MULTIPLE MECHANISMS CONTRIBUTE TO OSMOTIC INDUCIBILITY OF PROU OPERON EXPRESSION IN ESCHERICHIA-COLI - DEMONSTRATION OF 2 OSMORESPONSIVE PROMOTERS AND OF A NEGATIVE REGULATORY ELEMENT WITHIN THE 1ST STRUCTURAL GENE

被引:66
作者
DATTANANDA, CS [1 ]
RAJKUMARI, K [1 ]
GOWRISHANKAR, J [1 ]
机构
[1] CTR CELLULAR & MOLEC BIOL,HYDERABAD 500007,INDIA
关键词
D O I
10.1128/jb.173.23.7481-7490.1991
中图分类号
Q93 [微生物学];
学科分类号
071005 ; 100705 ;
摘要
Transcription of the proU operon in Escherichia coli is induced several hundredfold upon growth of cells in media of elevated osmolarity. A low-copy-number promoter-cloning plasmid vector, with lacZ as the reporter gene, was used for assaying the osmoresponsive promoter activity of each of various lengths of proU DNA, generated by cloning of discrete restriction fragments and by an exonuclease III-mediated deletion approach. The results indicate that expression of proU in E. coli is directed from two promoters, one (P2) characterized earlier by other workers with the start site of transcription 60 nucleotides upstream of the initiation codon of the first structural gene (proV), and the other (P1) situated 250 nucleotides upstream of proV. Furthermore, a region of DNA within proV was shown to be involved in negative regulation of proU transcription; phage Mu dII1681-generated lac fusions in the early region of proV also exhibited partial derepression of proU regulation, in comparison with fusions further downstream in the operon. Sequences around promoter P1, sequences around P2, and the promoter-downstream negative regulatory element, respectively, conferred approximately 5-, 8-, and 25-fold osmoresponsivity on proU expression. Within the region genetically defined to encode the negative regulatory element, there is a 116-nucleotide stretch that is absolutely conserved between the proU operons of E. coli and Salmonella typhimurium and has the capability of exhibiting alternative secondary structure. Insertion of this region of DNA into each of two different plasmid vectors was associated with a marked reduction in the mean topological linking number in plasmid molecules isolated from cultures grown in high-osmolarity medium. We propose that this region of DNA undergoes reversible transition to an underwound DNA conformation under high-osmolarity growth conditions and that this transition mediates its regulatory effect on proU expression.
引用
收藏
页码:7481 / 7490
页数:10
相关论文
共 48 条
  • [21] HISTONE-LIKE PROTEIN H1 (H-NS), DNA SUPERCOILING, AND GENE-EXPRESSION IN BACTERIA
    HULTON, CSJ
    SEIRAFI, A
    HINTON, JCD
    SIDEBOTHAM, JM
    WADDELL, L
    PAVITT, GD
    OWENHUGHES, T
    SPASSKY, A
    BUC, H
    HIGGINS, CF
    [J]. CELL, 1990, 63 (03) : 631 - 642
  • [22] SOME SIMPLE COMPUTATIONAL METHODS TO IMPROVE THE FOLDING OF LARGE RNAS
    JACOBSON, AB
    GOOD, L
    SIMONETTI, J
    ZUKER, M
    [J]. NUCLEIC ACIDS RESEARCH, 1984, 12 (01) : 45 - 52
  • [23] JAWORSKI A, 1991, J BIOL CHEM, V266, P2576
  • [24] JAYASHREE P, UNPUB
  • [25] JOVANOVICH SB, 1989, J BIOL CHEM, V264, P7821
  • [26] RAPID SITE-SPECIFIC DNA INVERSION IN ESCHERICHIA-COLI MUTANTS LACKING THE HISTONELIKE PROTEIN H-NS
    KAWULA, TH
    ORNDORFF, PE
    [J]. JOURNAL OF BACTERIOLOGY, 1991, 173 (13) : 4116 - 4123
  • [27] CHARACTERIZATION OF MUTATIONS AFFECTING THE OSMOREGULATED PROU PROMOTER OF ESCHERICHIA-COLI AND IDENTIFICATION OF 5' SEQUENCES REQUIRED FOR HIGH-LEVEL EXPRESSION
    LUCHT, JM
    BREMER, E
    [J]. JOURNAL OF BACTERIOLOGY, 1991, 173 (02) : 801 - 809
  • [28] PTAC-85, AN ESCHERICHIA-COLI VECTOR FOR EXPRESSION OF NON-FUSION PROTEINS
    MARSH, P
    [J]. NUCLEIC ACIDS RESEARCH, 1986, 14 (08) : 3603 - 3603
  • [29] THE OSMZ (BGLY) GENE ENCODES THE DNA-BINDING PROTEIN H-NS (H1A), A COMPONENT OF THE ESCHERICHIA-COLI K12 NUCLEOID
    MAY, G
    DERSCH, P
    HAARDT, M
    MIDDENDORF, A
    BREMER, E
    [J]. MOLECULAR & GENERAL GENETICS, 1990, 224 (01): : 81 - 90
  • [30] CHARACTERIZATION OF THE OSMOREGULATED ESCHERICHIA-COLI PROU PROMOTER AND IDENTIFICATION OF PROV AS A MEMBRANE-ASSOCIATED PROTEIN
    MAY, G
    FAATZ, E
    LUCHT, JM
    HAARDT, M
    BOLLIGER, M
    BREMER, E
    [J]. MOLECULAR MICROBIOLOGY, 1989, 3 (11) : 1521 - 1531