Nyla Vasquez

 Introduction. Salmonella typhimurium is a gram negative facultative anaerobe that causes substantial morbidity and mortality worldwide1. Infections with Salmonella when left untreated can progress to typhoid fever and can become fatal. Salmonella infections are prevalent in areas with poor sanitation where food and water are easily contaminated. Individuals who are over 70 years of age and under 20 are at an increased risk because of a compromised immunity and low gastric acidity, respectively1. In the past few decades there has been a rise in the increase in drug resistance against this pathogen sparking the search for new anti-virulent therapeutics1. Research has shown that Salmonella secretes effector proteins that manipulate hosts ubiquitin system to maximize its effector function and obtain nutrients, thereby promoting bacterial proliferation and survival within the host2-3. One effector in particular, SopB, is an example of such manipulation4. Once secreted by Salmonella via a T3SS mechanism, it is quickly ubiquitinated diversifying its function and propagated infection4. A study has shown that the own hosts TRAF6, an E3 ligase, is responsible for the ubiquitination of the effector molecule SopB secreted by Salmonella, allowing it survive within the host cell4. These findings suggest a potential therapeutic against Salmonella infection. Methods. To discover the E3 ligase, a Henle-407 S100 assay was used. In the assay, 3HA-SopB and FLAG-tagged ubiquitin were used as substrates where anti-HA and anti-FLAG immunoblotting of 3HA-SopB led to a new band and the ability to detect the activity and purification of E3 ubiquitin ligase. Potential E3 ubiquitin ligases were purified and subjected to sequential chromatography columns. The fractions from each column were assayed for the capability to ubiquitinate 3-HA-SopB.The largest peak was analyzed by tandem mass spectrometry analysis3. Results. After use of mass speak it was determined that the peak was TRAF6. TRAF6 is the E3 ligase that ubiquitinates SopB after translocation. It was further shown that traf6 mutants with no E3 ligase activity did not ubiquitinate SopB. Conclusion. S.typhimurium has evolved a mechanism of utilizing the host’s E3 ubiquitin ligase to modify and modulate the function of its effector proteins in order to secure pathogen and host cell survival4. Recent finds have shown that TRAF6 is responsible for the ubiquitination of SopB. Inhibitors of TRAF6 can act as potential therapeutics to combat this infection. A recent study has proposed a novel approach to screen for potential inhibitors of TRAF6 using a Time-Resolved Fluorescence Resonance Energy Transfer Assay5.

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2. Ashida H, Kim M, Sasakawa C Exploitation of the host ubiquitin system by human bacterial pathogens Nature Reviews Microbiology 2014; 12: 399–413
3. Narayanan, , Edelmann MJ. Ubiquitination as an Efficient Molecular Strategy Employed in Salmonella Infection Front Immunology 2014; 5(10): 558
4. Ruan HH, Li Y, Zhang XX, Liu Q, Ren H, Zhang KS, Zhao Identification of TRAF6 as a ubiquitin ligase engaged in the ubiquitination of SopB, a virulence effector protein secreted by Salmonella typhimurium. Biochemical and Biophysical Research Communications. 2014; 447(1): 172-177
5. Hong CA, Swearingen E, Mallari R, Gao R, Cao Z, North A, Young SW, Huang ASSAY and Drug Development Technologies. July 2004, 1(supplement 2): 175-180