Functional Analysis of the Polymorphic Membrane Protein Family of Chlamydia Item Type dissertation Authors Grinblat-Huse, Valerie Publication Date 2013 Abstract Chlamydia trachomatis is the most common sexually transmitted bacterial pathogen and the leading cause of infectious blindness. It is an obligate intracellular pathogen that has a biphasic lifecycle encompassing infectious elementary bodies (EBs) and... Keywords adhesion; polymorphic; Chlamydia trachomatis; Membranes; Proteins Download date 03/10/2021 18:41:02 Link to Item http://hdl.handle.net/10713/3641 CURRICULUM VITAE NAME: Valerie Ann Grinblat-Huse EDUCATION: University of Maryland at Baltimore, Baltimore, Maryland June 2008 – October 2013 Ph.D. in Molecular Medicine Passed qualifying examination, July 2010 Johns Hopkins University, Baltimore, Maryland December 2003 – September 2007 M.S. in Biotechnology James Madison University, Harrisonburg, Virginia August 1999 – May 2003 B.S. in Computer Science, Minor in Spanish ACADEMIC AWARDS and COMMUNITY ACTIVITIES: 2012 – 2013 Graduate Student Association Representative 2012 – 2013 Graduate Assistant Advisory Council (to UMB Executive Board) 2012 – 2013 The Graduate Journal Editor/Contributor 2011 – 2012 Dental School Training Grant 2009 Travel Grant, Annual Conference on Microbial Genomics 2008 – 2009 Meyerhoff Scholars Fellowship 2008 – 2013 Meyerhoff Scholar 2006 – 2013 Homeward Trails Animal Rescue, Adoption Coordinator RESEARCH EXPERIENCE: Doctoral Research: Department of Microbial Pathogenesis, University of Maryland at Baltimore, Oct. 2010 – present . Research focuses on virulence factors and methods of pathogen infectivity . Interpretation of technical data to a non-scientific audience . Experience with writing and editing technical documents including grant writing . Development of research plan and use of innovative approaches and techniques . Data extraction, analysis, and translation of results . Implementation and instruction of techniques new to laboratory members and collaborators . Training of collaborators, research associates, and other graduate students . Hands-on experience with cloning and genetic manipulation, along with RNA, protein, and cell culture techniques . Gaining experience with fluorescent, confocal, and electron microscopy techniques . Maintaining knowledge of new technologies through research and attendance at national and local professional meetings Doctoral Research: Department of Microbiology and Immunology, University of Maryland at Baltimore, Jan. 2009 – Oct. 2010 . Studied biofilm formation in Halorubrum lacusprofundi . Microarray development and analysis . Bioinformatics analysis and script writing Research Associate III-MB: Research and Development Team in the Pathogen Functional Genomics Resource Center, The J. Craig Venter Institute, July 2005 – June 2008 . Provided analysis and assessment of research and development projects to the management team . Wrote and established Standard Operating Procedures (SOPs) to include an all- encompassing SOP designed to automate a tracking system and improve efficiency . Oversaw high-throughput pipeline operations from start to finish . Research and Development projects included isothermal amplification of rare transcripts and testing multiplex PCR for use in a high-throughput cloning process . Evaluated new technologies and optimized protocols . Maintained and programmed Biomek Beckman-Coulter liquid-handling robots for entire department . Trained new hires and research associates . Supplied data for grant application COMPUTER EXPERIENCE and SKILLS: . Familiar with several languages including: PERL, JAVA, C, C++, Visual Basic, Visual C++, HTML, and SQL . Experienced in Microsoft and Apple applications . Knowledge of UNIX and LINUX Computer Specialist: System Source, Hunt Valley, Maryland, Oct. 2003 – Jan. 2004 . Installed computer hardware; imaged computers and installed commercial software, set up labs for BlueCross/Blue Shield; Updated software and removed viruses to assure security was not compromised Billing Consultant/Technical Support: Nexpoint Technologies Inc., Harrisonburg, Virginia, 2002 – 2003 . Provided technical support to clients in regards to code problems and server issues . Reconsolidated billing issues . Designed and implemented billing manual to facilitate the training of new hires Technical Assistant: ECI Telecom (formerly Wavepacer division of Hubbell Pulsecom), Herndon, Virginia, Summer 2000, Winter 1999, Summer 1999 . Member of transition team; set up database of schematics to be transferred overseas . Tested Network Management Systems and ADSL software systems; Set up computers: installed Windows 95, 98, and NT PUBLICATIONS and PRESENTATIONS: Mlacha SZK, Peret TCT, Kumar N, Romero-Steiner S, Hotopp JCD, Ishmael N, Grinblat-Huse V, Riley DR, Erdman DD, Carlone GM, Sampson J, Scott JAG, Tettelin H. (2013). Transcriptional adaptation of pneumococci and human pharyngeal cells in the presence of a virus infection. BMC Genomics, 14:378. Huse V, Drabek EF, Creasy HH, Daugherty S, Jones KM, Santana-Cruz I, Tallon LJ, Read TD, Hatch TP, Bavoil P, Meyers GSA*. (2011). Genome sequences of the zoonotic pathogens Chlamydia psittaci 6BC and Cal10. Journal of Bacteriology, Aug;193(15):4039-40. Capes MD, Coker JA, Gessler R, Grinblat-Huse V, DasSarma SL, Jacob CG, Kim JM, DasSarma P, DasSarma S. (2011). The information transfer system of halophilic archaea, Plasmid, v. 65:2, p. 77-101. Grinblat-Huse V*, Marques P, Shou H, Bavoil P. (2011). Polymorphic Membrane Protein-Mediated Adherence of Chlamydia trachomatis to Epithelial Cells. Poster session presented at: The Annual Meeting of the NIAID STI Cooperative Research Centers; Nov 1-2; Chapel Hill, NC. Carrasco-Lopez JA*, Tan C, Kennard A, Huse V, Shou H, Rank R, Bavoil PM. (2011). Expression of the polymorphic membrane protein gene family of Chlamydia caviae under conditions of penicillin-induced persistence. Poster session presented at: 5th Biennial Meeting of the Chlamydia Basic Research Society; Mar 18 – 21; Redondo Beach, CA. DasSarma PD, Coker JA, Huse V, DasSarma S. (2010). Halophiles, Industrial Applications, in Flickinger MC, ed., Encyclopedia of Industrial Biotechnology, Bioprocess, Bioseparation, and Cell Technology, Wiley. Huse V, DasSarma S. (2009). Evaluation, Faculty 1000, http://f1000.com/1163626#evaluations DasSarma S, Coker JA*, Huse V*, Gessler R, Capes M, Kim JM, Han J, DasSarma P, DasSarma S, Tatusov R, Anderson IJ, and Kyrpides NC. (2009). Comparative Genomics of Halophilic Archaea: The Conserved Predicted Proteins. Poster session presented at: 17th Annual Microbial Genomics Conference; Oct 11-15; Rocky Gap State Park, MD. ABSTRACT Title of Dissertation: Functional Analysis of the Polymorphic Membrane Protein Family of Chlamydia Valerie Grinblat-Huse, Doctor of Philosophy, 2013 Dissertation Directed by: Dr. Patrik Bavoil, Professor and Chair, Department of Microbial Pathogenesis Chlamydia trachomatis is the most common sexually transmitted bacterial pathogen and the leading cause of infectious blindness. It is an obligate intracellular pathogen that has a biphasic lifecycle encompassing infectious elementary bodies (EBs) and metabolically active reticulate bodies (RBs). C. trachomatis encodes a polymorphic membrane protein (Pmp) gene family whose nine members are differentially expressed under in vitro growth conditions. The observed “phase-variation-like” phenotype of the autotransported Pmps is consistent with observed variable Pmp subtype-specific antibody profiles in patients infected with C. trachomatis. The autotransported Pmps display repeated, often alternating motifs GGA(I,L,V) and FXXN within the predicted “passenger” domain that have been proposed to mediate adhesion of Chlamydia pneumoniae to susceptible cells. The identification of a “magic bullet” adhesin responsible for attachment and subsequent internalization by epithelial cells has eluded researchers for several decades, owing to the inherent difficulties of the Chlamydia system, in particular the genetic intractability of these organisms. Others have established that Pmps of C. pneumoniae and PmpD of C. trachomatis function as adhesins and that PmpD-specific antibodies block attachment. I hypothesize that antibodies against each of the six Pmp subtypes (A, B/C, D, E/F, G/I & H) can neutralize a C. trachomatis infection, and that the application of multiple Pmp subtype-specific antibodies will have a cumulative negative impact on infection. I additionally hypothesize that Pmps of C. trachomatis facilitate adhesion to epithelial cells. For Aim 1, I will investigate the neutralizing effects of single Pmp subtype-specific antibodies by infecting epithelial cells with C. trachomatis serovar E EBs that have been preincubated with antibodies against each of the Pmp subtypes. The inclusions will be visualized using immunofluorescence. For Aim 2, I will investigate Pmp-mediated adherence to epithelial cells using a Pmp-coated latex bead-binding assay. The neutralization assay results cast doubt on the ability of Pmp-specific antibodies to neutralize a chlamydial infection on their own. It is possible that the Pmp-specific antibodies are necessary for complete neutralization, but are not sufficient. Additionally, the adhesion assay results suggest that other functions of Pmps should still be considered, however, their role in adhesion should not be ruled out. Functional Analysis of the Polymorphic Membrane Protein Family of Chlamydia by Valerie Grinblat-Huse Dissertation submitted to the Faculty