Characterization of Type IV Pilus System Genes and Their Regulation in Clostridium perfringens

Clostridium perfringens is a Gram-positive (Gr+) anaerobic pathogen that was found to contain Type IV pilus (T4P) system genes within the genomes of all its sequenced strains. T4P are widely used in Gram-negative organisms for aggregation, biofilm formation, adherence, and DNA uptake. Because few examples of T4P-utilizing Gram-positive bacteria are studied to date, we wanted to characterize the T4P system in this Gr+ bacterium.

To understand the regulation of T4P genes and therefore better understand their expression, we employed the highly powerful next-generation sequencing tool RNA-seq in a variety of conditions. RNA-seq uncovered previously unknown regulatory mechanisms surrounding T4P genes as well as provided transcriptional information for most of the genes in the C. perfringens strain 13 genome. We also utilized reporter gene assays to look at post-transcriptional regulation of T4P promoters.

The wealth of RNA-seq data acted as a jumping-off point for many smaller projects involving transcriptional regulators that may influence T4P expression. We investigated a novel small RNA in close proximity to the major T4P operon, as well as two little-characterized transcriptional regulators that function in the same conditions as T4P genes. RNA-seq also provided data to develop a method for protein purification from C. perfringens without induction.