Post-translational modification (PTM) of proteins is vital for increasing proteome diversity and maintaining cellular homeostasis. If the writing, reading, and removal of modifications are not ...controlled, cancer can develop. Arginine methylation is an understudied modification that is increasingly associated with cancer progression. Consequently protein arginine methyltransferases (PRMTs), the writers of arginine methylation, have rapidly gained interest as novel drug targets. However, for clinical success a deep mechanistic understanding of the biology of PRMTs is required. In this review we focus on advances made regarding the role of PRMTs in stem cell biology, epigenetics, splicing, immune surveillance and the DNA damage response, and highlight the rapid rise of specific inhibitors that are now in clinical trials for cancer therapy.
Arginine methylation as a PTM has gained considerable interest since the recent discovery that solid and haematological cancers display elevated expression of PRMTs, which correlates with poor patient prognosis.Several new findings have cemented arginine methylation as a key regulator of processes hijacked by the cancer cell to ensure survival, including epigenetic-mediated gene expression, mRNA splicing, and the DNA damage response.Growing appreciation of the important role of PRMT5 in cancer stem cell function provides an exciting therapeutic prospect.The development of specific PRMT inhibitors has proceeded at an unprecedented pace, resulting in three major pharmaceutical companies entering their own PRMT inhibitors into Phase I trials. Drug targeting of arginine methylation is becoming a real clinical prospect.
Breast cancer progression, treatment resistance, and relapse are thought to originate from a small population of tumor cells, breast cancer stem cells (BCSCs). Identification of factors critical for ...BCSC function is therefore vital for the development of therapies. Here, we identify the arginine methyltransferase PRMT5 as a key in vitro and in vivo regulator of BCSC proliferation and self-renewal and establish FOXP1, a winged helix/forkhead transcription factor, as a critical effector of PRMT5-induced BCSC function. Mechanistically, PRMT5 recruitment to the FOXP1 promoter facilitates H3R2me2s, SET1 recruitment, H3K4me3, and gene expression. Our findings are clinically significant, as PRMT5 depletion within established tumor xenografts or treatment of patient-derived BCSCs with a pre-clinical PRMT5 inhibitor substantially reduces BCSC numbers. Together, our findings highlight the importance of PRMT5 in BCSC maintenance and suggest that small-molecule inhibitors of PRMT5 or downstream targets could be an effective strategy eliminating this cancer-causing population.
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•PRMT5 expression is elevated in breast cancer stem cells (BCSCs)•PRMT5 is required for BCSC function and self-renewal in vitro and in vivo•Targeting PRMT5 in an established tumor reduces stem cell numbers and tumor growth•FOXP1 promoter is methylated by PRMT5 and is an effector of PRMT5-driven BCSC function
Chiang et al. show that the arginine methyltransferase PRMT5 contributes to breast cancer stem cell function, in part through histone methylation regulating FOXP1 expression. Targeting of PRMT5 through depletion or inhibition reduces stem cell frequency in vitro and in vivo, implicating PRMT5 as important in breast cancer pathogenesis.
Protein post-translation modification plays an important role in regulating DNA repair; however, the role of arginine methylation in this process is poorly understood. Here we identify the arginine ...methyltransferase PRMT5 as a key regulator of homologous recombination (HR)-mediated double-strand break (DSB) repair, which is mediated through its ability to methylate RUVBL1, a cofactor of the TIP60 complex. We show that PRMT5 targets RUVBL1 for methylation at position R205, which facilitates TIP60-dependent mobilization of 53BP1 from DNA breaks, promoting HR. Mechanistically, we demonstrate that PRMT5-directed methylation of RUVBL1 is critically required for the acetyltransferase activity of TIP60, promoting histone H4K16 acetylation, which facilities 53BP1 displacement from DSBs. Interestingly, RUVBL1 methylation did not affect the ability of TIP60 to facilitate ATM activation. Taken together, our findings reveal the importance of PRMT5-mediated arginine methylation during DSB repair pathway choice through its ability to regulate acetylation-dependent control of 53BP1 localization.
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•PRMT5 is a regulator of homologous recombination-mediated double-strand break repair•PRMT5 methylates RUVBL1 at R205, regulating TIP60-mediated histone acetylation•Loss of RUVBL1 methylation leads to 53BP1 retention at break ends•Arginine methylation crosstalks with histone acetylation to regulate repair pathway choice
Clarke et al. show that methylation of RUVBL1 by the arginine methyltransferase PRMT5 is required for homologous recombination-mediated double-strand break repair by promoting TIP60-mediated histone H4K16 acetylation. Loss of PRMT5 activity and defective RUVBL1 methylation leads to 53BP1 retention, increased sensitivity to DNA damaging agents, and genome instability.
First-line cancer treatments successfully eradicate the differentiated tumour mass but are comparatively ineffective against cancer stem cells (CSCs), a self-renewing subpopulation thought to be ...responsible for tumour initiation, metastasis, heterogeneity, and recurrence. CSCs are thus presented as the principal target for elimination during cancer treatment. However, CSCs are challenging to drug target because of numerous intrinsic and extrinsic mechanisms of drug resistance. One such mechanism that remains relatively understudied is the DNA damage response (DDR). CSCs are presumed to possess properties that enable enhanced DNA repair efficiency relative to their highly proliferative bulk progeny, facilitating improved repair of double-strand breaks induced by radiotherapy and most chemotherapeutics. This can occur through multiple mechanisms, including increased expression and splicing fidelity of DNA repair genes, robust activation of cell cycle checkpoints, and elevated homologous recombination-mediated DNA repair. Herein, we summarise the current knowledge concerning improved genome integrity in non-transformed stem cells and CSCs, discuss therapeutic opportunities within the DDR for re-sensitising CSCs to genotoxic stressors, and consider the challenges posed regarding unbiased identification of novel DDR-directed strategies in CSCs. A better understanding of the DDR mediating chemo/radioresistance mechanisms in CSCs could lead to novel therapeutic approaches, thereby enhancing treatment efficacy in cancer patients.
Chronic inflammation is a hallmark of many cancers, yet the pathogenic mechanisms that distinguish cancer-associated inflammation from benign persistent inflammation are still mainly unclear. Here, ...we report that the protein kinase ERK5 controls the expression of a specific subset of inflammatory mediators in the mouse epidermis, which triggers the recruitment of inflammatory cells needed to support skin carcinogenesis. Accordingly, inactivation of ERK5 in keratinocytes prevents inflammation-driven tumorigenesis in this model. In addition, we found that anti-ERK5 therapy cooperates synergistically with existing antimitotic regimens, enabling efficacy of subtherapeutic doses. Collectively, our findings identified ERK5 as a mediator of cancer-associated inflammation in the setting of epidermal carcinogenesis. Considering that ERK5 is expressed in almost all tumor types, our findings suggest that targeting tumor-associated inflammation via anti-ERK5 therapy may have broad implications for the treatment of human tumors.
Cross-talk between distinct protein post-translational modifications is critical for an effective DNA damage response. Arginine methylation plays an important role in maintaining genome stability, ...but how this modification integrates with other enzymatic activities is largely unknown. Here, we identify the deubiquitylating enzyme USP11 as a previously uncharacterised PRMT1 substrate, and demonstrate that the methylation of USP11 promotes DNA end-resection and the repair of DNA double strand breaks (DSB) by homologous recombination (HR), an event that is independent from another USP11-HR activity, the deubiquitylation of PALB2. We also show that PRMT1 is a ubiquitylated protein that it is targeted for deubiquitylation by USP11, which regulates the ability of PRMT1 to bind to and methylate MRE11. Taken together, our findings reveal a specific role for USP11 during the early stages of DSB repair, which is mediated through its ability to regulate the activity of the PRMT1-MRE11 pathway.
•New methods for isolating protein–protein and nucleic acid–protein interactions.•Novel techniques for improving stringency and specificity in interactomics.•An overview of online tools for analysing ...interactomics data.
The technological revolution in high-throughput nucleic acid and protein analysis in the last 15years has launched the field of ‘omics’ and led to great advances in our understanding of cell biology. Consequently the study of the cellular proteome and protein dynamics, in particular interactomics, has been a matter of intense investigation, specifically the determination and description of complex protein interaction networks in the cell, not only with other proteins but also with RNA and DNA. The analysis of these interactions, beginning with their identification and ultimately resulting in structural level examination, is one of the cornerstones of modern biological science underpinning basic research and impacting on applied biology, biomedicine and drug discovery. In this review we summarise a selection of emerging and established techniques currently being applied in this field with a particular focus on affinity-based purification systems and their optimisation, including tandem affinity purification (TAP) tagging, isolation of proteins on nascent DNA (IPOND) and RNA–protein immunoprecipitation in tandem (RIPiT). The recent application of quantitative proteomics to improve stringency and specificity is also discussed, including the use of metabolic labelling by stable isotope labelling by amino acids in cell culture (SILAC), localization of organelle proteins by isotope tagging (LOPIT) and proximity-dependent biotin identification (BioID). Finally, we describe a range of software resources that can be applied to interactomics, both to handle raw data and also to scrutinise its broader biological context. In this section we focus especially on open-access online interactomic databases such as Reactome and IntAct.
Attenuating the function of protein arginine methyltransferases (PRMTs) is an objective for the investigation and treatment of several diseases including cardiovascular disease and cancer. ...Bisubstrate inhibitors that simultaneously target binding sites for arginine substrate and the cofactor (S-adenosylmethionine (SAM)) have potential utility, but structural information on their binding is required for their development. Evaluation of bisubstrate inhibitors featuring an isosteric guanidine replacement with two prominent enzymes PRMT1 and CARM1 (PRMT4) by isothermal titration calorimetry (ITC), activity assays and crystallography are reported. Key findings are that 2-aminopyridine is a viable replacement for guanidine, providing an inhibitor that binds more strongly to CARM1 than PRMT1. Moreover, a residue around the active site that differs between CARM1 (Asn-265) and PRMT1 (Tyr-160) is identified that affects the side chain conformation of the catalytically important neighbouring glutamate in the crystal structures. Mutagenesis data supports its contribution to the difference in binding observed for this inhibitor. Structures of CARM1 in complex with a range of seven inhibitors reveal the binding modes and show that inhibitors with an amino acid terminus adopt a single conformation whereas the electron density for equivalent amine-bearing inhibitors is consistent with preferential binding in two conformations. These findings inform the molecular basis of CARM1 ligand binding and identify differences between CARM1 and PRMT1 that can inform drug discovery efforts.
The arginine methyltransferase PRMT5 has been increasingly associated with cancer development. Here we describe our recent findings that PRMT5 is a critical regulator of breast cancer stem cell ...survival via the epigenetic regulation of FOXP1. Consequently, PRMT5 inhibitors could potentially eradicate cancer stem cells thereby preventing tumour relapse.
c‐Jun, the major component of the AP‐1 transcription factor complex, has important functions in cellular proliferation and oncogenic transformation. The RING domain‐containing protein RACO‐1 ...functions as a c‐Jun coactivator that molecularly links growth factor signalling to AP‐1 transactivation. Here we demonstrate that RACO‐1 is present as a nuclear dimer and that c‐Jun specifically interacts with dimeric RACO‐1. Moreover, RACO‐1 is identified as a substrate of the arginine methyltransferase PRMT1, which methylates RACO‐1 on two arginine residues. Arginine methylation of RACO‐1 promotes a conformational change that stabilises RACO‐1 by facilitating K63‐linked ubiquitin chain formation, and enables RACO‐1 dimerisation and c‐Jun interaction. Abrogation of PRMT1 function impairs AP‐1 activity and results in decreased expression of a large percentage of c‐Jun target genes. These results demonstrate that arginine methylation of RACO‐1 is required for efficient transcriptional activation by c‐Jun/AP‐1 and thus identify PRMT1 as an important regulator of c‐Jun/AP‐1 function.
Protein arginine methylation of the recently identified c‐Jun coactivator protein RACO‐1 establishes PRMT1 as new regulator of c‐Jun/AP‐1 function.
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