Thesis Title
Using Fluorescence Nanoscopy to Study RNA Localization in Borrelia burgdorferi, the Spirochete that Causes Lyme Disease
Department or Program
Biological Chemistry
Abstract
The spatial arrangement of RNA localization plays a role in the expression and turnover of transcripts in B. burgdorferi, including those that are involved in its virulence and pathogenicity. RNA localization was experimented with by using techniques such as fluorescence in situ hybridization with RNA polymerase subunits and regulatory Factors, using RNA aptamers through induction, and imaging using stimulated emission depletion nanoscopy. Throughout experimentation, it will be determined whether RNA transcripts distribute themselves along the full length of B. burgdorferi cells or localize at the poles, septa of dividing cells, or are concentrated at other foci within the cells. In both fluorescence in situ hybridization and RNA aptamers a fluorescence signal was observed.
Level of Access
Restricted: Archival Copy [No Access]
First Advisor
Schlax, Paula
Date of Graduation
5-2021
Degree Name
Bachelor of Science
Recommended Citation
1. Irastortza-Olaziregi, M. and O. Amster-Choder, RNA localization in prokaryotes: Where, when, how, and why. WIREs RNA, 2021. 12(2): p. e1615. 2. Motaleb, M.A., J. Liu, and R.M. Wooten, Spirochetal motility and chemotaxis in the natural enzootic cycle and development of Lyme disease. Curr Opin Microbiol, 2015. 28: p. 106-13. 3. Malge, A., et al., mRNA transcript distribution bias between Borrelia burgdorferi bacteria and their outer membrane vesicles. FEMS Microbiology Letters, 2018. 365(13). 4. Burgdorfer, W., et al., Lyme disease-a tick-borne spirochetosis? Science, 1982. 216(4552): p. 1317-9. 5. Sapi, E., et al., Improved culture conditions for the growth and detection of Borrelia from human serum. Int J Med Sci, 2013. 10(4): p. 362-76. 6. Shapiro, E.D., Borrelia burgdorferi (Lyme disease). Pediatrics in review, 2014. 35(12): p. 500-509. 7. Tilly, K., P.A. Rosa, and P.E. Stewart, Biology of infection with Borrelia burgdorferi. Infectious disease clinics of North America, 2008. 22(2): p. 217-v. 8. Fleshman, A.C., et al., Reported County-Level Distribution of Lyme Disease Spirochetes, Borrelia burgdorferi sensu stricto and Borrelia mayonii (Spirochaetales: Spirochaetaceae), in Host-Seeking Ixodes scapularis and Ixodes pacificus Ticks (Acari: Ixodidae) in the Contiguous United States. Journal of Medical Entomology, 2021. 58(3): p. 1219-1233. 9. Dumes, A.A., Lyme Disease Outside In, in Divided Bodies: Lyme Disease, Contested Illness, and Evidence-Based Medicine. 2020, Duke University Press. p. 0. 10. Lacout, A., et al., The Persistent Lyme Disease: "True Chronic Lyme Disease" rather than "Post-treatment Lyme Disease Syndrome". J Glob Infect Dis, 2018. 10(3): p. 170- 171. 11. Nelson, C.A., et al., Incidence of Clinician-Diagnosed Lyme Disease, United States, 2005-2010. Emerg Infect Dis, 2015. 21(9): p. 1625-31. 12. Samuels, D.S., Gene regulation in Borrelia burgdorferi. Annu Rev Microbiol, 2011. 65: p. 479-99. 13. Snow, S., et al., Transcript decay mediated by RNase III in Borrelia burgdorferi. Biochem Biophys Res Commun, 2020. 529(2): p. 386-391. 14. Jutras, B.L., et al., Posttranscriptional self-regulation by the Lyme disease bacterium's BpuR DNA/RNA-binding protein. J Bacteriol, 2013. 195(21): p. 4915-23. 15. Jutras, B.L., et al., The Lyme disease spirochete's BpuR DNA/RNA-binding protein is differentially expressed during the mammal-tick infectious cycle, which affects translation of the SodA superoxide dismutase. Mol Microbiol, 2019. 112(3): p. 973-991. 16. Dong, T., R. Yu, and H. Schellhorn, Antagonistic regulation of motility and transcriptome expression by RpoN and RpoS in Escherichia coli. Molecular Microbiology, 2011. 79(2): p. 375-386. 17. Buck, M. and W. Cannon, Specific binding of the transcription factor sigma-54 to promoter DNA. Nature, 1992. 358(6385): p. 422-422. 18. Ouyang, Z., J.S. Blevins, and M.V. Norgard, Transcriptional interplay among the regulators Rrp2, RpoN and RpoS in Borrelia burgdorferi. Microbiology (Reading), 2008. 154(Pt 9): p. 2641-2658. 34 19. Ouyang, Z., et al., Activation of the RpoN-RpoS regulatory pathway during the enzootic life cycle of Borrelia burgdorferi. BMC Microbiol, 2012. 12: p. 44. 20. Fisher, M.A., et al., Borrelia burgdorferi σ54 is required for mammalian infection and vector transmission but not for tick colonization. Proceedings of the National Academy of Sciences of the United States of America, 2005. 102(14): p. 5162-5167. 21. Li, Y., K. Ke, and R.C. Spitale, Biochemical Methods To Image and Analyze RNA Localization: From One to Many. Biochemistry, 2019. 58(5): p. 379-386. 22. Levsky, J.M. and R.H. Singer, Fluorescence in situ hybridization: past, present and future. Journal of Cell Science, 2003. 116(14): p. 2833-2838. 23. Cawte, A.D., P.J. Unrau, and D.S. Rueda, Live cell imaging of single RNA molecules with fluorogenic Mango II arrays. Nature Communications, 2020. 11(1): p. 1283. 24. Swetha, P., et al., Genetically encoded light-up RNA aptamers and their applications for imaging and biosensing. J Mater Chem B, 2020. 8(16): p. 3382-3392. 25. Hein, B., K.I. Willig, and S.W. Hell, Stimulated emission depletion (STED) nanoscopy of a fluorescent protein-labeled organelle inside a living cell. Proceedings of the National Academy of Sciences, 2008. 105(38): p. 14271. 26. Persson, F., et al., Fluorescence Nanoscopy of Single DNA Molecules by Using Stimulated Emission Depletion (STED). Angewandte Chemie International Edition, 2011. 50(24): p. 5581-5583.
Number of Pages
35