Background VirtUaL ChIP-seq Analysis through Networks (VULCAN) infers regulatory interactions of transcription factors by overlaying networks generated from publicly available tumor expression data onto ChIP-seq data. We apply our method to dissect the regulation of estrogen receptor-alpha activation in breast cancer to identify potential co-regulators of the estrogen receptor’s transcriptional response. Results VULCAN analysis of estrogen receptor activation in breast cancer highlights the key components of the estrogen receptor complex alongside a novel interaction with GRHL2. We demonstrate that GRHL2 is recruited to a subset of estrogen receptor binding sites and regulates transcriptional output, as evidenced by changes in estrogen receptor-associated eRNA expression and stronger estrogen receptor binding at active enhancers after GRHL2 knockdown. Conclusions Our findings provide new insight into the role of GRHL2 in regulating eRNA transcription as part of estrogen receptor signaling. These results demonstrate VULCAN, available from Bioconductor, as a powerful predictive tool. Electronic supplementary material The online version of this article (10.1186/s13059-019-1698-z) contains supplementary material, which is available to authorized users.

The development of commercially available transfection reagents for gene transfer applications has revolutionized the field of molecular biology and scientific research. However, the challenge remains in ensuring that they are efficient, safe, reproducible and cost effective. Bacteriophage (phage)-based viral vectors have the potential to be utilized for general gene transfer applications within research and industry. Yet, they require adaptations in order to enable them to efficiently enter cells and overcome mammalian cellular barriers, as they infect bacteria only; furthermore, limited progress has been made at increasing their efficiency. The production of a novel hybrid nanocomplex system consisting of two different nanomaterial systems, phage vectors and conventional transfection reagents, could overcome these limitations. Here we demonstrate that the combination of cationic lipids, cationic polymers or calcium phosphate with M13 bacteriophage-derived vectors, engineered to carry a mammalian transgene cassette, resulted in increased cellular attachment, entry and improved transgene expression in human cells. Moreover, addition of a targeting ligand into the nanocomplex system, through genetic engineering of the phage capsid further increased gene expression and was effective in a stable cell line generation application. Overall, this new hybrid nanocomplex system (i) provides enhanced phage-mediated gene transfer; (ii) is applicable for laboratory transfection processes and (iii) shows promise within industry for large-scale gene transfer applications.

Background Estrogen Receptor-alpha (ER) is the main driver of~75% of all breast cancers.

In recent years, transcriptional roadblocking has emerged as a crucial regulatory mechanism in gene expression, whereby other DNA-bound obstacles can block the progression of transcribing RNA polymerase (RNAP), leading to RNAP pausing and ultimately dissociation from the DNA template. In this review, we discuss the mechanisms by which transcriptional roadblocks can impede RNAP progression, as well as how RNAP can overcome these obstacles to continue transcription. We examine different DNA-binding proteins involved in transcriptional roadblocking and their biophysical properties that determine their effectiveness in blocking RNAP progression. The catalytically dead CRISPR-Cas (dCas) protein is used as an example of an engineered programmable roadblock, and the current literature in understanding the polarity of dCas roadblocking is also discussed. Finally, we delve into a stochastic model of transcriptional roadblocking and highlight the importance of transcription factor binding kinetics and its resistance to dislodgement by an elongating RNAP in determining the strength of a roadblock.