Sequence-independent upstream DNA-alphaCTD interactions strongly stimulate Escherichia coli RNA polymerase-lacUV5 promoter association.
Sequence-independent upstream DNA-alphaCTD interactions strongly stimulate Escherichia coli RNA polymerase-lacUV5 promoter association.
复制标题
序列独立的上游 DNA-αCTD 相互作用强烈刺激大肠杆菌 RNA 聚合酶-lacUV5 启动子关联。
DOI：
10.1073/pnas.0405814102
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发表时间：
2005
影响因子：
11.1
通讯作者：
Gourse,RichardL
中科院分区：
文献类型：
--
作者：
Ross,Wilma;Gourse,RichardL
The C-terminal domains of the two α-subunits (αCTD) inEscherichia coliRNA polymerase (RNAP) recognize specific sequences called UP elements in some promoters. These interactions can increase transcription dramatically. Previously, effects of upstream DNA–αCTD interactions on transcription were quantified relative to control promoters with nonspecific DNA sequences substituted for UP elements. However, contributions of nonspecific upstream DNA–αCTD interactions to promoter activity have not been evaluated extensively. Here, we examine effects of removal of αCTD, upstream promoter DNA, or both on the rate of open-complex formation with promoters that lack UP elements. Deletion of αCTD decreased the composite second-order association rate constant,ka, of RNAP for thelacUV5promoter by ≈10-fold. Much of this effect was attributable to a decrease in the isomerization rate constant,k2. Removal of promoter DNA upstream of the -35 element also decreased bothkaandk2≈10-fold. Upstream DNA extending approximately to base pair -100 was sufficient for maximal association rates of wild-type RNAP withlacUV5promoter fragments. The αCTD and upstream DNA did not affect dissociation rates from the open complex. We suggest that sequence-independent upstream DNA interactions with αCTD are major contributors to initiation at many (or all) promoters (not merely promoters containing UP elements) and that these interactions facilitate isomerization events occurring well downstream of the α-binding sites. In addition to highlighting the functional importance of nonspecific protein–DNA interactions, these results suggest also that UP element–αCTD interactions play an even larger role in transcription initiation than appreciated previously.
