The yeast CDC9 gene encodes both a nuclear and a mitochondrial form of DNA ligase I.
BACKGROUND
The yeast CDC9 gene encodes a DNA ligase I activity required during nuclear DNA replication to ligate the Okazaki fragments formed when the lagging DNA strand is synthesised. The only other DNA ligase predicted from the yeast genome sequence, DNL4/LIG4, is specifically involved in a non-homologous DNA end-joining reaction. What then is the source of the DNA ligase activity required for replication of the yeast mitochondrial genome?


RESULTS
We report that CDC9 encodes two distinct polypeptides expressed from consecutive in-frame AUG codons. Translational initiation at these two sites gives rise to polypeptides differing by a 23 residue amino-terminal extension, which corresponds to a functional mitochondrial pre-sequence sufficient to direct import into yeast mitochondria. Initiation at the first AUG codon results in a 755 amino-acid polypeptide that is imported into mitochondria, whereupon the pre-sequence is proteolytically removed to yield the mature mitochondrial form of Cdc9p. Initiation at the second AUG codon produces a 732 amino-acid polypeptide, which is localised to the nucleus. Cells expressing only the nuclear isoform were found to be specifically defective in the maintenance of the mitochondrial genome.


CONCLUSIONS
CDC9 encodes two distinct forms of DNA ligase I. The first is targeted to the mitochondrion and is required for propagation and maintenance of mitochondrial DNA, the second localises to the nucleus and is sufficient for the essential cell-division function associated with this gene.
Willer
M
M
School of Biological Sciences 2.205 Stopford Building University of Manchester Oxford Road, Manchester, M13 9PT, UK.
Rainey
M
M
Pullen
T
T
Stirling
C J
CJ
D000595:Amino Acid Sequence; D015153:Blotting, Western; D002467:Cell Nucleus / Q000201:enzymology* / Q000235:genetics; D003062:Codon; D000072481:DNA Ligase ATP; D011088:DNA Ligases / Q000235:genetics / Q000378:metabolism*; D018604:Epitope Mapping; D015966:Gene Expression Regulation, Fungal; D008856:Microscopy, Fluorescence; D008928:Mitochondria / Q000201:enzymology* / Q000235:genetics; D008969:Molecular Sequence Data; D016297:Mutagenesis, Site-Directed; D011993:Recombinant Fusion Proteins / Q000378:metabolism; D012441:Saccharomyces cerevisiae / Q000201:enzymology* / Q000235:genetics
C000607890:CDC9 protein, S cerevisiae; D003062:Codon; C000607891:DNL4 protein, S cerevisiae; D011993:Recombinant Fusion Proteins; D011088:DNA Ligases; D000072481:DNA Ligase ATP