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Nickels, B.

Publications and source records attributed to Nickels, B..

2 recordsLinked to original sources

A role for GrgA in regulation of σ28-dependent transcription in the obligate intracellular bacterial pathogen Chlamydia trachomatis

The sexually transmitted obligate intracellular bacterial pathogen Chlamydia trachomatis has a unique developmental cycle consisting of two contrasting cellular forms. Whereas the primary Chlamydia sigma factor, {sigma}66, is involved in the expression of the majority of chlamydial genes throughout the developmental cycle, expression of several late genes requires the alternative sigma factor {sigma}28. In prior work we identified GrgA as a Chlamydia-specific transcription factor that activates {sigma}66-dependent transcription by binding DNA and interacting with a non-conserved region (NCR) of {sigma}66. Here, we extend these findings by showing GrgA can also activate {sigma}28-dependent transcription through direct interaction with {sigma}28. We measure the binding affinity of GrgA for both {sigma}66and {sigma}28, and we identify regions of GrgA important for {sigma}28-dependent transcription. Similar to results obtained with {sigma}66, we find that GrgAs interaction with {sigma}28 involves a NCR located upstream of conserved region 2 of {sigma}28. Our findings suggest GrgA is an important regulator of both {sigma}66- and {sigma}28-dependent transcription in C. trachomatis and further highlight NCRs of bacterial RNA polymerase as targets for regulatory factors unique to particular organisms.

microbiology

The mechanism of transcription start site selection

During transcription initiation, RNA polymerase (RNAP) binds to promoter DNA, unwinds promoter DNA to form an RNAP-promoter open complex (RPo) containing a single-stranded \"transcription bubble,\" and selects a transcription start site (TSS). TSS selection occurs at different positions within the promoter region, depending on promoter sequence and initiating-substrate concentration. Variability in TSS selection has been proposed to involve DNA \"scrunching\" and \"antiscrunching,\" the hallmarks of which are: (i) forward and reverse movement of the RNAP leading edge, but not trailing edge, relative to DNA, and (ii) expansion and contraction of the transcription bubble. Here, using in vitro and in vivo protein-DNA photocrosslinking and single-molecule nanomanipulation, we show bacterial TSS selection exhibits both hallmarks of scrunching and anti-scrunching, and we define energetics of scrunching and anti-scrunching. The results establish the mechanism of TSS selection by bacterial RNAP and suggest a general mechanism for TSS selection by bacterial, archaeal, and eukaryotic RNAP.

molecular biology