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Regulation of gene expression at the transcriptional level is achieved by complex interactions of transcription factors operating at their target genes. Dissecting the specific combination of factors that bind each target is a significant challenge. Here, we describe in detail the Allele Binding Cooperativity test, which uses variation in transcription factor binding among individuals to discover combinations of factors and their targets. We developed the ALPHABIT (a large-scale process to hunt for allele binding interacting transcription factors) pipeline, which includes statistical analysis of binding sites followed by experimental validation, and demonstrate that this method predicts transcription factors that associate with NFκB. Our method successfully identifies factors that have been known to work with NFκB (E2A, STAT1, IRF2), but whose global coassociation and sites of cooperative action were not known. In addition, we identify a unique coassociation (EBF1) that had not been reported previously. We present a general approach for discovering combinatorial models of regulation and advance our understanding of the genetic basis of variation in transcription factor binding. [ABSTRACT FROM AUTHOR]

Small noncoding regulatory RNAs (sRNAs) play a key role in the posttranscriptional regulation of many bacterial genes. The genome of Caulobacter crescentus encodes at least 31 sRNAs, and 27 of these sRNAs are of unknown function. An overexpression screen for sRNA-induced growth inhibition along with sequence conservation in a related Caulobacter species led to the identification of a novel sRNA, CrfA, that is specifically induced upon carbon starvation. Twenty-seven genes were found to be strongly activated by CrfA accumulation. One-third of these target genes encode putative TonB-dependent receptors, suggesting CrfA plays a role in the surface modification of C. crescentus, facilitating the uptake of nutrients during periods of carbon starvation. The mechanism of CrfA-mediated gene activation was investigated for one of the genes predicted to encode a TonB-dependent receptor, CC3461. CrfA functions to stabilize the CC3461 transcript. Complementarity between a region of CrfA and the terminal region of the CC3461 5'-untranslated region (5'-UTR) and also the behavior of a deletion of this region and a site-specific base substitution and a 3-base deletion in the CrfA complementary sequence suggest that CrfA binds to a stem-loop structure upstream of the CC3461 Shine-Dalgarno sequence and stabilizes the transcript. [ABSTRACT FROM AUTHOR]