Kormanec J, Homerová D, Potúcková L, Nováková R, Rezuchová B
Institute of Molecular Biology, Slovak Academy of Sciences, Bratislava, Slovak Republic.
Gene. 1996 Nov 28;181(1-2):19-27. doi: 10.1016/s0378-1119(96)00395-2.
In previous experiments, the Streptomyces aureofaciens (Sa) rpoZ, and sigF genes have been shown to encode putative sigma factors essential in differentiation. In an attempt to investigate expression of these genes during differentiation, we have performed S1 nuclease mapping using RNA prepared from Sa in various developmental stages. Two putative promoters were identified upstream of the rpoZ coding region. The promoters significantly differed in their strength, and were active in distinct developmental stages; the weaker, rpoZ-P1, was active only in substrate mycelium, and the stronger, rpoZ-P2, was induced at the beginning of aerial mycelium formation. Transcriptional analysis of sigF revealed two apparent transcription start points, both being detectable only in the late phase of aerial mycelium formation. No sigF transcription was detected in an rpoZ-disrupted Sa strain. Promoter-bearing DNA fragments from rpoZ and sigF were inserted into several promoter-probe vectors, to give expression patterns consistent with the results of direct RNA analysis. The results implicate temporally different expression of rpoZ and sigF during the differentiation of Sa, and direct or indirect dependence of sigF expression on the rpoZ-encoded putative sigma factor, thus indicating a cascade of sigma factors in Streptomyces development.
在先前的实验中，金色链霉菌（Sa）的rpoZ和sigF基因已被证明编码在分化过程中必不可少的假定σ因子。为了研究这些基因在分化过程中的表达情况，我们使用从处于不同发育阶段的Sa制备的RNA进行了S1核酸酶图谱分析。在rpoZ编码区上游鉴定出两个假定的启动子。这些启动子的强度有显著差异，且在不同的发育阶段具有活性；较弱的rpoZ - P1仅在基内菌丝中具有活性，而较强的rpoZ - P2在气生菌丝形成开始时被诱导。对sigF的转录分析揭示了两个明显的转录起始点，二者仅在气生菌丝形成的后期才能检测到。在rpoZ缺失的Sa菌株中未检测到sigF转录。将来自rpoZ和sigF的含启动子DNA片段插入到几个启动子探针载体中，得到与直接RNA分析结果一致的表达模式。结果表明，在Sa分化过程中rpoZ和sigF在时间上存在不同表达，且sigF表达直接或间接依赖于rpoZ编码的假定σ因子，从而表明链霉菌发育过程中存在σ因子级联反应。
