Ambulos N P, Duvall E J, Lovett P S
J Bacteriol. 1987 Mar;169(3):967-72. doi: 10.1128/jb.169.3.967-972.1987.
cat-86 is a promoter-deficient plasmid gene that encodes chloramphenicol acetyltransferase (CAT). Insertion of a promoter at a site 144 base pairs 5' to the cat-86 coding sequence activates transcription of the gene and allows cat-86 to specify chloramphenicol-inducible CAT activity in Bacillus subtilis. Induction of cat-86 by chloramphenicol has been shown to result from a regulatory event that activates translation of cat-86 mRNA that is present in cells before the addition of inducer (E. J. Duvall and P. S. Lovett, Proc. Natl. Acad. Sci. USA 83:3939-3943, 1986). In the present study we show an unusual property of cat-86 mRNA. Full-length cat-86 transcripts, consisting of 920 nucleotides (nt), are cleaved in B. subtilis to yield two predominant fragmentation products: an 810-nt species that lacks sequences present at the 5' end of the 920-nt species and a 720-nt species that lacks sequences present at the 3' end of the 920-nt species. A third fragmentation product consisting of 620 nt may result from the cleavage of a single 920-nt transcript at both the 5' and 3' ends. The sequences which are missing from the 720- and 620-nt species suggest that these transcripts cannot be translated into functional CAT. The 810-nt species lacks sequences from the 5' regulatory region, and it is not yet certain whether or not translation of this species can be induced by chloramphenicol. The ratio of 920-nt molecules/720-nt molecules in rifampin-treated cells is increased when the cells are grown in chloramphenicol. Therefore, induction may partially stabilize full-length cat-86 transcripts against inactivation by a novel processing-like system.
cat - 86是一种启动子缺陷型质粒基因,编码氯霉素乙酰转移酶(CAT)。在cat - 86编码序列5'端144个碱基对处插入一个启动子可激活该基因的转录,并使cat - 86在枯草芽孢杆菌中产生氯霉素诱导型CAT活性。已证明氯霉素对cat - 86的诱导是由一种调节事件导致的,该事件激活了在添加诱导剂之前细胞中就已存在的cat - 86 mRNA的翻译(E. J. 杜瓦尔和P. S. 洛维特,《美国国家科学院院刊》83:3939 - 3943,1986)。在本研究中,我们展示了cat - 86 mRNA的一个不寻常特性。由920个核苷酸(nt)组成的全长cat - 86转录本在枯草芽孢杆菌中被切割,产生两种主要的片段化产物:一种810 - nt的物种,其缺少920 - nt物种5'端的序列;另一种720 - nt的物种,其缺少920 - nt物种3'端的序列。由620 nt组成的第三种片段化产物可能是单个920 - nt转录本在5'端和3'端都被切割的结果。720 - nt和620 - nt物种中缺失的序列表明这些转录本不能被翻译成有功能的CAT。810 - nt物种缺少5'调节区域的序列,目前尚不确定该物种的翻译是否能被氯霉素诱导。当细胞在氯霉素中生长时,利福平处理的细胞中920 - nt分子/720 - nt分子的比例会增加。因此,诱导可能会部分稳定全长cat - 86转录本,使其免受一种类似新加工系统的失活作用。