows it forms a tetrameric structure for substrate recognition and specificity (Lee et al., 2005Lee J. Sayegh J. Daniel J. Clarke S. Bedford M.T. PRMT8, a new membrane-bound tissue-specific member of the protein arginine methyltransferase family.J. Biol. Chem. 2005; 280: 32890-32896Crossref PubMed Scopus (145) Google Scholar, Lee et al., 2015Lee W.C. Lin W.L. Matsui T. Chen E.S. Wei T.Y. Lin W.H. Hu H. Zheng Y.G. Tsai M.D. Ho M.C. Protein Arginine Methyltransferase 8: Tetrameric Structure and Protein Substrate Specificity.Biochemistry. 2015; 54: 7514-7523Crossref PubMed Scopus (5) Google Scholar). PRMT8 also has been reported to harbor non-canonical phospholipase D activity (Kim et al., 2015bKim J.D. Park K.E. Ishida J. Kako K. Hamada J. Kani S. Takeuchi M. Namiki K. Fukui H. Fukuhara S. et al.PRMT8 as a phospholipase regulates Purkinje cell dendritic arborization and motor coordination.Sci. Adv. 2015; 1: e1500615Crossref PubMed Google Scholar). In mammals, PRMT5 is the main type II enzyme responsible for the majority of sDMA formation in polypeptides. It methylates in a distributive, rather than processive, manner (Wang et al., 2014bWang M. Fuhrmann J. Thompson P.R. Protein arginine methyltransferase 5 catalyzes substrate dimethylation in a distributive fashion.Biochemistry. 2014; 53: 7884-7892Crossref PubMed Scopus (7) Google Scholar), meaning PRMT5 releases MMA before the second methylation event. In vitro, PRMT5 activity requires the formation of a hetero-octameric complex with MEP50, which mediates substrate specificity and interaction with binding partners (Antonysamy et al., 2012Antonysamy S. Bonday Z. Campbell R.M. Doyle B. Druzina Z. Gheyi T. Han B. Jungheim L.N. Qian Y. Rauch C. et al.Crystal structure of the human PRMT5:MEP50 complex.Proc. Natl. Acad. Sci. USA. 2012; 109: 17960-17965Crossref PubMed Scopus (96) Google Scholar). Although MEP50 is required to activate PRMT5 for substrate recognition in vitro (Wang et al., 2014bWang M. Fuhrmann J. Thompson P.R. Protein arginine methyltransferase 5 catalyzes substrate dimethylation in a distributive fashion.Biochemistry. 2014; 53: 7884-7892Crossref PubMed Scopus (7) Google Scholar), the absolute requirement of MEP50 for PRMT5 function in vivo remains unclear (Tee et al., 2010Tee W.W. Pardo M. Theunissen T.W. Yu L. Choudhary J.S. Hajkova P. Surani M.A. Prmt5 is essential for early mouse development and acts in the cytoplasm to maintain ES cell pluripotency.Genes Dev. 2010; 24: 2772-2777Crossref PubMed Scopus (115) Google Scholar). PRMT5 interaction with various partners, including plCln, and the kinase RioK1, is important to regulate its catalytic activity. These partners bind PRMT5 in a mutually exclusive manner to control substrate specificity (Guderian et al., 2011Guderian G. Peter C. Wiesner J. Sickmann A. Schulze-Osthoff K. Fischer U. Grimmler M. RioK1, a new interactor of protein arginine methyltransferase 5 (PRMT5), competes with pICln for binding and modulates PRMT5 complex composition and substrate specificity.J. Biol. Chem. 2011; 286: 1976-1986Crossref PubMed Scopus (43) Google Scholar). As a cautionary note, anti-FLAG antibodies are known to non-specifically immunopurify PRMT5 (Nishioka and Reinberg, 2003Nishioka K. Reinberg D. Methods and tips for the purification of human histone methyltransferases.Methods. 2003; 31: 49-58Crossref PubMed Scopus (0) Google Scholar). Besides methylating histones and functioning, in general, as a transcriptional corepressor, PRMT5 methylates Sm proteins for their assembly into mature small nuclear ribonucleoproteins (snRNPs) (Meister et al., 2001Meister G. Eggert C. Bühler D. Brahms H. Kambach C. Fischer U. Methylation of Sm proteins by a complex containing PRMT5 and the putative U snRNP assembly factor pICln.Curr. Biol. 2001; 11: 1990-1994Abstract Full Text Full Text PDF PubMed Scopus (193) Google Scholar). Thus, it is not surprising that PRMT5-deficient cells harbor numerous defects in splicing (Bezzi et al., 2013Bezzi M. Teo S.X. Muller J. Mok W.C. Sahu S.K. Vardy L.A. Bonday Z.Q. Guccione E. Regulation of constitutive and alternative splicing by PRMT5 reveals a role for Mdm4 pre-mRNA in sensing defects in the spliceosomal machinery.Genes Dev. 2013; 27: 1903-1916Crossref PubMed Scopus (0) Google Scholar). PRMT9 has recently been shown to be a type II enzyme that is not redundant with PRMT5 (Hadjikyriacou et al., 2015Hadjikyriacou A. Yang Y. Espejo A. Bedford M.T. Clarke S.G. Unique Features of Human Protein Arginine Methyltransferase 9 (PRMT9) and Its Substrate RNA Splicing Factor SF3B2.J. Biol. Chem. 2015; 290: 16723-16743Crossref PubMed Scopus (7) Google Scholar, Yang et al., 2015Yang Y. Hadjikyriacou A. Xia Z. Gayatri S. Kim D. Zurita-Lopez C. Kelly R. Guo A. Li W. Clarke S.G. Bedford M.T. PRMT9 is a type II methyltransferase that methylates the splicing factor SAP145.Nat. Commun. 2015; 6: 6428Crossref PubMed Scopus (53) Google Scholar). PRMT9 methylates spliceosome-associated protein 145 (SAP145) to regulate alternative splicing (Yang et al., 2015Yang Y. Hadjikyriacou A. Xia Z. Gayatri S. Kim D. Zurita-Lopez C. Kelly R. Guo A. Li W. Clarke S.G. Bedford M.T. PRMT9 is a type II methyltransferase that methylates the splicing factor SAP145.Nat. Commun. 2015; 6: 6428Crossref PubMed Scopus (53) Google Scholar). PRMT9, like PRMT7, contains two catalytic domains with conserved sequences in the double E-loop important for substrate specificity and activity (Hadjikyriacou et al., 2015Hadjikyriacou A. Yang Y. Espejo A. Bedford M.T. Clarke S.G. Unique Features of Human Protein Arginine Methyltransferase 9 (PRMT9) and Its Substrate RNA Splicing Factor SF3B2.J. Biol. Chem. 2015; 290: 16723-16743Crossref PubMed Scopus (7) Google Scholar, Jain et al., 2016Jain K. Warmack R.A. Debler E.W. Hadjikyriacou A. Stavropoulos P. Clarke S.G. Protein Arginine Methyltransferase Product Specificity Is Mediated by Distinct Active-site Architectures.J. Biol. Chem. 2016; 291: 18299-18308Crossref PubMed Scopus (0) Google Scholar). Mutations within the catalytic domain can switch PRMT9 activity from a type II to a type III enzyme (Figure 1) (Jain et al., 2016Jain K. Warmack R.A. Debler E.W. Hadjikyriacou A. Stavropoulos P. Clarke S.G. Protein Arginine Methyltransferase Product Specificity Is Mediated by Distinct Active-site Architectures.J. Biol. Chem. 2016; 291: 18299-18308Crossref PubMed Scopus (0) Google Scholar). PRMT7 plays a role in transcriptional regulation, snRNP biogenesis, and splicing. PRMT7 was first identified in a screen where its depletion leads to increase sensitivity to topoisomerase II inhibitors (Gros et al., 2006Gros L. Renodon-Cornière A. de Saint Vincent B.R. Feder M. Bujnicki J.M. Jacquemin-Sablon A. Characterization of prmt7alpha and beta isozymes from Chinese hamster cells sensitive and resistant to topoisomerase II inhibitors.Biochim. Biophys. Acta. 2006; 1760: 1646-1656Crossref PubMed Scopus (0) Google Scholar). Although the mechanism of action of PRMT7 in DNA damage signaling remains to be defined, it was shown that cells depleted of PRMT7 upregulate genes of the DNA repair machinery such as POLD1, POLD2, ALKBH5, and APEX2 (Karkhanis et al., 2012Karkhanis V. Wang L. Tae S. Hu Y.J. Imbalzano A.N. Sif S. Protein arginine methyltransferase 7 regulates cellular response to DNA damage by methylating promoter histones H2A and H4 of the polymerase δ catalytic subunit gene, POLD1.J. Biol. Chem. 2012; 287: 29801-29814Crossref PubMed Scopus (53) Google Scholar). The catalytic activity of PRMT7 is dependent on the highly conserved double E-loop containing two residues critical for substrate preference; mutating these residues switches PRMT7 to a type I or II enzyme (Figure 1) (Debler et al., 2016Debler E.W. Jain K. Warmack R.A. Feng Y. Clarke S.G. Blobel G. Stavropoulos P. A glutamate/aspartate switch controls product specificity in a protein arginine methyltransferase.Proc. Natl. Acad. Sci. USA. 2016; 113: 2068-2073Crossref PubMed Google Scholar, Jain et al., 2016Jain K. Warmack R.A. Debler E.W. Hadjikyriacou A. Stavropoulos P. Clarke S.G. Protein Arginine Methyltransferase Product Specificity Is Mediated by Distinct Active-site Architectures.J. Biol. Chem. 2016; 291: 18299-18308Crossref PubMed Scopus (0) Google Scholar). Presently, the relationship between type III and type I and II enzymes is not understood, and there is no evidence of priming by PRMT7 monomethylation for su