Interaction between PCNA and Diubiquitinated Mcm10 Is Essential for Cell Growth in Budding Yeast

Das-Bradoo, Sapna; Ricke, Robin M.; Bielinsky, Anja‐Katrin

doi:10.1128/mcb.02062-05

Cited by 69 publications

(124 citation statements)

References 69 publications

(110 reference statements)

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“…For example, RPA, a eukaryotic recruiting and scaffolding protein critical to DNA replication, has been shown to bind both oligonucleotides and peptides through its six OB-fold domains (66 -70). Mcm10 appears to exhibit similar behavior by binding to DNA, pol ␣, Dbf4-dependent kinase, and PCNA (24,25,31), although the role of the OB-fold in Mcm10 interactions with Dbf4-dependent kinase and PCNA remains to be determined.…”

Section: Chemical Nature Of Mcm10-id Interactions With Dna Andmentioning

confidence: 99%

“…This follows a common strategy for numerous modular proteins involved in DNA processing; there is a significant kinetic advantage to deconstructing protein interactions into two or more weak binding sites (68). The recruitment of pol ␣ to origins of replication by Mcm10 would be a significant step to signal nascent DNA synthesis and contribute to fork stability (6,12,16,24,26,27,29,71). Indeed, Mcm10 has been shown to be necessary for pol ␣ loading onto chromatin (4).…”

Section: A Molecular Mechanism For Mcm10mentioning

confidence: 99%

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Physical Interactions between Mcm10, DNA, and DNA Polymerase α

Warren

Huang²,

Fanning³

et al. 2009

Journal of Biological Chemistry

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Mcm10 is an essential eukaryotic protein required for the initiation and elongation phases of chromosomal replication. Specifically, Mcm10 is required for the association of several replication proteins, including DNA polymerase ␣ (pol ␣), with chromatin. We showed previously that the internal (ID) and C-terminal (CTD) domains of Mcm10 physically interact with both single-stranded (ss) DNA and the catalytic p180 subunit of pol ␣. However, the mechanism by which Mcm10 interacts with pol ␣ on and off DNA is unclear. As a first step toward understanding the structural details for these critical intermolecular interactions, x-ray crystallography and NMR spectroscopy were used to map the binary interfaces between Mcm10-ID, ssDNA, and p180. The crystal structure of an Mcm10-ID⅐ssDNA complex confirmed and extended our previous evidence that ssDNA binds within the oligonucleotide/oligosaccharide binding-fold cleft of Mcm10-ID. We show using NMR chemical shift perturbation and fluorescence spectroscopy that p180 also binds to the OB-fold and that ssDNA and p180 compete for binding to this motif. In addition, we map a minimal Mcm10 binding site on p180 to a small region within the p180 N-terminal domain (residues 286 -310). These findings, together with data for DNA and p180 binding to an Mcm10 construct that contains both the ID and CTD, provide the first mechanistic insight into how Mcm10 might use a handoff mechanism to load and stabilize pol ␣ within the replication fork.

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Section: Chemical Nature Of Mcm10-id Interactions With Dna Andmentioning

confidence: 99%

Section: A Molecular Mechanism For Mcm10mentioning

confidence: 99%

Physical Interactions between Mcm10, DNA, and DNA Polymerase α

Warren

Huang²,

Fanning³

et al. 2009

Journal of Biological Chemistry

View full text Add to dashboard Cite

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“…Because Mcm10 is necessary for chromatin association of the entire pol-␣/primase complex in budding yeast (Ricke and Bielinsky, 2004), it probably has a crucial role in lagging strand synthesis. The initiation of individual Okazaki fragments requires the repeated action of pol-␣/primase (Nasheuer and Grosse, 1987) to generate a primed template onto which PCNA can be loaded, and it is conceivable that Mcm10 facilitates the RNA-DNA primer synthesis by pol-␣/primase and may also be involved in the subsequent recruitment of PCNA (Das-Bradoo et al, 2006). Alternatively, Mcm10 itself may function as a primase.…”

Section: Introductionmentioning

confidence: 99%

“…Thus, it is not surprising that, in vitro, Mcm10 prefers ss over double-stranded (ds) DNA (Fien et al, 2004). Within the central OB-fold resides a proliferating cell nuclear antigen (PCNA) interacting protein motif (Fien et al, 2004;Das-Bradoo et al, 2006), or PIP-box, as well as a hydrophobic stretch, similar to the one found in the mobile loop of heat-shock factor (Hsp) 10 (Ricke and . Although the PIP box mediates the interaction with PCNA (Das-Bradoo et al, 2006), the Hsp10-like domain forms part of the interaction site with the catalytic subunit of DNA polymerase (pol)-␣ ( Bielinsky, 2004, 2006).…”

Section: Introductionmentioning

confidence: 99%

“…Within the central OB-fold resides a proliferating cell nuclear antigen (PCNA) interacting protein motif (Fien et al, 2004;Das-Bradoo et al, 2006), or PIP-box, as well as a hydrophobic stretch, similar to the one found in the mobile loop of heat-shock factor (Hsp) 10 (Ricke and . Although the PIP box mediates the interaction with PCNA (Das-Bradoo et al, 2006), the Hsp10-like domain forms part of the interaction site with the catalytic subunit of DNA polymerase (pol)-␣ ( Bielinsky, 2004, 2006). Because purified S. pombe Mcm10 coimmunoprecipitates purified p180, the catalytic subunit of pol-␣ in humans, it is reasonable to conclude that the binding between Mcm10 and pol-␣ is direct and that it does not require any other mediators.…”

Section: Introductionmentioning

confidence: 99%

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Human Mcm10 Regulates the Catalytic Subunit of DNA Polymerase-α and Prevents DNA Damage during Replication

Chattopadhyay

Bielinsky

2007

MBoC

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In Saccharomyces cerevisiae, minichromosome maintenance protein (Mcm) 10 interacts with DNA polymerase (pol)-␣ and functions as a nuclear chaperone for the catalytic subunit, which is rapidly degraded in the absence of Mcm10. We report here that the interaction between Mcm10 and pol-␣ is conserved in human cells. We used a small interfering RNA-based approach to deplete Mcm10 in HeLa cells, and we observed that the catalytic subunit of pol-␣, p180, was degraded with similar kinetics as Mcm10, whereas the regulatory pol-␣ subunit, p68, remained unaffected. Simultaneous loss of Mcm10 and p180 inhibited S phase entry and led to an accumulation of already replicating cells in late S/G2 as a result of DNA damage, which triggered apoptosis in a subpopulation of cells. These phenotypes differed considerably from analogous studies in Drosophila embryo cells that did not exhibit a similar arrest. To further dissect the roles of Mcm10 and p180 in human cells, we depleted p180 alone and observed a significant delay in S phase entry and fork progression but little effect on cell viability. These results argue that cells can tolerate low levels of p180 as long as Mcm10 is present to "recycle" it. Thus, human Mcm10 regulates both replication initiation and elongation and maintains genome integrity. INTRODUCTIONDNA replication is a highly regulated process in which multiprotein complexes generate an exact copy of a cell's DNA. These multiprotein complexes are assembled into replication forks. Fork assembly takes place at replication origins that are spread throughout the genome in all eukaryotic cells. The first step is the formation of a prereplicative complex (pre-RC) (Diffley et al., 1994), which is subsequently transformed into a preinitiation complex, and finally into a pair of functional replication forks that have the ability to unwind parental DNA and synthesize new daughter strands (Mendez and Stillman, 2003). DNA unwinding is likely catalyzed by the minichromosome maintenance (Mcm) 2-7 complex (Aparicio et al., 1997;Labib et al., 2000;Shechter et al., 2004) in conjunction with two coactivators, Cdc45 and GINS (Pacek and Walter, 2004;Gambus et al., 2006;Moyer et al., 2006;Pacek et al., 2006). The association of Cdc45 and GINS with DNA is interdependent (Kubota et al., 2003;Takayama et al., 2003), and it requires yet another factor, Mcm10 (Wohlschlegel et al., 2002;Gregan et al., 2003;Sawyer et al., 2004). Despite its name, Mcm10 is not a member of the MCM protein family, although it was isolated in the same genetic screen in budding yeast that identified the MCM2-7 genes (Solomon et al., 1992;Merchant et al., 1997).Mcm10 is a constitutively nuclear DNA binding protein (Merchant et al., 1997;Burich and Lei, 2003) that is essential in yeast (Burich and Lei, 2003). Besides its role in DNA replication, Mcm10 has also been implicated in transcriptional silencing (Liachko and Tye, 2005). It is important to note that the general protein domain structure of Mcm10 is not unique in Saccharomyces cerevisiae, but it is highly con...

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Current awareness on yeast

2007

Yeast

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In order to keep subscribers up‐to‐date with the latest developments in their field, this current awareness service is provided by John Wiley & Sons and contains newly‐published material on yeasts. Each bibliography is divided into 10 sections. 1 Books, Reviews & Symposia; 2 General; 3 Biochemistry; 4 Biotechnology; 5 Cell Biology; 6 Gene Expression; 7 Genetics; 8 Physiology; 9 Medical Mycology; 10 Recombinant DNA Technology. Within each section, articles are listed in alphabetical order with respect to author. If, in the preceding period, no publications are located relevant to any one of these headings, that section will be omitted. (4 weeks journals ‐ search completed 11th. Oct. 2006)

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Interaction between PCNA and Diubiquitinated Mcm10 Is Essential for Cell Growth in Budding Yeast

Cited by 69 publications

References 69 publications

Physical Interactions between Mcm10, DNA, and DNA Polymerase α

Physical Interactions between Mcm10, DNA, and DNA Polymerase α

Human Mcm10 Regulates the Catalytic Subunit of DNA Polymerase-α and Prevents DNA Damage during Replication

Current awareness on yeast

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