A possible functional role for a new class of eukaryotic DNA polymerases.

The recent Correspondence from Sonnhammer and Wootton [1xWidespread eukaryotic sequences, highly similar to bacterial DNA polymerase I, looking for functions. Sonnhammer, EL and Wootton, JC. Curr Biol. 1997; 7: R463–R465Abstract | Full Text | Full Text PDF | PubMedSee all References[1] regarding a new family of eukaryotic proteins similar to bacterial DNA polymerase I raises a number of interesting questions. Our report on the characterization of the Drosophila DNA repair gene mus308[2xMolecular cloning of Drosophila mus308, a gene involved in DNA cross-link repair with homology to prokaryotic DNA polymerase I genes. Harris, PV, Mazina, OM, Leonhardt, EA, Case, RB, Boyd, JB, and Burtis, KC. Mol Cell Biol. 1996; 16: 5764–5771PubMedSee all References[2] and a related Caenorhabditis elegans gene addresses several of these questions, including the possible role of this gene family.The mus308 gene (Genbank L76559) encodes a 229 kDa polypeptide, the carboxy-terminal domain of which is closely related to the polymerase domain of the bacterial DNA polymerase I enzymes. Remarkably, the amino-terminal domain of the Mus308 polypeptide also includes the seven amino-acid sequence motifs that are characteristic of the ‘superfamily 2’ DNA and RNA helicases [3xTwo related superfamilies of putative helicases involved in replication, recombination, repair and expression of DNA and RNA genomes. Gorbalenya, AE, Koonin, EV, Donchenko, AP, and Blinov, VM. Nucleic Acids Res. 1989; 17: 4713–4730Crossref | PubMed | Scopus (679)See all References[3]. This finding makes mus308 not only the first characterized gene encoding a eukaryotic homolog of the bacterial DNA polymerase I family, but also the first reported gene encoding both helicase and polymerase motifs in a single polypeptide.A gene very similar to mus308, which we have designated mus-1, encoded by genomic sequences in the cosmids R12B2 (U00066) and W03A3 (U50184), is predicted to exist in C. elegans. A combination of genomic and partial cDNA sequences indicate that the carboxy-terminal domain of the Mus-1 polypeptide is highly homologous to the polymerase domains of Mus308 and other members of the DNA polymerase I family [2xMolecular cloning of Drosophila mus308, a gene involved in DNA cross-link repair with homology to prokaryotic DNA polymerase I genes. Harris, PV, Mazina, OM, Leonhardt, EA, Case, RB, Boyd, JB, and Burtis, KC. Mol Cell Biol. 1996; 16: 5764–5771PubMedSee all References[2], as noted by Sonnhammer and Wootton [1xWidespread eukaryotic sequences, highly similar to bacterial DNA polymerase I, looking for functions. Sonnhammer, EL and Wootton, JC. Curr Biol. 1997; 7: R463–R465Abstract | Full Text | Full Text PDF | PubMedSee all References[1]. Upon comparing the remainder of the mus308 coding sequences and C. elegans genomic sequences located upstream of the Mus-1 polymerase domain, a helicase domain closely related to the Mus308 helicase domain was immediately apparent [2xMolecular cloning of Drosophila mus308, a gene involved in DNA cross-link repair with homology to prokaryotic DNA polymerase I genes. Harris, PV, Mazina, OM, Leonhardt, EA, Case, RB, Boyd, JB, and Burtis, KC. Mol Cell Biol. 1996; 16: 5764–5771PubMedSee all References[2]. Further limited sequence homology between the Mus308 and predicted Mus-1 polypeptides was also seen in the region between the helicase and polymerase domains (P.H., unpublished observations). The juxtaposition of Mus308-related helicase and polymerase domains and sequence similarities in the region between the two domains strongly support the hypothesis that the mus-1 gene encodes a protein homologous to Mus308. The existence of an evolutionarily conserved enzyme family with this unique combination of helicase and polymerase motifs suggests a conserved functional role in DNA repair for these proteins. Direct evidence for such a role is available, however, only for Mus308 [4xThird-chromosome mutagen-sensitive mutants of Drosophila melanogaster. Boyd, JB, Golino, MD, Shaw, KE, Osgood, CJ, and Green, MM. Genetics. 1981; 97: 607–623PubMedSee all References[4], which is required to repair the damage caused by mutagens such as nitrogen mustard and cisplatin that result in interstrand cross-links [5xmus308 mutants of Drosophila exhibit hypersensitivity to DNA cross-linking agents and are defective in a deoxyribonuclease. Boyd, JB, Sakaguchi, K, and Harris, PV. Genetics. 1990; 125: 813–819PubMedSee all References[5].As reported by Sonnhammer and Wootton [1xWidespread eukaryotic sequences, highly similar to bacterial DNA polymerase I, looking for functions. Sonnhammer, EL and Wootton, JC. Curr Biol. 1997; 7: R463–R465Abstract | Full Text | Full Text PDF | PubMedSee all References[1], two distinct human sequences encoding polypeptides with similarity to the polymerase domain of the bacterial DNA polymerase I family are also present in the sequence databases. We have also noted the existence of these coding sequences, which encode polypeptides related to the Mus308 polymerase domain. The sequence most closely related to mus308 is represented by human cDNA clone za38h12.r1 (Genbank W00829. Sequence information from partial cDNAs and RT-PCR products derived from this gene show additional homology to mus308 in sequences upstream of the polymerase domain (P.V.H. and K.C.B., unpublished observations), suggesting homology to the mus308/mus-1 family; however, the presence of helicase motifs remains to be established. We have also noted the sequences homologous to the Mus308 polymerase domain in the Huntington's disease region of the genome, as reported by Sonnhammer and Wootton [1xWidespread eukaryotic sequences, highly similar to bacterial DNA polymerase I, looking for functions. Sonnhammer, EL and Wootton, JC. Curr Biol. 1997; 7: R463–R465Abstract | Full Text | Full Text PDF | PubMedSee all References[1]. In this case, however, the availability of almost 2 megabases of sequence information upstream of this domain reveals the absence of any recognizable helicase motifs (P.V.H. and K.C.B., unpublished observations), suggesting that this gene is not likely to be a functional homolog of the mus308/mus-1 family.Sonnhammer and Wootton [1xWidespread eukaryotic sequences, highly similar to bacterial DNA polymerase I, looking for functions. Sonnhammer, EL and Wootton, JC. Curr Biol. 1997; 7: R463–R465Abstract | Full Text | Full Text PDF | PubMedSee all References[1] note that it is ‘tempting to consider DNA repair’ as a possible function of these DNA polymerases, but that ‘experimental verification of polymerase activity is the next step.’ Our previous report [2xMolecular cloning of Drosophila mus308, a gene involved in DNA cross-link repair with homology to prokaryotic DNA polymerase I genes. Harris, PV, Mazina, OM, Leonhardt, EA, Case, RB, Boyd, JB, and Burtis, KC. Mol Cell Biol. 1996; 16: 5764–5771PubMedSee all References[2] indeed confirms a role in DNA repair for at least the prototypical member of this new gene family, mus308. Furthermore, we have recently determined that the polymerase domain of the polypeptide encoded by the za38h12.r1 human cDNA possesses enzymatic polymerase activity, confirming the prediction based on amino-acid sequence (P.V.H. and K.C.B., unpublished observations). In the light of this enzymatic activity, we have termed this new human DNA polymerase gene POLH. The next step is to determine the functional role of the polymerase (and possibly helicase) domains of this new class of eukaryotic DNA polymerases in DNA repair and other cellular functions.