An integrative computational approach to effectively guide experimental identification of regulatory elements in promoters

Abstract

Background: Transcriptional activity of genes depends on many factors like DNA motifs, conformational characteristics of DNA, melting etc. and there are computational approaches for their identification. However, in real applications, the number of predicted, for example, DNA motifs may be considerably large. In cases when various computational programs are applied, systematic experimental knock out of each of the potential elements obviously becomes nonproductive. Hence, one needs an approach that is able to integrate many heterogeneous computational methods and upon that suggest selected regulatory elements for experimental verification.

Results: Here, we present an integrative bioinformatic approach aimed at the discovery of regulatory modules that can be effectively verified experimentally. It is based on combinatorial analysis of known and novel binding motifs, as well as of any other known features of promoters. The goal of this method is the identification of a collection of modules that are specific for an established dataset and at the same time are optimal for experimental verification. The method is particularly effective on small datasets, where most statistical approaches fail. We apply it to promoters that drive tumor-specific gene expression in tumor-colonizing Gram-negative bacteria. The method successfully identified a number of potential modules, which required only a few experiments to be verified. The resulting minimal functional bacterial promoter exhibited high specificity of expression in cancerous tissue.

Conclusions: Experimental analysis of promoter structures guided by bioinformatics has proved to be efficient. The developed computational method is able to include heterogeneous features of promoters and suggest combinatorial modules for experimental testing. Expansibility and robustness of the methodology implemented in the approach ensures good results for a wide range of problems.

Figure 1

Workflow diagrams for the experimental discovery of cis- regulatory modules. A) Conventional workflow used to identify regulatory modules in promoters. B) Workflow based exclusively on experimental research. C) Integrative approach for identification of regulatory modules.

Figure 2

Tumor specific regulatory module on the sequence of fragment 212_1. Schematic representation of the regulatory module on the DNA sequence of the fragment 212_1 (A). Expression of the GFP_OVA gene under control of the fragment 212_1 in tumor (B) and spleen (C).