Neshat Armin, Mentz Almut, Rückert Christian, Kalinowski Jörn
Microbial Genomics and Biotechnology, Center for Biotechnology, Bielefeld University, Universitätsstraße 27, 33615 Bielefeld, Germany.
Microbial Genomics and Biotechnology, Center for Biotechnology, Bielefeld University, Universitätsstraße 27, 33615 Bielefeld, Germany; Technology Platform Genomics, Center for Biotechnology, Bielefeld University, Universitätsstraße 27, 33615 Bielefeld, Germany.
J Biotechnol. 2014 Nov 20;190:55-63. doi: 10.1016/j.jbiotec.2014.05.033. Epub 2014 Jun 6.
The Gram-positive bacterium Corynebacterium glutamicum belongs to the order Corynebacteriales and is used as a producer of amino acids at industrial scales. Due to its economic importance, gene expression and particularly the regulation of amino acid biosynthesis has been investigated extensively. Applying the high-resolution technique of transcriptome sequencing (RNA-seq), recently a vast amount of data has been generated that was used to comprehensively analyze the C. glutamicum transcriptome. By analyzing RNA-seq data from a small RNA cDNA library of C. glutamicum, short transcripts in the known transcriptional attenuators sites of the trp operon, the ilvBNC operon and the leuA gene were verified. Furthermore, whole transcriptome RNA-seq data were used to elucidate the transcriptional organization of these three amino acid biosynthesis operons. In addition, we discovered and analyzed the novel attenuator aroR, located upstream of the aroF gene (cg1129). The DAHP synthase encoded by aroF catalyzes the first step in aromatic amino acid synthesis. The AroR leader peptide contains the amino acid sequence motif F-Y-F, indicating a regulatory effect by phenylalanine and tyrosine. Analysis by real-time RT-PCR suggests that the attenuator regulates the transcription of aroF in dependence of the cellular amount of tRNA loaded with phenylalanine when comparing a phenylalanine-auxotrophic C. glutamicum mutant fed with limiting and excess amounts of a phenylalanine-containing dipeptide. Additionally, the very interesting finding was made that all analyzed attenuators are leaderless transcripts.
革兰氏阳性菌谷氨酸棒杆菌属于棒杆菌目，在工业规模上用作氨基酸生产菌。由于其经济重要性，人们对其基因表达，尤其是氨基酸生物合成的调控进行了广泛研究。应用转录组测序（RNA-seq）这一高分辨率技术，最近已产生了大量数据，用于全面分析谷氨酸棒杆菌的转录组。通过分析谷氨酸棒杆菌小RNA cDNA文库的RNA-seq数据，验证了色氨酸操纵子、ilvBNC操纵子和亮氨酸A基因已知转录衰减子位点处的短转录本。此外，利用全转录组RNA-seq数据阐明了这三个氨基酸生物合成操纵子的转录组织。另外，我们发现并分析了位于aroF基因（cg1129）上游的新型衰减子aroR。aroF编码的DAHP合酶催化芳香族氨基酸合成的第一步。AroR前导肽含有氨基酸序列基序F-Y-F，表明受苯丙氨酸和酪氨酸的调节作用。实时RT-PCR分析表明，在比较用限量和过量含苯丙氨酸二肽喂养的苯丙氨酸营养缺陷型谷氨酸棒杆菌突变体时，衰减子根据负载苯丙氨酸的tRNA的细胞量调节aroF的转录。此外，还发现了一个非常有趣的现象，即所有分析的衰减子都是无前导序列的转录本。
