Abstract
Exonuclease VII (ExoVII) of Escherichia coli is a single strandspecific DNA nuclease composed of two different subunits: the large subunit, XseA, and the small subunit, XseB. In this study, we found that multicopy single-stranded DNAs (msDNAs), Ec83 and Ec78, are the in vivo substrates of ExoVII; the enzyme cuts the phosphodiester bond between the fourth and fifth nucleotides from the 5′end. We used this msDNA cleavage to assess ExoVII activity in vivo. Both subunits were required for enzyme activity. Expression of XseA without XseB caused cell death, even though no ExoVII activity was detected. The lethality caused by XseA was rescued by surplus XseB. In XseA-induced death, cells were elongated and multinucleated, and their chromosomes were fragmented and condensed; these are the morphological hallmarks of apoptotic cell death in bacteria. A putative caspase recognition sequence (FVAD) was found in XseA, and its hypothetical caspase product with 257 amino acids was as active as the intact protein in inducing cell death. We propose that under ordinary conditions, XseA protects chromosome as a component of the ExoVII enzyme, but in some conditions, the protein causes cell death; the destruction of cell is probably carried out by the amino terminal fragment derived from the cleavage of XseA by caspase-like enzyme.
This is a preview of subscription content, log in via an institution to check access.
Similar content being viewed by others
References
Baba, T., Ara, T., Hasegawa, M., Takai, Y., Okumura, Y., Baba, M., Datsenko, K.A., Tomita, M., Wanner, B.L., and Mori, H. 2006. Construction of Escherichia coli K-12 in-frame, single-gene knockout mutants: the Keio collection. Mol. Syst. Biol. 2, 2006.0008.
Bayles, K.W. 2014. Bacterial programmed cell death: making sense of a paradox. Nat. Rev. Microbiol. 12, 63–69.
Burdett, V., Baitinger, C., Viswanathan, M., Lovett, S.T., and Modrich, P. 2001. In vivo requirement for RecJ, ExoVII, ExoI, and ExoX in methyl-directed mismatch repair. Proc. Natl. Acad. Sci. USA 98, 6765–6770.
Carmona-Gutierrez, D., Kroemer, G., and Madeo, F. 2012. When death was young: an ancestral apoptotic network in bacteria. Mol. Cell 46, 552–554.
Chase, J.W., Rabin, B.A., Murphy, J.B., Stone, K.L., and Williams, K.R. 1986. Escherichia coli exonuclease VII. Cloning and sequencing of the gene encoding the large subunit (xseA). J. Biol. Chem. 261, 14929–14935.
Chase, J.W. and Richardson, C.C. 1974a. Exonuclease VII of Escherichia coli. Mechanism of action. J. Biol. Chem. 249, 4553–4561.
Chase, J.W. and Richardson, C.C. 1974b. Exonuclease VII of Escherichia coli. Purification and properties. J. Biol. Chem. 249, 4545–4552.
Datsenko, K.A. and Wanner, B.L. 2000. One-step inactivation of chromosomal genes in Escherichia coli K-12 using PCR products. Proc. Natl. Acad. Sci. USA 97, 6640–6645.
Dermic, D. 2006. Functions of multiple exonucleases are essential for cell viability, DNA repair and homologous recombination in recD mutants of Escherichia coli. Genetics 172, 2057–2069.
Dwyer, D.J., Camacho, D.M., Kohanski, M.A., Callura, J.M., and Collins, J.J. 2012. Antibiotic-induced bacterial cell death exhibits physiological and biochemical hallmarks of apoptosis. Mol. Cell 46, 561–572.
Erental, A., Kalderon, Z., Saada, A., Smith, Y., and Engelberg-Kulka, H. 2014. Apoptosis-like death, an extreme SOS response in Escherichia coli. mBio 5, e01426–01414.
Farzadfard, F. and Lu, T.K. 2014. Genomically encoded analog memory with precise in vivo DNA writing in living cell populations. Science 346, 1256272.
Hakansson, A.P., Roche-Hakansson, H., Mossberg, A.K., and Svanborg, C. 2011. Apoptosis-like death in bacteria induced by HAMLET, a human milk lipid-protein complex. PLoS One 6, e17717.
Harris, R.S., Ross, K.J., Lombardo, M.J., and Rosenberg, S.M. 1998. Mismatch repair in Escherichia coli cells lacking single-strand exonucleases ExoI, ExoVII, and RecJ. J. Bacteriol. 180, 989–993.
Kidd, V.J. 1998. Proteolytic activities that mediate apoptosis. Annu. Rev. Physiol. 60, 533–573.
Kim, K., Jeong, D., and Lim, D. 1997. A mutational study of the site-specific cleavage of EC83, a multicopy single-stranded DNA (msDNA): nucleotides at the msDNA stem are important for its cleavage. J. Bacteriol. 179, 6518–6521.
Larrea, A.A., Pedroso, I.M., Malhotra, A., and Myers, R.S. 2008. Identification of two conserved aspartic acid residues required for DNA digestion by a novel thermophilic Exonuclease VII in Thermotoga maritima. Nucleic Acids Res. 36, 5992–6003.
Lim, D. 1992. Structure and biosynthesis of unbranched multicopy single-stranded DNA by reverse transcriptase in a clinical Escherichia coli isolate. Mol. Microbiol. 6, 3531–3542.
Lim, D. and Maas, W.K. 1989. Reverse transcriptase-dependent synthesis of a covalently linked, branched DNA-RNA compound in E. coli B. Cell 56, 891–904.
Lima, T.M. and Lim, D. 1997. A novel retron that produces RNAless msDNA in Escherichia coli using reverse transcriptase. Plasmid 38, 25–33.
Morelle, S., Carbonnelle, E., Matic, I., and Nassif, X. 2005. Contact with host cells induces a DNA repair system in pathogenic Neisseriae. Mol. Microbiol. 55, 853–861.
Poleszak, K., Kaminska, K.H., Dunin-Horkawicz, S., Lupas, A., Skowronek, K.J., and Bujnicki, J.M. 2012. Delineation of structural domains and identification of functionally important residues in DNA repair enzyme exonuclease VII. Nucleic Acids Res. 40, 8163–8174.
Repar, J., Briski, N., Buljubasic, M., Zahradka, K., and Zahradka, D. 2013. Exonuclease VII is involved in “reckless” DNA degradation in UV-irradiated Escherichia coli. Mutat. Res. 750, 96–104.
Sherratt, D.J. 1995 Mobile genetic elements. IRL Press at Oxford University Press, Oxford.
Vales, L.D., Rabin, B.A., and Chase, J.W. 1982. Subunit structure of Escherichia coli exonuclease VII. J. Biol. Chem. 257, 8799–8805.
Viswanathan, M. and Lovett, S.T. 1998. Single-strand DNA-specific exonucleases in Escherichia coli. Roles in repair and mutation avoidance. Genetics 149, 7–16.
Wadhawan, S., Gautam, S., and Sharma, A. 2013. A component of gamma-radiation-induced cell death in E. coli is programmed and interlinked with activation of caspase-3 and SOS response. Arch. Microbiol. 195, 545–557.
Wild, J., Hradecna, Z., and Szybalski, W. 2002. Conditionally amplifiable BACs: switching from single-copy to high-copy vectors and genomic clones. Genome Res. 12, 1434–1444.
Wild, J. and Szybalski, W. 2004. Copy-control tightly regulated expression vectors based on pBAC/oriV. Methods Mol. Biol. 267, 155–167.
Zahradka, K., Buljubasic, M., Petranovic, M., and Zahradka, D. 2009. Roles of ExoI and SbcCD nucleases in “reckless” DNA degradation in recA mutants of Escherichia coli. J. Bacteriol. 191, 1677–1687.
Author information
Authors and Affiliations
Corresponding author
Additional information
Supplemental material for this article may be found at http://www.springerlink.com/content/120956.
Electronic supplementary material
Rights and permissions
About this article
Cite this article
Jung, H., Liang, J., Jung, Y. et al. Characterization of cell death in Escherichia coli mediated by XseA, a large subunit of exonuclease VII. J Microbiol. 53, 820–828 (2015). https://doi.org/10.1007/s12275-015-5304-0
Received:
Revised:
Accepted:
Published:
Issue Date:
DOI: https://doi.org/10.1007/s12275-015-5304-0