Expression of the Bacillus subtilis trpEDCFBA operon is influenced by translational coupling and Rho termination factor

被引:41
作者
Yakhnin, H [1 ]
Babiarz, JE [1 ]
Yakhnin, AV [1 ]
Babitzke, P [1 ]
机构
[1] Penn State Univ, Dept Biochem & Mol Biol, University Pk, PA 16802 USA
关键词
D O I
10.1128/JB.183.20.5918-5926.2001
中图分类号
Q93 [微生物学];
学科分类号
071005 ; 100705 ;
摘要
The trp RNA-binding attenuation protein (TRAP) regulates expression of the Bacillus subtilis trpEDCFBA operon by transcription attenuation and translational control mechanisms. Both mechanisms require binding of tryptophan-activated TRAP to 11 (G/U)AG repeats in the tip leader transcript. trpE translational control involves formation of a TRAP-dependent RNA structure that sequesters the trpE Shine-Dalgarno (SD) sequence (the SD blocking hairpin). By comparing expression levels from trpE'-'lacZ translational fusions controlled by the wild-type leader or by a leader that cannot form the SD blocking hairpin, we found that translational control requires a tryptophan concentration higher than that required for transcription attenuation. We also found that inhibition of trpE translation by the SD blocking hairpin does not alter the stability of the downstream message. Since the coding sequences for trpE and trpD overlap by 29 nucleotides, we examined expression levels from trpED'-'lacZ translational fusions to determine if these two genes are translationally coupled. We found that introduction of a UAA stop codon in trpE resulted in a substantial reduction in expression. Since expression was partially restored in the presence of a tRNA suppressor, our results indicate that trpE and trpD are translationally coupled. We determined that the coupling mechanism is TRAP independent and that formation of the SD blocking hairpin regulates trpD translation via translational coupling. We also constructed a rho mutation to investigate the role of Rho-dependent termination in trp operon expression. We found that TRAP-dependent formation of the SD blocking hairpin allows Rho access to the nascent transcript, causing transcriptional polarity.
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页码:5918 / 5926
页数:9
相关论文
共 33 条
[1]   REQUIREMENTS FOR TRANSFORMATION IN BACILLUS SUBTILIS [J].
ANAGNOSTOPOULOS, C ;
SPIZIZEN, J .
JOURNAL OF BACTERIOLOGY, 1961, 81 (05) :741-&
[2]   Structure of the trp RNA-binding attenuation protein, TRAP, bound to RNA [J].
Antson, AA ;
Dodson, EJ ;
Dodson, G ;
Greaves, RB ;
Chen, XP ;
Gollnick, P .
NATURE, 1999, 401 (6750) :235-242
[3]   THE STRUCTURE OF TRP RNA-BINDING ATTENUATION PROTEIN [J].
ANTSON, AA ;
OTRIDGE, J ;
BRZOZOWSKI, AM ;
DODSON, EJ ;
DODSON, GG ;
WILSON, KS ;
SMITH, TM ;
YANG, M ;
KURECKI, T ;
GOLLNICK, P .
NATURE, 1995, 374 (6524) :693-700
[4]   STRUCTURAL FEATURES OF L-TRYPTOPHAN REQUIRED FOR ACTIVATION OF TRAP, THE TRP RNA-BINDING ATTENUATION PROTEIN OF BACILLUS-SUBTILIS [J].
BABITZKE, P ;
YANOFSKY, C .
JOURNAL OF BIOLOGICAL CHEMISTRY, 1995, 270 (21) :12452-12456
[5]   RECONSTITUTION OF BACILLUS-SUBTILIS TRP ATTENUATION INVITRO WITH TRAP, THE TRP RNA-BINDING ATTENUATION PROTEIN [J].
BABITZKE, P ;
YANOFSKY, C .
PROCEEDINGS OF THE NATIONAL ACADEMY OF SCIENCES OF THE UNITED STATES OF AMERICA, 1993, 90 (01) :133-137
[6]  
BABITZKE P, 1994, J BIOL CHEM, V269, P16597
[7]   Regulation of tryptophan biosynthesis: Trp-ing the TRAP or how Bacillus subtilis reinvented the wheel [J].
Babitzke, P .
MOLECULAR MICROBIOLOGY, 1997, 26 (01) :1-9
[8]   Kinetic and thermodynamic analysis of the interaction between TRAP (trp RNA-binding attenuation protein) of Bacillus subtilis and trp leader RNA [J].
Baumann, C ;
Otridge, J ;
Gollnick, P .
JOURNAL OF BIOLOGICAL CHEMISTRY, 1996, 271 (21) :12269-12274
[9]   GENOMIC SEQUENCING [J].
CHURCH, GM ;
GILBERT, W .
PROCEEDINGS OF THE NATIONAL ACADEMY OF SCIENCES OF THE UNITED STATES OF AMERICA-BIOLOGICAL SCIENCES, 1984, 81 (07) :1991-1995
[10]   trp RNA-binding attenuation protein-5′ stem-loop RNA interaction is required for proper transcription attenuation control of the Bacillus subtilis trpEDCFBA operon [J].
Du, H ;
Yakhnin, AV ;
Dharmaraj, S ;
Babitzke, P .
JOURNAL OF BACTERIOLOGY, 2000, 182 (07) :1819-1827