The trp RNA-binding attenuation protein of Bacillus subtilis regulates translation of the tryptophan transport gene trpP (yhaG) by blocking ribosome binding.
Expression of the Bacillus subtilis tryptophan biosynthetic genes (trpEDCFBA and pabA [trpG]) is regulated in response to tryptophan by TRAP, the trp RNA-binding attenuation protein. TRAP-mediated regulation of the tryptophan biosynthetic genes includes a transcription attenuation and two distinct translation control mechanisms. TRAP also regulates translation of trpP (yhaG), a single-gene operon that encodes a putative tryptophan transporter. Its translation initiation region contains triplet repeats typical of TRAP-regulated mRNAs. We found that regulation of trpP and pabA is unaltered in a rho mutant strain. Results from filter binding and gel mobility shift assays demonstrated that TRAP binds specifically to a segment of the trpP transcript that includes the untranslated leader and translation initiation region. While the affinities of TRAP for the trpP and pabA transcripts are similar, TRAP-mediated translation control of trpP is much more extensive than for pabA. RNA footprinting revealed that the trpP TRAP binding site consists of nine triplet repeats (five GAG, three UAG, and one AAG) that surround and overlap the trpP Shine-Dalgarno (S-D) sequence and translation start codon. Results from toeprint and RNA-directed cell-free translation experiments indicated that tryptophan-activated TRAP inhibits TrpP synthesis by preventing binding of a 30S ribosomal subunit. Taken together, our results establish that TRAP regulates translation of trpP by blocking ribosome binding. Thus, TRAP coordinately regulates tryptophan synthesis and transport by three distinct mechanisms: attenuation transcription of the trpEDCFBA operon, promoting formation of the trpE S-D blocking hairpin, and blocking ribosome binding to the pabA and trpP transcripts.
Yakhnin
Helen
H
Department of Biochemistry and Molecular Biology, The Pennsylvania State University, University Park, Pennsylvania 16802, USA.
Zhang
Hong
H
Yakhnin
Alexander V
AV
Babitzke
Paul
P
D026905:Amino Acid Transport Systems / Q000235:genetics*; D001412:Bacillus subtilis / Q000235:genetics*; D001426:Bacterial Proteins / Q000235:genetics* / Q000502:physiology*; D001483:Base Sequence; D019755:Carbon-Carbon Lyases / Q000235:genetics; D029968:Escherichia coli Proteins / Q000235:genetics; D008969:Molecular Sequence Data; D009876:Operon; D014176:Protein Biosynthesis; D012329:RNA, Bacterial / Q000737:chemistry / Q000378:metabolism; D016601:RNA-Binding Proteins / Q000502:physiology*; D012270:Ribosomes / Q000378:metabolism*; D000637:Transaminases; D014157:Transcription Factors / Q000502:physiology*; D018911:Trinucleotide Repeats; D014364:Tryptophan / Q000378:metabolism*
D026905:Amino Acid Transport Systems; D001426:Bacterial Proteins; D029968:Escherichia coli Proteins; C079092:MtrB protein, Bacteria; D012329:RNA, Bacterial; D016601:RNA-Binding Proteins; D014157:Transcription Factors; C406408:yhaG protein, Bacillus subtilis; D014364:Tryptophan; D000637:Transaminases; C000707653:aminodeoxychorismate synthase protein, E coli; D019755:Carbon-Carbon Lyases