My research focuses on the effects of iron and heme on the physiology and expression of virulence traits in Pseudomonas aeruginosa. P. aeruginosa is an important human pathogen of great concern for nosocomial exposure, causing serious infections in compromised individuals, especially burn victims and people with Cystic Fibrosis. In order to establish successful infection, P. aeruginosa requires iron and employs several strategies for its acquisition. One of these involves the uptake and degradation of iron-porphyrin, or heme, an abundant source of iron in the human body. P. aeruginosa acquires extracellular heme via the Phu (Pseudomonas heme uptake) and Has (heme assimilation system) proteins. The heme can then be degraded to biliverdin and carbon monoxide by use of a heme oxygenase (HemO), releasing the iron for intracellular functions. Although required for survival, surplus iron or heme can lead to oxidative stress; thus, the ferric uptake regulator (Fur) protein tightly regulates the uptake of these nutrients. In iron-replete environments, Fur blocks expression of genes required for iron and heme uptake, as well as two nearly identical genes encoding the PrrF1 and PrrF2 small regulatory RNAs (Wilderman et al., 2004). The PrrF RNAs negatively affect the expression of a large number of genes whose products function in key metabolic pathways, one of which profoundly affects production of quorum sensing molecules (Oglesby et al., 2008). Consequently, the PrrF RNAs are capable of exerting wide-ranging effects on multiple aspects of P. aeruginosa physiology and virulence.
Heme is an abundant source of iron in the human body, and its acquisition by P. aeruginosa is hypothesized to play a significant role in infection. Because of the potentially toxic effects of heme, however, it is also expected that a heme regulatory system coordinates expression of genes for heme uptake, degradation, and biosynthesis. I and others in Dr. Vasil's lab previously showed that the prrF locus encodes an additional, longer RNA, designated PrrH, which is repressed by heme (Ochsner et al., 2000; Oglesby-Sherrouse and Vasil, 2010) and believed to impart additional regulatory activities to P. aeruginosa. My own studies demonstrate the PrrH RNA is 325 nt in length, and its transcription initiates at the 5’ end of prrF1, proceeds through the prrF1-prrF2 intergenic sequence (95 nt), and terminates at the 3’ end of the prrF2 gene (Oglesby-Sherrouse and Vasil, 2010). Thus, expression of prrH is dependent on read-through transcription at the prrF1 Rho-independent (intrinsic) terminator. My preliminary data suggest heme regulation of prrH is achieved by modulating prrF1 termination efficiency. Although the Rho-independent transcriptional terminators of prrF1 and prrF2 are identical, the nucleotide sequences both upstream and downstream of these structures differ. Therefore, I hypothesize a heme-regulated anti-terminator binds either the nascent PrrF1 RNA at sequence upstream or the DNA sequence downstream of the prrF1 terminator, stabilizing the interaction of RNA polymerase with DNA and allowing for prrH expression. Studies are currently underway to test this hypothesis.
and Vasil ML. (2010). Characterization of a heme-regulated non-coding RNA encoded by the prrF
locus of Pseudomonas aeruginosa
. PLoS One
, Farrow JM, Lee JH, Tomaras AP, Greenberg EP, Pesci EC, and Vasil ML. (2008). The influence of iron on Pseudomonas aeruginosa
physiology: a regulatory link between iron and quorum sensing. J. Biol. Chem.
Cited in Faculty of 1000 Biology: www.f1000biology.com/article/id/1119759/evaluation
, Murphy ER, Iyer VR, Payne SM. (2005). Fur regulates acid resistance in Shigella flexneri
via RyhB and ydeP
. Mol. Microbiol.
Mey AR, Wyckoff EE, Oglesby AG
, Rab E, Taylor RK, Payne SM. (2002). Identification of the Vibrio cholerae
enterobactin receptors VctA and IrgA: IrgA is not required for virulence. Infect. Immun.
Payne SM, Wyckoff EE, Murphy ER, Oglesby AG
, Boulette M, and Davies N. (2006). Iron and pathogenesis of Shigella
: iron acquisition in the intracellular environment. Biometals