Cloning and Chromosomal Mapping of the Human DNA Polymerase θ (POLQ), the Eighth Human DNA Polymerase

Sharief, Farida S.; Vojta, Patrick J.; Ropp, Patricia A.; Copeland, William C.

doi:10.1006/geno.1999.5843

Cited by 108 publications

(71 citation statements)

References 30 publications

Supporting

Mentioning

Contrasting

Order By: Relevance

“…These authors designated human DinB1 protein as DNA polymerase (pol). In view of the fact that this name has been assigned to the product of another gene (27), we suggest that the DinB1 gene product be referred to as pol, a designation approved by the Human Genome Organization nomenclature committee.…”

Section: Discussionmentioning

confidence: 99%

Purification and Characterization of polκ, a DNA Polymerase Encoded by the Human DINB1 Gene

Gerlach¹,

Feaver²,

Fischhaber³

et al. 2001

Journal of Biological Chemistry

115

103

View full text Add to dashboard Cite

The Escherichia coli dinB gene encodes DNA polymerase (pol) IV, a protein involved in increasing spontaneous mutations in vivo. The protein-coding region of DINB1, the human ortholog of DNA pol IV, was fused to glutathione S-transferase and expressed in insect cells. The purified fusion protein was shown to be a templatedirected DNA polymerase that we propose to designate pol. Human pol lacks detectable 3 3 5 proofreading exonuclease activity and is not stimulated by recombinant human proliferating cell nuclear antigen in vitro. Between pH 6.5 and 8.5, human pol possesses optimal activity at 37°C over the pH range 6.5-7.5, and is insensitive to inhibition by aphidicolin, dideoxynucleotides, or NaCl up to 50 mM. Either Mg 2؉ or Mn 2؉ can satisfy a metal cofactor requirement for pol activity, with Mg 2؉ being preferred. Human pol is unable to bypass a cisplatin adduct in the template. However, pol shows limited bypass of an 2-acetylaminofluorene lesion and can incorporate dCTP or dTTP across from this lesion, suggesting that the bypass is potentially mutagenic. These results are consistent with a model in which pol acts as a specialized DNA polymerase whose possible role is to facilitate the replication of templates containing abnormal bases, or possessing structurally aberrant replication forks that inhibit normal DNA synthesis.We previously reported the cloning and characterization of the human DINB1 and mouse Dinb1 genes, mammalian orthologs of the Escherichia coli dinB gene (1) and members of the UmuC/DinB superfamily of DNA polymerases (2). Expression of the E. coli dinB gene is tightly regulated by the SOS system (3). Following exposure of E. coli cells to DNA-damaging agents such as ultraviolet (UV) radiation, induction of dinB results in enhanced spontaneous (untargeted) mutagenesis of phage DNA introduced into the bacteria subsequent to irradiation (4). Increased spontaneous mutagenesis is also observed following overexpression of dinB in cells transfected with FЈlac plasmids, with the most prevalent mutations detected being Ϫ1 frameshifts (5). Recombinant E. coli DinB protein carrying a 6-histidine tag was purified and shown to be a DNA polymerase, designated DNA pol IV of E. coli, which is devoid of detectable exonuclease activity (6). Consistent with its apparent ability to generate frameshift mutations in vivo, DNA pol IV is able to extend a misaligned primer-template in vitro, resulting in a Ϫ1 frameshift mutation (6). More recently, DNA pol IV has been shown to be unable to efficiently bypass an abasic site, thymine dimer, or 6-4 photoproduct in vitro (7). Based on these observations, it has been suggested that DNA pol IV is a specialized enzyme whose role is to negotiate sites of stalled or arrested DNA replication caused by structurally abnormal replication forks, such as those caused by slippage at repeated sequences (2, 6, 7).Human DINB1 cDNA is predicted to encode a polypeptide with a molecular mass of 99 kDa, which shares extensive amino acid sequence homology with E. coli DNA pol IV, includin...

show abstract

Section: Discussionmentioning

confidence: 99%

Purification and Characterization of polκ, a DNA Polymerase Encoded by the Human DINB1 Gene

Gerlach¹,

Feaver²,

Fischhaber³

et al. 2001

Journal of Biological Chemistry

115

103

View full text Add to dashboard Cite

show abstract

“…Among these, 30% were directly involved in DNA replication, cell cycle progression and chromosomal segregation, indicating that MAPK1 indeed promotes expressions of genes essential for cell proliferation. They included genes encoding cyclins such as CCNA2, CCNB1, CCNB2 and CCNE2 (Hunter and Pines, 1991), kinetocore-related molecules such as AURKA, AURKB, CENPA and CENPF (Hussein and Taylor, 2002;Kunitoku et al, 2003;Marumoto et al, 2005), microtubule-associated molecules such as TPX2 and KIF2C (Kufer et al, 2002;Hunter et al, 2003), RRM1 encoding ribonucleotide reductase (Parker et al, 1995), POLQ encoding DNA polymerase theta (Sharief et al, 1999), PRIM1 encoding DNA primase (Stadlbauer et al, 1994), MCM4 encoding DNA replication-initiation factor (Musahl et al, 1995) and RFC4 encoding DNA replication factor (Wang et al, 2000). Genes of interest in terms of implication in cancer progression were MMP1 and EPHA2; both are associated with invasion and metastasis (Kinch and Carles-Kinch, 2003;Seiki and Yana, 2003).…”

Section: Targets Of Mapk1 In Pancreatic Cancer T Furukawa Et Almentioning

confidence: 99%

AURKA is one of the downstream targets of MAPK1/ERK2 in pancreatic cancer

et al. 2006

View full text Add to dashboard Cite

DUSP6/MKP-3, a specific inhibitor of MAPK1/ERK2, frequently loses its expression in primary pancreatic cancer tissues. This evidence suggests that constitutive activation of MAPK1 synergistically induced by frequent mutation of KRAS2 and the loss of function of DUSP6 plays key roles in pancreatic carcinogenesis and progression. By profiling of gene expressions associated with downregulation of MAPK1 induced by exogenous overexpression of DUSP6 in pancreatic cancer cells, we found that AURKA/STK15, the gene encoding Aurora-A kinase, which plays key roles in cellular mitosis, was among the downregulated genes along with its related genes, which included AURKB, TPX2 and CENPA. An association of expression and promoter activity of AURKA with MAPK activity was verified. Knockdown of ETS2 resulted in a reduction of AURKA expression. These results indicate that AURKA is a direct target of the MAPK pathway and that its overexpression in pancreatic cancer is induced by hyperactivation of the pathway, at least via ETS2.

show abstract

“…To specifically inactivate the DNA polymerase activity of Pol while leaving the helicase and other potentially important domains intact, we deleted exons 25 (185 base pairs) and 26 (112 base pairs), which encode a conserved aspartic acid residue known to be essential for the polymerase catalytic activity (23,24) and the tyrosine residue that binds the incoming nucleotide (11,14), respectively ( Fig. 1 A and B).…”

Section: Generation Of Mice Specifically Devoid Of Pol Polymerase Actmentioning

confidence: 99%

“…DNA polymerase (Pol ) is a Ϸ300-kDa family A polymerase with a unique structure, having a helicase domain in its N-terminal portion and a polymerase domain in its C terminus (11)(12)(13). Polq, the gene encoding Pol , has a homolog in Drosophila, the mus308 gene (14).…”

mentioning

confidence: 99%

DNA polymerase θ contributes to the generation of C/G mutations during somatic hypermutation of Ig genes

Masuda

Ouchida

Takeuchi

et al. 2005

Proc. Natl. Acad. Sci. U.S.A.

103

View full text Add to dashboard Cite

Somatic hypermutation of Ig variable region genes is initiated by activation-induced cytidine deaminase; however, the activity of multiple DNA polymerases is required to ultimately introduce mutations. DNA polymerase (Pol ) has been implicated in mutations at A͞T, but polymerases involved in C͞G mutations have not been identified. We have generated mutant mice expressing DNA polymerase (Pol ) specifically devoid of polymerase activity. Compared with WT mice, Polq-inactive (Polq, the gene encoding Pol ) mice exhibited a reduced level of serum IgM and IgG1. The mutant mice mounted relatively normal primary and secondary immune responses to a T-dependent antigen, but the production of high-affinity specific antibodies was partially impaired. Analysis of the J H4 intronic sequences revealed a slight reduction in the overall mutation frequency in Polq-inactive mice. Remarkably, although mutations at A͞T were unaffected, mutations at C͞G were significantly decreased, indicating an important, albeit not exclusive, role for Pol activity. The reduction of C͞G mutations was particularly focused on the intrinsic somatic hypermutation hotspots and both transitions and transversions were similarly reduced. These findings, together with the recent observation that Pol efficiently catalyzes the bypass of abasic sites, lead us to propose that Pol introduces mutations at C͞G by replicating over abasic sites generated via uracil-DNA glycosylase.abasic site ͉ low-fidelity DNA polymerase ͉ activation-induced cytidine deaminase ͉ uracil-DNA glycosylase

show abstract

Cloning and Chromosomal Mapping of the Human DNA Polymerase θ (POLQ), the Eighth Human DNA Polymerase

Cited by 108 publications

References 30 publications

Purification and Characterization of polκ, a DNA Polymerase Encoded by the Human DINB1 Gene

Purification and Characterization of polκ, a DNA Polymerase Encoded by the Human DINB1 Gene

AURKA is one of the downstream targets of MAPK1/ERK2 in pancreatic cancer

DNA polymerase θ contributes to the generation of C/G mutations during somatic hypermutation of Ig genes

Contact Info

Product

Resources

About