Skip to main content
PMC home page
Journal of Bacteriology logoLink to Journal of Bacteriology
. 1996 Jun;178(12):3572–3577. doi: 10.1128/jb.178.12.3572-3577.1996

Evidence that the hanA gene coding for HU protein is essential for heterocyst differentiation in, and cyanophage A-4(L) sensitivity of, Anabaena sp. strain PCC 7120.

I Khudyakov 1, C P Wolk 1
PMCID: PMC178128  PMID: 8655556

Abstract

The highly pleiotropic, transposon-generated mutant AB22 of Anabaena sp. strain PCC 7120 exhibits slow growth, altered pigmentation, cellular fragility, resistance to phage A-4(L), and the inability to differentiate heterocysts. Reconstruction of the transposon mutation in the wild-type strain reproduced the phenotype of the original mutant. Sequencing of the flanking DNA showed that the transposon had inserted at the beginning of a gene, which we call hanA, that encodes Anabaena HU protein (R. Nagaraja and R. Haselkorn, Biochimie 76:1082-1089, 1994). Mapping of the transposon insertion by pulsed-field gel electrophoresis showed that hanA is located at ca. 4.76 Mb on the physical map of the chromosome and is transcribed clockwise. Repeated subculturing of AB22 resulted in improved growth and loss of filament fragmentation, presumably because of one or more compensatory mutations; however, the mutant retained its A-4(L)r Het- phenotype. The mutation in strain AB22 could be complemented by a fragment of wild-type DNA bearing hanA as its only open reading frame.

Full Text

The Full Text of this article is available as a PDF (270.2 KB).

Selected References

These references are in PubMed. This may not be the complete list of references from this article.

  1. Allen M. B., Arnon D. I. Studies on Nitrogen-Fixing Blue-Green Algae. I. Growth and Nitrogen Fixation by Anabaena Cylindrica Lemm. Plant Physiol. 1955 Jul;30(4):366–372. doi: 10.1104/pp.30.4.366. [DOI] [PMC free article] [PubMed] [Google Scholar]
  2. Altschul S. F., Gish W., Miller W., Myers E. W., Lipman D. J. Basic local alignment search tool. J Mol Biol. 1990 Oct 5;215(3):403–410. doi: 10.1016/S0022-2836(05)80360-2. [DOI] [PubMed] [Google Scholar]
  3. Bianchi M. E. Prokaryotic HU and eukaryotic HMG1: a kinked relationship. Mol Microbiol. 1994 Oct;14(1):1–5. doi: 10.1111/j.1365-2958.1994.tb01261.x. [DOI] [PubMed] [Google Scholar]
  4. Black T. A., Cai Y., Wolk C. P. Spatial expression and autoregulation of hetR, a gene involved in the control of heterocyst development in Anabaena. Mol Microbiol. 1993 Jul;9(1):77–84. doi: 10.1111/j.1365-2958.1993.tb01670.x. [DOI] [PubMed] [Google Scholar]
  5. Black T. A., Wolk C. P. Analysis of a Het- mutation in Anabaena sp. strain PCC 7120 implicates a secondary metabolite in the regulation of heterocyst spacing. J Bacteriol. 1994 Apr;176(8):2282–2292. doi: 10.1128/jb.176.8.2282-2292.1994. [DOI] [PMC free article] [PubMed] [Google Scholar]
  6. Borthakur D., Haselkorn R. Tn5 mutagenesis of Anabaena sp. strain PCC 7120: isolation of a new mutant unable to grow without combined nitrogen. J Bacteriol. 1989 Oct;171(10):5759–5761. doi: 10.1128/jb.171.10.5759-5761.1989. [DOI] [PMC free article] [PubMed] [Google Scholar]
  7. Buikema W. J., Haselkorn R. Characterization of a gene controlling heterocyst differentiation in the cyanobacterium Anabaena 7120. Genes Dev. 1991 Feb;5(2):321–330. doi: 10.1101/gad.5.2.321. [DOI] [PubMed] [Google Scholar]
  8. Buikema W. J., Haselkorn R. Isolation and complementation of nitrogen fixation mutants of the cyanobacterium Anabaena sp. strain PCC 7120. J Bacteriol. 1991 Mar;173(6):1879–1885. doi: 10.1128/jb.173.6.1879-1885.1991. [DOI] [PMC free article] [PubMed] [Google Scholar]
  9. Cai Y. P., Wolk C. P. Use of a conditionally lethal gene in Anabaena sp. strain PCC 7120 to select for double recombinants and to entrap insertion sequences. J Bacteriol. 1990 Jun;172(6):3138–3145. doi: 10.1128/jb.172.6.3138-3145.1990. [DOI] [PMC free article] [PubMed] [Google Scholar]
  10. Carrasco C. D., Buettner J. A., Golden J. W. Programmed DNA rearrangement of a cyanobacterial hupL gene in heterocysts. Proc Natl Acad Sci U S A. 1995 Jan 31;92(3):791–795. doi: 10.1073/pnas.92.3.791. [DOI] [PMC free article] [PubMed] [Google Scholar]
  11. Drlica K., Rouviere-Yaniv J. Histonelike proteins of bacteria. Microbiol Rev. 1987 Sep;51(3):301–319. doi: 10.1128/mr.51.3.301-319.1987. [DOI] [PMC free article] [PubMed] [Google Scholar]
  12. Elhai J., Wolk C. P. Conjugal transfer of DNA to cyanobacteria. Methods Enzymol. 1988;167:747–754. doi: 10.1016/0076-6879(88)67086-8. [DOI] [PubMed] [Google Scholar]
  13. Elhai J., Wolk C. P. Developmental regulation and spatial pattern of expression of the structural genes for nitrogenase in the cyanobacterium Anabaena. EMBO J. 1990 Oct;9(10):3379–3388. doi: 10.1002/j.1460-2075.1990.tb07539.x. [DOI] [PMC free article] [PubMed] [Google Scholar]
  14. Ernst A., Black T., Cai Y., Panoff J. M., Tiwari D. N., Wolk C. P. Synthesis of nitrogenase in mutants of the cyanobacterium Anabaena sp. strain PCC 7120 affected in heterocyst development or metabolism. J Bacteriol. 1992 Oct;174(19):6025–6032. doi: 10.1128/jb.174.19.6025-6032.1992. [DOI] [PMC free article] [PubMed] [Google Scholar]
  15. Fernández-Piñas F., Leganés F., Wolk C. P. A third genetic locus required for the formation of heterocysts in Anabaena sp. strain PCC 7120. J Bacteriol. 1994 Sep;176(17):5277–5283. doi: 10.1128/jb.176.17.5277-5283.1994. [DOI] [PMC free article] [PubMed] [Google Scholar]
  16. Fernández-Piñas F., Wolk C. P. Expression of luxCD-E in Anabaena sp. can replace the use of exogenous aldehyde for in vivo localization of transcription by luxAB. Gene. 1994 Dec 2;150(1):169–174. doi: 10.1016/0378-1119(94)90879-6. [DOI] [PubMed] [Google Scholar]
  17. Fleming H., Haselkorn R. The program of protein synthesis during heterocyst differentiation in nitrogen-fixing blue-green algae. Cell. 1974 Oct;3(2):169–170. doi: 10.1016/0092-8674(74)90121-4. [DOI] [PubMed] [Google Scholar]
  18. Frías J. E., Flores E., Herrero A. Requirement of the regulatory protein NtcA for the expression of nitrogen assimilation and heterocyst development genes in the cyanobacterium Anabaena sp. PCC 7120. Mol Microbiol. 1994 Nov;14(4):823–832. doi: 10.1111/j.1365-2958.1994.tb01318.x. [DOI] [PubMed] [Google Scholar]
  19. Frías J. E., Mérida A., Herrero A., Martín-Nieto J., Flores E. General distribution of the nitrogen control gene ntcA in cyanobacteria. J Bacteriol. 1993 Sep;175(17):5710–5713. doi: 10.1128/jb.175.17.5710-5713.1993. [DOI] [PMC free article] [PubMed] [Google Scholar]
  20. Fürste J. P., Pansegrau W., Frank R., Blöcker H., Scholz P., Bagdasarian M., Lanka E. Molecular cloning of the plasmid RP4 primase region in a multi-host-range tacP expression vector. Gene. 1986;48(1):119–131. doi: 10.1016/0378-1119(86)90358-6. [DOI] [PubMed] [Google Scholar]
  21. Greene J. R., Brennan S. M., Andrew D. J., Thompson C. C., Richards S. H., Heinrikson R. L., Geiduschek E. P. Sequence of the bacteriophage SP01 gene coding for transcription factor 1, a viral homologue of the bacterial type II DNA-binding proteins. Proc Natl Acad Sci U S A. 1984 Nov;81(22):7031–7035. doi: 10.1073/pnas.81.22.7031. [DOI] [PMC free article] [PubMed] [Google Scholar]
  22. Grisolia V., Carlomagno M. S., Nappo A. G., Bruni C. B. Cloning, structure, and expression of the Escherichia coli K-12 hisC gene. J Bacteriol. 1985 Dec;164(3):1317–1323. doi: 10.1128/jb.164.3.1317-1323.1985. [DOI] [PMC free article] [PubMed] [Google Scholar]
  23. Haselkorn R. Developmentally regulated gene rearrangements in prokaryotes. Annu Rev Genet. 1992;26:113–130. doi: 10.1146/annurev.ge.26.120192.000553. [DOI] [PubMed] [Google Scholar]
  24. Henner D. J., Band L., Flaggs G., Chen E. The organization and nucleotide sequence of the Bacillus subtilis hisH, tyrA and aroE genes. Gene. 1986;49(1):147–152. doi: 10.1016/0378-1119(86)90394-x. [DOI] [PubMed] [Google Scholar]
  25. Higgins C. F., Hinton J. C., Hulton C. S., Owen-Hughes T., Pavitt G. D., Seirafi A. Protein H1: a role for chromatin structure in the regulation of bacterial gene expression and virulence? Mol Microbiol. 1990 Dec;4(12):2007–2012. doi: 10.1111/j.1365-2958.1990.tb00559.x. [DOI] [PubMed] [Google Scholar]
  26. Hillyard D. R., Edlund M., Hughes K. T., Marsh M., Higgins N. P. Subunit-specific phenotypes of Salmonella typhimurium HU mutants. J Bacteriol. 1990 Sep;172(9):5402–5407. doi: 10.1128/jb.172.9.5402-5407.1990. [DOI] [PMC free article] [PubMed] [Google Scholar]
  27. Hu N. T., Thiel T., Giddings T. H., Jr, Wolk C. P. New Anabaena and Nostoc cyanophages from sewage settling ponds. Virology. 1981 Oct 15;114(1):236–246. doi: 10.1016/0042-6822(81)90269-5. [DOI] [PubMed] [Google Scholar]
  28. Huisman O., Faelen M., Girard D., Jaffé A., Toussaint A., Rouvière-Yaniv J. Multiple defects in Escherichia coli mutants lacking HU protein. J Bacteriol. 1989 Jul;171(7):3704–3712. doi: 10.1128/jb.171.7.3704-3712.1989. [DOI] [PMC free article] [PubMed] [Google Scholar]
  29. Kano Y., Goshima N., Wada M., Imamoto F. Participation of hup gene product in replicative transposition of Mu phage in Escherichia coli. Gene. 1989;76(2):353–358. doi: 10.1016/0378-1119(89)90175-3. [DOI] [PubMed] [Google Scholar]
  30. Khudiakov I. Ia, Gromov B. V. Umerennyi tsianofag A-4 (L) cine-zelenoi vodorosli Anabaena variabilis. Mikrobiologiia. 1973 Sep-Oct;42(5):904–907. [PubMed] [Google Scholar]
  31. Koz'iakov S. Ia, Gromov B. V., Khudiakov I. Ia. A-1 (L)-tsianofag sinezelenoi vodorosli Anabaena variabilis. Mikrobiologiia. 1972 May-Jun;41(3):555–559. [PubMed] [Google Scholar]
  32. Kuritz T., Ernst A., Black T. A., Wolk C. P. High-resolution mapping of genetic loci of Anabaena PCC 7120 required for photosynthesis and nitrogen fixation. Mol Microbiol. 1993 Apr;8(1):101–110. doi: 10.1111/j.1365-2958.1993.tb01207.x. [DOI] [PubMed] [Google Scholar]
  33. Lavoie B. D., Chaconas G. Site-specific HU binding in the Mu transpososome: conversion of a sequence-independent DNA-binding protein into a chemical nuclease. Genes Dev. 1993 Dec;7(12B):2510–2519. doi: 10.1101/gad.7.12b.2510. [DOI] [PubMed] [Google Scholar]
  34. Leganés F., Fernández-Piñas F., Wolk C. P. Two mutations that block heterocyst differentiation have different effects on akinete differentiation in Nostoc ellipsosporum. Mol Microbiol. 1994 May;12(4):679–684. doi: 10.1111/j.1365-2958.1994.tb01055.x. [DOI] [PubMed] [Google Scholar]
  35. Liang J., Scappino L., Haselkorn R. The patA gene product, which contains a region similar to CheY of Escherichia coli, controls heterocyst pattern formation in the cyanobacterium Anabaena 7120. Proc Natl Acad Sci U S A. 1992 Jun 15;89(12):5655–5659. doi: 10.1073/pnas.89.12.5655. [DOI] [PMC free article] [PubMed] [Google Scholar]
  36. Liang J., Scappino L., Haselkorn R. The patB gene product, required for growth of the cyanobacterium Anabaena sp. strain PCC 7120 under nitrogen-limiting conditions, contains ferredoxin and helix-turn-helix domains. J Bacteriol. 1993 Mar;175(6):1697–1704. doi: 10.1128/jb.175.6.1697-1704.1993. [DOI] [PMC free article] [PubMed] [Google Scholar]
  37. Luque I., Flores E., Herrero A. Molecular mechanism for the operation of nitrogen control in cyanobacteria. EMBO J. 1994 Jun 15;13(12):2862–2869. doi: 10.1002/j.1460-2075.1994.tb06580.x. [DOI] [PMC free article] [PubMed] [Google Scholar]
  38. Lynn M. E., Bantle J. A., Ownby J. D. Estimation of gene expression in heterocysts of Anabaena variabilis by using DNA-RNA hybridization. J Bacteriol. 1986 Sep;167(3):940–946. doi: 10.1128/jb.167.3.940-946.1986. [DOI] [PMC free article] [PubMed] [Google Scholar]
  39. Nagaraja R., Haselkorn R. Protein HU from the cyanobacterium Anabaena. Biochimie. 1994;76(10-11):1082–1089. doi: 10.1016/0300-9084(94)90034-5. [DOI] [PubMed] [Google Scholar]
  40. Pridmore R. D. New and versatile cloning vectors with kanamycin-resistance marker. Gene. 1987;56(2-3):309–312. doi: 10.1016/0378-1119(87)90149-1. [DOI] [PubMed] [Google Scholar]
  41. Pruss G. J., Drlica K. DNA supercoiling and prokaryotic transcription. Cell. 1989 Feb 24;56(4):521–523. doi: 10.1016/0092-8674(89)90574-6. [DOI] [PubMed] [Google Scholar]
  42. Ramasubramanian T. S., Wei T. F., Golden J. W. Two Anabaena sp. strain PCC 7120 DNA-binding factors interact with vegetative cell- and heterocyst-specific genes. J Bacteriol. 1994 Mar;176(5):1214–1223. doi: 10.1128/jb.176.5.1214-1223.1994. [DOI] [PMC free article] [PubMed] [Google Scholar]
  43. Rothfield L. I. Bacterial chromosome segregation. Cell. 1994 Jul 1;77(7):963–966. doi: 10.1016/0092-8674(94)90435-9. [DOI] [PubMed] [Google Scholar]
  44. Schmid M. B. More than just "histone-like" proteins. Cell. 1990 Nov 2;63(3):451–453. doi: 10.1016/0092-8674(90)90438-k. [DOI] [PubMed] [Google Scholar]
  45. Schneider G. J., Lang J. D., Haselkorn R. Promoter recognition by the RNA polymerase from vegetative cells of the cyanobacterium Anabaena 7120. Gene. 1991 Aug 30;105(1):51–60. doi: 10.1016/0378-1119(91)90513-b. [DOI] [PubMed] [Google Scholar]
  46. Steck T. R., Drlica K. Bacterial chromosome segregation: evidence for DNA gyrase involvement in decatenation. Cell. 1984 Apr;36(4):1081–1088. doi: 10.1016/0092-8674(84)90058-8. [DOI] [PubMed] [Google Scholar]
  47. Travers A. A., Ner S. S., Churchill M. E. DNA chaperones: a solution to a persistence problem? Cell. 1994 Apr 22;77(2):167–169. doi: 10.1016/0092-8674(94)90306-9. [DOI] [PubMed] [Google Scholar]
  48. Vieira J., Messing J. The pUC plasmids, an M13mp7-derived system for insertion mutagenesis and sequencing with synthetic universal primers. Gene. 1982 Oct;19(3):259–268. doi: 10.1016/0378-1119(82)90015-4. [DOI] [PubMed] [Google Scholar]
  49. Wada M., Kano Y., Ogawa T., Okazaki T., Imamoto F. Construction and characterization of the deletion mutant of hupA and hupB genes in Escherichia coli. J Mol Biol. 1988 Dec 5;204(3):581–591. doi: 10.1016/0022-2836(88)90357-9. [DOI] [PubMed] [Google Scholar]
  50. Wei T. F., Ramasubramanian T. S., Golden J. W. Anabaena sp. strain PCC 7120 ntcA gene required for growth on nitrate and heterocyst development. J Bacteriol. 1994 Aug;176(15):4473–4482. doi: 10.1128/jb.176.15.4473-4482.1994. [DOI] [PMC free article] [PubMed] [Google Scholar]
  51. Wolk C. P., Cai Y., Panoff J. M. Use of a transposon with luciferase as a reporter to identify environmentally responsive genes in a cyanobacterium. Proc Natl Acad Sci U S A. 1991 Jun 15;88(12):5355–5359. doi: 10.1073/pnas.88.12.5355. [DOI] [PMC free article] [PubMed] [Google Scholar]
  52. Wolk C. P., Vonshak A., Kehoe P., Elhai J. Construction of shuttle vectors capable of conjugative transfer from Escherichia coli to nitrogen-fixing filamentous cyanobacteria. Proc Natl Acad Sci U S A. 1984 Mar;81(5):1561–1565. doi: 10.1073/pnas.81.5.1561. [DOI] [PMC free article] [PubMed] [Google Scholar]

Articles from Journal of Bacteriology are provided here courtesy of American Society for Microbiology (ASM)

RESOURCES