Article

Phosphorylation of MLL by ATR is Required for Execution of Mammalian S Phase Checkpoint

Department of Medicine, Washington University School of Medicine, St Louis, Missouri 63110, USA.
Nature (Impact Factor: 41.46). 09/2010; 467(7313):343-6. DOI: 10.1038/nature09350
Source: PubMed

ABSTRACT

Cell cycle checkpoints are implemented to safeguard the genome, avoiding the accumulation of genetic errors. Checkpoint loss results in genomic instability and contributes to the evolution of cancer. Among G1-, S-, G2- and M-phase checkpoints, genetic studies indicate the role of an intact S-phase checkpoint in maintaining genome integrity. Although the basic framework of the S-phase checkpoint in multicellular organisms has been outlined, the mechanistic details remain to be elucidated. Human chromosome-11 band-q23 translocations disrupting the MLL gene lead to poor prognostic leukaemias. Here we assign MLL as a novel effector in the mammalian S-phase checkpoint network and identify checkpoint dysfunction as an underlying mechanism of MLL leukaemias. MLL is phosphorylated at serine 516 by ATR in response to genotoxic stress in the S phase, which disrupts its interaction with, and hence its degradation by, the SCF(Skp2) E3 ligase, leading to its accumulation. Stabilized MLL protein accumulates on chromatin, methylates histone H3 lysine 4 at late replication origins and inhibits the loading of CDC45 to delay DNA replication. Cells deficient in MLL showed radioresistant DNA synthesis and chromatid-type genomic abnormalities, indicative of S-phase checkpoint dysfunction. Reconstitution of Mll(-/-) (Mll also known as Mll1) mouse embryonic fibroblasts with wild-type but not S516A or ΔSET mutant MLL rescues the S-phase checkpoint defects. Moreover, murine myeloid progenitor cells carrying an Mll-CBP knock-in allele that mimics human t(11;16) leukaemia show a severe radioresistant DNA synthesis phenotype. MLL fusions function as dominant negative mutants that abrogate the ATR-mediated phosphorylation/stabilization of wild-type MLL on damage to DNA, and thus compromise the S-phase checkpoint. Together, our results identify MLL as a key constituent of the mammalian DNA damage response pathway and show that deregulation of the S-phase checkpoint incurred by MLL translocations probably contributes to the pathogenesis of human MLL leukaemias.

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    • "Thus, reduction of MLL1 expression or activity prevents DDR activation and SASP expression through its direct inhibition of the expression of key genes necessary for proliferation, orderly cell cycle progression, and triggering of the DDR (Fig. 7E). Specifically, in contrast to normal OIS cells, MLL1-inhib- ited cells are unable to have a normal progression through the S-phase checkpoint due to the reduced expression of proproliferative cell cycle genes and MLL1's known required role for normal S-phase progression (Liu et al. 2010 ). In turn, these cells are unable to trigger the activation of the DDR and its key mediator for expression of the SASP, ATM phospho-S1981. "

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    • "Phosphorylation of RbBP5 on S350 potentiates WRAD assembly MLL1 is tightly regulated by various mechanisms, including allosteric regulation by the WRAD complex (Dou et al. 2006), deposition of other post-translational modifications on histone proteins (Southall et al. 2009), and phosphorylation of MLL1 by ATR (Liu et al. 2010). In the RbBP5 D/E box (Supplemental Fig. S4), an evolutionarily conserved serine residue (S350) is found in the center of the Ash2L SPRY concave surface (Fig. 3A). "
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    ABSTRACT: The methyltransferase activity of the trithorax group (TrxG) protein MLL1 found within its COMPASS (complex associated with SET1)-like complex is allosterically regulated by a four-subunit complex composed of WDR5, RbBP5, Ash2L, and DPY30 (also referred to as WRAD). We report structural evidence showing that in WRAD, a concave surface of the Ash2L SPIa and ryanodine receptor (SPRY) domain binds to a cluster of acidic residues, referred to as the D/E box, in RbBP5. Mutational analysis shows that residues forming the Ash2L/RbBP5 interface are important for heterodimer formation, stimulation of MLL1 catalytic activity, and erythroid cell terminal differentiation. We also demonstrate that a phosphorylation switch on RbBP5 stimulates WRAD complex formation and significantly increases KMT2 (lysine [K] methyltransferase 2) enzyme methylation rates. Overall, our findings provide structural insights into the assembly of the WRAD complex and point to a novel regulatory mechanism controlling the activity of the KMT2/COMPASS family of lysine methyltransferases. © 2015 Zhang et al; Published by Cold Spring Harbor Laboratory Press.
    Full-text · Article · Jan 2015 · Genes & Development
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    • "Consistently, ChIP analysis of H3K4me3 at the B-globin replication origin in Jurkat cells shows that H3K4me3 does not correlate with CDC45 enrichment [20]. Notably, in cells expressing a catalytic dead mutant of MLL, the H3K4me3 methyltransferase, CDC45 does accumulate at this origin [20]. Thus, H3K4me3 could prevent the binding of CDC45. "
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    ABSTRACT: In eukaryotic organisms, the replication of the DNA sequence and its organization into chromatin is critical to maintain genome integrity. Chromatin components, such as histone variants and histone post-translational modifications, along with the higher-order chromatin structure, impact several DNA metabolic processes, including replication, transcription, and repair. In this review we focus on lysine methylation and the relationships between this histone mark and chromatin replication. We first describe studies implicating lysine methylation in regulating early steps in the replication process. We then discuss chromatin reassembly following replication fork passage, where the incorporation of a combination of newly synthesized histones and parental histones can impact the inheritance of lysine methylation marks on the daughter strands. Finally, we elaborate on how the inheritance of lysine methylation can impact maintenance of the chromatin landscape, using heterochromatin as a model chromatin domain, and we discuss the potential mechanisms involved in this process. This article is part of a Special Issue entitled: Methylation Multifaceted Modification - looking at transcription and beyond, edited by Dr. Johnathan Whetstine.
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