Dual localization of human DNA topoisomerase IIIα to mitochondria and nucleus

The human TOP3α gene encoding DNA topoisomerase IIIα (hTop3α) has two potential start codons for the synthesis of proteins 1,001 and 976 aa residues in length. The sequence of the N-terminal region of the 1,001-residue form resembles signal peptide sequences for mitochondrial import, and fluorescence microscopy shows that the addition of as few as the first 34 aa of the 1,001-residue form of hTop3α to a green fluorescent protein can direct the chimeric protein to mitochondria. Biochemical analyses of subcellular fractions of HeLa cells further demonstrate that a distinctive fraction of hTop3α is present inside mitochondria, as evidenced by its resistance to proteinase K. This fraction constitutes several percent of the enzyme in the nuclear fraction, suggesting that the distribution of the mitochondrial and nuclear forms of hTop3α is roughly in proportion to the DNA contents of these cellular compartments. The presence of a type IA DNA topoisomerase in the mitochondria of other eukaryotes is supported by an examination of the amino acid sequences of mouse and Drosophila DNA topoisomerase IIIα and Schizosaccharomyces pombe DNA topoisomerase III. Given the presence of at least one type IA DNA topoisomerase in all forms of life examined to date, the finding of a type IA enzyme in mitochondria further supports the notion of a key role of such enzymes in DNA transactions.

[1]  W. Neupert,et al.  Protein import into mitochondria. , 2005, Biochemical Society transactions.

[2]  JAMES C. Wang,et al.  Cellular roles of DNA topoisomerases: a molecular perspective , 2002, Nature Reviews Molecular Cell Biology.

[3]  P. Wilairat,et al.  Partial purification of mitochondrial DNA topoisomerase II from Plasmodium falciparum and its sensitivity to inhibitors. , 2001, The Southeast Asian journal of tropical medicine and public health.

[4]  D. Busija,et al.  Isolation and characterization of intact mitochondria from neonatal rat brain. , 2001, Brain research. Brain research protocols.

[5]  R. Bennett,et al.  Association of yeast DNA topoisomerase III and Sgs1 DNA helicase: Studies of fusion proteins , 2001, Proceedings of the National Academy of Sciences of the United States of America.

[6]  Zefeng Wang,et al.  RNA interference of a trypanosome topoisomerase II causes progressive loss of mitochondrial DNA , 2001, The EMBO journal.

[7]  Y. Pommier,et al.  Human mitochondrial topoisomerase I , 2001, Proceedings of the National Academy of Sciences of the United States of America.

[8]  H. Prydz,et al.  Cloning and Characterization of MDDX28, a Putative DEAD-box Helicase with Mitochondrial and Nuclear Localization* , 2001, The Journal of Biological Chemistry.

[9]  P. Pongpech,et al.  Type I topoisomerase activity is required for proper chromosomal segregation in Escherichia coli , 2001, Proceedings of the National Academy of Sciences of the United States of America.

[10]  E. Ladoukakis,et al.  Direct evidence for homologous recombination in mussel (Mytilus galloprovincialis) mitochondrial DNA. , 2001, Molecular biology and evolution.

[11]  P. Pandolfi,et al.  Evidence for BLM and Topoisomerase IIIα interaction in genomic stability , 2001 .

[12]  H. Hayashi,et al.  Nuclear Localization of Yeast Nfs1p Is Required for Cell Survival* , 2001, Journal of Biological Chemistry.

[13]  D. Ramotar,et al.  Pir1p Mediates Translocation of the Yeast Apn1p Endonuclease into the Mitochondria To Maintain Genomic Stability , 2001, Molecular and Cellular Biology.

[14]  Michael R. Olson,et al.  Mitochondria in apoptosis and human disease. , 2001, Current molecular medicine.

[15]  K. Mihara,et al.  Characterization of the Signal That Directs Tom20 to the Mitochondrial Outer Membrane , 2000, The Journal of cell biology.

[16]  Colin R Campbell,et al.  An alternate form of Ku80 is required for DNA end-binding activity in mammalian mitochondria. , 2000, Nucleic acids research.

[17]  J. Wang,et al.  Interaction between yeast sgs1 helicase and DNA topoisomerase III. , 2000, The Journal of biological chemistry.

[18]  I. Hansen,et al.  Localization of glucocorticoid hormone receptors in mitochondria of human cells. , 2000, European journal of cell biology.

[19]  Z. Darżynkiewicz,et al.  Differential expression of human topoisomerase IIIalpha during the cell cycle progression in HL-60 leukemia cells and human peripheral blood lymphocytes. , 2000, Experimental cell research.

[20]  K. Gatter,et al.  The Bloom's Syndrome Gene Product Interacts with Topoisomerase III* , 2000, The Journal of Biological Chemistry.

[21]  R. Jaenisch,et al.  Association of the Bloom syndrome protein with topoisomerase IIIalpha in somatic and meiotic cells. , 2000, Cancer research.

[22]  J. Sedman,et al.  A DNA Helicase Required for Maintenance of the Functional Mitochondrial Genome in Saccharomyces cerevisiae , 2000, Molecular and Cellular Biology.

[23]  L. Langston,et al.  The top3(+) gene is essential in Schizosaccharomyces pombe and the lethality associated with its loss is caused by Rad12 helicase activity. , 1999, Nucleic acids research.

[24]  A. Mondragón,et al.  The structure of Escherichia coli DNA topoisomerase III. , 1999, Structure.

[25]  C. J. Stirling,et al.  The yeast CDC9 gene encodes both a nuclear and a mitochondrial form of DNA ligase I , 1999, Current Biology.

[26]  S. W. Wang,et al.  Topoisomerase III is essential for accurate nuclear division in Schizosaccharomyces pombe. , 1999, Nucleic acids research.

[27]  R. Stierum,et al.  Mitochondrial DNA repair pathways. , 1999, Journal of bioenergetics and biomembranes.

[28]  W. Neupert,et al.  The DNA Helicase, Hmi1p, Is Transported into Mitochondria by a C-terminal Cleavable Targeting Signal* , 1999, The Journal of Biological Chemistry.

[29]  M. Kozak.  Initiation of translation in prokaryotes and eukaryotes. , 1999, Gene.

[30]  U. Lakshmipathy,et al.  The Human DNA Ligase III Gene Encodes Nuclear and Mitochondrial Proteins , 1999, Molecular and Cellular Biology.

[31]  D. Wallace.  Mitochondrial diseases in man and mouse. , 1999, Science.

[32]  J. Nitiss.  Investigating the biological functions of DNA topoisomerases in eukaryotic cells. , 1998, Biochimica et biophysica acta.

[33]  Wei Li,et al.  Mammalian DNA topoisomerase IIIα is essential in early embryogenesis , 1998 .

[34]  A. Tua,et al.  Mitochondrial DNA topoisomerase I of Saccharomyces cerevisiae. , 1997, Biochimie.

[35]  K. Komori,et al.  Cloning and characterization of the gene encoding a mitochondrially localized DNA topoisomerase II in Dictyostelium discoideum. Western blot analysis. , 1997, Biochimica et biophysica acta.

[36]  A. Nicolas,et al.  An atypical topoisomerase II from archaea with implications for meiotic recombination , 1997, Nature.

[37]  S. Keeney,et al.  Meiosis-Specific DNA Double-Strand Breaks Are Catalyzed by Spo11, a Member of a Widely Conserved Protein Family , 1997, Cell.

[38]  B. Thyagarajan,et al.  Mammalian Mitochondria Possess Homologous DNA Recombination Activity* , 1996, The Journal of Biological Chemistry.

[39]  P Vincens,et al.  Computational method to predict mitochondrially imported proteins and their targeting sequences. , 1996, European journal of biochemistry.

[40]  J. Wang,et al.  Human TOP3: a single-copy gene encoding DNA topoisomerase III. , 1996, Proceedings of the National Academy of Sciences of the United States of America.

[41]  F. Castora,et al.  Mammalian mitochondrial DNA topoisomerase I preferentially relaxes supercoils in plasmids containing specific mitochondrial DNA sequences. , 1995, Biochimica et biophysica acta.

[42]  S Gangloff,et al.  The yeast type I topoisomerase Top3 interacts with Sgs1, a DNA helicase homolog: a potential eukaryotic reverse gyrase , 1994, Molecular and cellular biology.

[43]  B. Rost,et al.  Combining evolutionary information and neural networks to predict protein secondary structure , 1994, Proteins.

[44]  Alfonso Mondragón,et al.  Three-dimensional structure of the 67K N-terminal fragment of E. coli DNA topoisomerase I , 1994, Nature.

[45]  B. Rost,et al.  Prediction of protein secondary structure at better than 70% accuracy. , 1993, Journal of molecular biology.

[46]  W. Neupert,et al.  Targeting of the master receptor MOM19 to mitochondria. , 1991, Science.

[47]  A K Hopper,et al.  mRNA leader length and initiation codon context determine alternative AUG selection for the yeast gene MOD5. , 1991, Proceedings of the National Academy of Sciences of the United States of America.

[48]  G. Soslau,et al.  Mitochondrial DNA topoisomerase I from human platelets. , 1991, Biochimica et biophysica acta.

[49]  G von Heijne,et al.  Cleavage-site motifs in mitochondrial targeting peptides. , 1990, Protein engineering.

[50]  J. Wallis,et al.  A hyper-recombination mutation in S. cerevisiae identifies a novel eukaryotic topoisomerase , 1989, Cell.

[51]  C. Sheline,et al.  Localization of a type II DNA topoisomerase to two sites at the periphery of the kinetoplast DNA of Crithidia fasciculata , 1988, Cell.

[52]  M. Kozak.  An analysis of 5'-noncoding sequences from 699 vertebrate messenger RNAs. , 1987, Nucleic acids research.

[53]  W. G. Kelly,et al.  Purification and characterization of a type I DNA topoisomerase from calf thymus mitochondria. , 1987, Biochemistry.

[54]  R. Low.  Isolation of DNA topoisomerases I and II activities from mitochondria of mammalian heart. , 2002, Methods in molecular biology.

[55]  S. Keeney,et al.  Mechanism and control of meiotic recombination initiation. , 2001, Current topics in developmental biology.

[56]  J. Champoux.  DNA topoisomerases: structure, function, and mechanism. , 2001, Annual review of biochemistry.

[57]  K. Nakai,et al.  PSORT: a program for detecting sorting signals in proteins and predicting their subcellular localization. , 1999, Trends in biochemical sciences.

[58]  G. Schneider,et al.  Feature‐extraction from endopeptidase cleavage sites in mitochondrial targeting peptides , 1998 .

[59]  G. Shadel,et al.  Mitochondrial DNA maintenance in vertebrates. , 1997, Annual review of biochemistry.

[60]  T. Shapiro,et al.  The structure and replication of kinetoplast DNA. , 1995, Annual review of microbiology.

[61]  B. Storrie,et al.  Isolation of subcellular organelles. , 1990, Methods in enzymology.