Poster Presentation BacPath 13: Molecular Analysis of Bacterial Pathogens Conference 2015

The role of the GcvB sRNA in the regulation of protein production in Pasteurella multocida (#200)

Emily Gulliver 1 , Amy Wright 1 , Oded Kleifeld 2 , David Powell 3 , Marina Harper 1 , John Boyce 1
  1. Department of Microbiology, Monash University, Clayton, VIC, Australia
  2. Monash Bioinformatics Platform, Monash University, Clayton, VIC, Australia
  3. Department of Biochemistry and Molecular Biology, Monash University, Clayton, VIC, Australia

P. multocida causes several diseases of economic importance to agricultural industries, including fowl cholera in chickens, haemorrhagic septicaemia in cattle and atrophic rhinitis in pigs. The primary virulence factors of P. multocida, namely capsule, filamentous haemagglutinin and LPS, have been well characterised, but the mechanisms of gene regulation in the organism are poorly understood. Small RNA (sRNA) molecules are critical players in bacterial gene regulation. These non-coding sRNAs bind to mRNA targets, often in association with the RNA chaperone Hfq, to inhibit or activate protein production. Through bioinformatics analysis of the P. multocida transcriptome, we identified an sRNA that is a putative homolog of the E. coli and Salmonella GcvB. In E. coli, GcvB has been shown to regulate production of amino acid transport and biosynthesis proteins. To determine if P. multocida GcvB acts similarly, a gcvB mutant strain of P. multocida was constructed. High throughput quantitative liquid proteomics was used to compare the proteomes of the gcvB mutant and wild-type. This analysis identified 25 proteins as produced at increased levels in the gcvB mutant; 18 of which are predicted to be involved in amino acid biosynthesis and 6 in amino acid transport. The gene sequences of each putative GcvB target were then compared using Clustal Omega and Meme motif finder and a consensus sequence of 5’-ACACAACA-3’ was identified. This sequence closely matches the GcvB seed region identified in E. coli and Salmonella GcvB targets. Therefore, P. multocida GcvB acts to decrease the production of amino acid biosynthesis and transport proteins with a binding mechanism that likely includes complementary base pairing to a highly conserved 8 nucleotide seed region on target mRNAs.