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Shin, Jongoh,Song, Yoseb,Jin, Sangrak,Lee, Jung-Kul,Kim, Dong Rip,Kim, Sun Chang,Cho, Suhyung,Cho, Byung-Kwan Cold Spring Harbor Laboratory Press 2018 RNA Vol.24 No.12
<P>Acetogens synthesize acetyl-CoA via CO<SUB>2</SUB> or CO fixation, producing organic compounds. Despite their ecological and industrial importance, their transcriptional and post-transcriptional regulation has not been systematically studied. With completion of the genome sequence of <I>Acetobacterium bakii</I> (4.28-Mb), we measured changes in the transcriptome of this psychrotolerant acetogen in response to temperature variations under autotrophic and heterotrophic growth conditions. Unexpectedly, acetogenesis genes were highly up-regulated at low temperatures under heterotrophic, as well as autotrophic, growth conditions. To mechanistically understand the transcriptional regulation of acetogenesis genes via changes in RNA secondary structures of 5′-untranslated regions (5′-UTR), the primary transcriptome was experimentally determined, and 1379 transcription start sites (TSS) and 1100 5′-UTR were found. Interestingly, acetogenesis genes contained longer 5′-UTR with lower RNA-folding free energy than other genes, revealing that the 5′-UTRs control the RNA abundance of the acetogenesis genes under low temperature conditions. Our findings suggest that post-transcriptional regulation via RNA conformational changes of 5′-UTRs is necessary for cold-adaptive acetogenesis.</P>
Applications of CRISPR/Cas System to Bacterial Metabolic Engineering
Cho, Suhyung,Shin, Jongoh,Cho, Byung-Kwan MDPI 2018 INTERNATIONAL JOURNAL OF MOLECULAR SCIENCES Vol.19 No.4
<P>The clustered regularly interspaced short palindromic repeats/CRISPR-associated (CRISPR/Cas) adaptive immune system has been extensively used for gene editing, including gene deletion, insertion, and replacement in bacterial and eukaryotic cells owing to its simple, rapid, and efficient activities in unprecedented resolution. Furthermore, the CRISPR interference (CRISPRi) system including deactivated Cas9 (dCas9) with inactivated endonuclease activity has been further investigated for regulation of the target gene transiently or constitutively, avoiding cell death by disruption of genome. This review discusses the applications of CRISPR/Cas for genome editing in various bacterial systems and their applications. In particular, CRISPR technology has been used for the production of metabolites of high industrial significance, including biochemical, biofuel, and pharmaceutical products/precursors in bacteria. Here, we focus on methods to increase the productivity and yield/titer scan by controlling metabolic flux through individual or combinatorial use of CRISPR/Cas and CRISPRi systems with introduction of synthetic pathway in industrially common bacteria including <I>Escherichia coli</I>. Further, we discuss additional useful applications of the CRISPR/Cas system, including its use in functional genomics.</P>