Title

Role of IVS in Increased Degradation of 23S rRNA during SCV phase of Coxiella

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Presentation

Abstract

Coxiella burnetii is one of the most infectious pathogens known (ID50=1-10 bacteria). It is able to infect both humans and livestock; in humans it causes Q fever. Due to its low infection dose as well as its incredible resistance to environmental factors, Coxiella is recognized as a potential bio-terrorism agent (class B select agent) and thus is important to study. Coxiella has a biphasic development cycle; it cycles from a stationary phase, small cell variant (SCV), to a developmentally active, log phase, large cell variant (LCV). In this project, I have been investigating Coxiella’s intervening sequence (IVS) within its rRNA gene. IVS’s are selfish genetic elements that disrupt 23S rRNA genes and have to be excised in order to yield a mature, fragmented 23S rRNA. Previous work with Salmonella showed that fragmentation of 23S rRNA by an IVS correlated with enhanced degradation of its 23S rRNA during stationary phase. I hypothesize that IVS-mediated fragmentation of 23S rRNA enhances its degradation during Coxiella’s transition from log-phase (LCV) to stationary-phase growth (SCV). The 23S rRNA of Coxiella was amplified using polymerase chain reaction (PCR). The PCR product was transcribed in vitro and the IVS sequence was excised using RNase III, a cellular rRNA processing enzyme. The fragmented 23S rRNA was used in an in vitro rRNA degradation assay with RNase A. As a control, intact 23S rRNA was also degraded with RNase A. It is hypothesized that because the fragmented 23S rRNA has four termini as a substrate for RNase A, degradation will occur at an increased rate compared to intact 23S rRNA. The results of this experiment will further clarify the purpose of the highly conserved IVS sequence in the Coxiella genome and its role in the development cycle of the bacterium.

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Apr 12th, 4:00 PM Apr 12th, 4:20 PM

Role of IVS in Increased Degradation of 23S rRNA during SCV phase of Coxiella

UC 332

Coxiella burnetii is one of the most infectious pathogens known (ID50=1-10 bacteria). It is able to infect both humans and livestock; in humans it causes Q fever. Due to its low infection dose as well as its incredible resistance to environmental factors, Coxiella is recognized as a potential bio-terrorism agent (class B select agent) and thus is important to study. Coxiella has a biphasic development cycle; it cycles from a stationary phase, small cell variant (SCV), to a developmentally active, log phase, large cell variant (LCV). In this project, I have been investigating Coxiella’s intervening sequence (IVS) within its rRNA gene. IVS’s are selfish genetic elements that disrupt 23S rRNA genes and have to be excised in order to yield a mature, fragmented 23S rRNA. Previous work with Salmonella showed that fragmentation of 23S rRNA by an IVS correlated with enhanced degradation of its 23S rRNA during stationary phase. I hypothesize that IVS-mediated fragmentation of 23S rRNA enhances its degradation during Coxiella’s transition from log-phase (LCV) to stationary-phase growth (SCV). The 23S rRNA of Coxiella was amplified using polymerase chain reaction (PCR). The PCR product was transcribed in vitro and the IVS sequence was excised using RNase III, a cellular rRNA processing enzyme. The fragmented 23S rRNA was used in an in vitro rRNA degradation assay with RNase A. As a control, intact 23S rRNA was also degraded with RNase A. It is hypothesized that because the fragmented 23S rRNA has four termini as a substrate for RNase A, degradation will occur at an increased rate compared to intact 23S rRNA. The results of this experiment will further clarify the purpose of the highly conserved IVS sequence in the Coxiella genome and its role in the development cycle of the bacterium.