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Summer 2007

	Aarti Chawla Home school: Beloit College  Loyola mentor:  Dr Katherine Knight This summer I worked with Kari Severson, a graduate student in the Knight lab. One of the aims of the Knight lab is to understand the mechanism by which intestinal bacteria promote gut-associated lymphoid tissue (GALT) development. There are a number of processes by which the intestinal microflora stimulate GALT development, but this summer I focused on one of these potential mechanisms: M cell transport. M cells are specialized epithelial cells that transcytose bacteria from the lumen into their pocket where they can facilitate interaction with the immune cells present there. It has earlier been shown by Rhee et al. that certain bacterial species induce rabbit GALT development better than others, in particular a combination of B. subtilis and B. fragilis induce B cell proliferation and follicle formation much better than either bacterium does individually. I hypothesized that there is a correlation between bacterial uptake by the M cells and GALT development. To test this hypothesis, the goal of my project was to generate M cells in vitro using a previously described system, and upon generating M cells, test for bacterial uptake using B subtilis and/or B. fragilis. My data showed that neither B. subtilis nor B. fragilis alone were sampled by M cells, but B. fragilis was taken up by M cells in the presence of B. subtilis. Although preliminary, these data suggest that B. subtilis may facilitate the uptake of B. fragilis through M cells, and offer an interesting possibility as to how commensal bacteria may work together to promote rabbit GALT development.
	Kate Eaton Home school: Beloit College  Loyola mentor:  Dr Nancy Zeleznik-Le I worked in Dr Nancy Zeleznik-Le laboratory this summer. My summer research project was involved with beginning to explore a novel mechanistic pathway that might be relevant to MLL leukemia. Dr Zeleznik-Le had recently determined that expression of a particular microRNA (small non-coding RNA), mir196b, was regulated by MLL. This same microRNA was also over-expressed in MLL fusion leukemias. I cloned several different sizes of precursors for this microRNA into an expression vector. We first used PCR amplification to obtain the fragments to clone, then ligated them into a retroviral vector, and screened for plasmids containing the insert. I was successful in obtaining the correct clones and prepared the DNA for subsequent functional analyses.
	Niyati Parikh Home school: Loyola University Chicago Loyola mentor:  Dr Alan Wolfe I had the wonderful opportunity to work in Dr. Alan Wolfe's lab this summer. My project focused on investigating the relationship between the pta-ack pathway and the cpxAR pathway in E.coli bacteria. The cpxAR pathway enables bacteria to respond to various stress signals from the envelope surrounding the bacteria. The pta-ack pathway produces acetyl phosphate and acetate. During the course of the ten weeks, I identified a linkage between central metabolism and the cpxAR signaling pathway. By performing epitasis experiments (making combinations of double mutants in two different pathways and determining what is required for activation of the cpxAR pathway), I discovered that pta-ack may be another input into cpx signaling. Contributing to the discovery of this alternate mechanism by which cpx signaling may be activated was a unique opportunity and I enjoyed learning skills and techniques involved in research.
	Caitlin Paul Home school: Kalamazoo College Loyola mentor:  Dr Katherine Knight This summer I worked with Venkata Yeramilli, a graduate student in the Knight lab. One of the focuses of the Knight lab is to understand the mechanism by which B cells are stimulated in the Gut Associated Lymphoid Tissues (GALT). My project for the summer was to test if a cytokine known as B cell activating factor (BAFF) can stimulate B cells isolated from rabbit GALT. To address this question, I cloned, expressed and purified rabbit BAFF from E.coli. I then set up in vitro cultures with splenocytes and B cells isolated from GALT to test if BAFF can induce these cells to undergo proliferation. From my preliminary experiments, it appeared that BAFF only induced proliferation in splenocytes, but not in B cells isolated from GALT. Perhaps BAFF induces a survival signal for GALT B cells. I really enjoyed my time in the lab this summer and I learned so much during this 10 week program.
	Enya Qing Home school: Beloit College  Loyola mentor: Dr. Karen Visick and Dr. Alan Wolfe This summer I had an amazing time working on a combined project between the labs of Dr. Karen Visick and Dr. Alan Wolfe. The goal of my project was to address the role of cyclic-di-GMP, a novel second messenger, in the regulation of flagella assembly by Vibrio fischeri. Previous studies by the two labs had shown that the bacteria’s flagellation was dependent on the presence of magnesium. They also found out that loss of the MifA protein, which synthesizes cyclic-di-GMP, restores some motility to cells grown in the absence of magnesium. These data suggested that c-di-GMP produced by MifA inhibits flagellation; however, it is unknown at what stage in flagellar biogenesis this inhibition occurs. In my project, I searched for and isolated mutants that could overcome MifA-mediated inhibition. Some of these mutations mapped to mifA itself, while others mapped elsewhere on the V. fischeri chromosome. Although I was not able to identify the chromosomal mutations I obtained, my work revealed that such an approach is feasible.
	Jenny Wu Home school: University of IL - Urbana-Champaign Loyola mentor: Dr. Karen Visick   I spent this summer working at Dr. Karen Visick's lab, which studies the symbiosis between Euprymna scolopes (a Hawaiian squid) and Vibrio fischeri (a bioluminescent bacterium). Previous experiments from the Visick lab supported the model that an interaction between a sensor kinase and a response regulator promotes transcription of the symbiosis polysaccharide (syp) cluster, a group of genes necessary for the colonization of E. scolopes by V. fischeri. The goal of my project was to determine which response regulators were required (or partially required) for interacting with a sensor kinase, called RscS, for the induction of this syp cluster and for the regulation of downstream phenotypes (including attachment to glass biofilms, pellicles, and wrinkled colonies). By the end of the summer, I had tested 36 out of the 40 putative response regulators and determined that SypG was the only response regulator involved in syp transcription, while a number of other response regulators, including SypG, were involved in downstream phenotype regulation. The future direction of my project is to investigate whether RscS indeed donates its phosphate to SypG, as expected from my results..

Summer 2006

	Katherine Houser Home school: Illinois Wesleyan College Loyola mentor:  Dr. Susan Baker This summer I worked in Dr. Susan Baker’s lab.  The research I worked on involved testing antiviral drugs against the mouse coronavirus MHV.  These drugs targeted the active site of one of the two proteases of the virus, 3CLPro, halting the enzymatic activity. The drugs had already been proven effective in purified protease samples, but had yet to be tested in living cells.  I tested 35 drugs, and while 22 displayed no viral inhibition and 5 exhibited cell toxicity, 8 drugs did exhibit viral inhibition.  One drug, GRL-0346S, was shown to inhibit viral replication at a concentration of 7mM.  The drugs that were found to be effective at inhibiting the viral replication in the cell cultures will continue on to be tested against SARS-HCoV in the Biosafety Level 3 lab at UIC.  I also used the effective drugs in order to create drug resistant variants of the virus.  These mutants can then be sequenced so that the drugs can be improved and the resistance avoided.
	Michael Misale Home school: Carthage College Loyola mentor:  Dr. Karen Visick This summer I worked with Dr. Kati Geszvain in the lab of Dr. Karen Visick.  Dr. Visick’s lab studies the symbiosis between the squid Euprymna scolopes and the bioluminescent bacteria Vibrio fischeri.  The primary focus of our experiments was to answer the question: which region of Rscs, a sensor kinase in Vibrio fischeri, is responsible for regulating a group of genes known as the symbiosis polysaccharide cluster (syp cluster).  The syp cluster is a group of eighteen genes that is believed to produce a polysaccharide to help induce the aggregation of bacteria on the squid’s light organ.  Past experiments have shown that rscS is also an important factor in promoting symbiosis; bacteria which lack rscS fail to colonize the squid.  Our approach to determine which region of the protein RscS is responsible for its function was to mutagenize the gene and screen Vibrio fischeri colonies for those with increased transcription of the syp cluster. We then determined if these putative rscS mutations had any effect on other bacterial phenotypes, such as biofilm and pellicle formation.  We found that those mutations that increased syp transcription also increased the bacteria’s capability to synthesize biofilms and pellicles.  The next step is to determine where the mutations are located.
	Deborah Muganda Home school: North Central College Loyola mentor:  Dr. Karen Visick This summer I worked with Beth Hussa, a graduate student in the Visick lab. The Visick lab studies the symbiotic relationship between the Hawaiian squid Euprymna scolopes and the bacterium Vibrio fisheri.  The lab specifically looks at the bacterial factors that allow colonization of the squid host. They have discovered a gene cluster known as the symbiosis polysaccharide (syp) cluster which is required for colonization. The syp cluster is believed to alter the surface of the cells in a way that allows them to enter the squid.  My project this summer studied the regulation of the syp cluster. The syp cluster is regulated by 2-component signal transduction. In this type of signaling system, the sensor kinase transfers a phosphate to a response regulator in response to environmental signal. My goal was to study the effects of  several 2-component regulators on syp expression and syp-related phenotypes, such as biofilm formation. Based on the work that I did, we conclude that regulation of the syp cluster is controlled by more than one response regulator.
	Courtney St. Clair Home school: The College of William and Mary Loyola mentor: Dr. Chris Wiethoff This summer I worked in the laboratory of Dr. Chris Wiethoff.   His lab studies the cell entry of Adenovirus type 5, a nonenveloped virus infecting the upper respiratory tract. My project this summer was to develop an assay that allows one to view Ad5 cell entry using fluorescent microscopy. I fluorescently labeled both viral particles and cellular membranes. This labeling allowed us to visualize the location of viral particles with respect to endosomes at specific times after a synchronous infection. Hopefully, my assay will eventually be used to identify both viral and cellular proteins that affect Ad5 cell entry.
	Kathleen Starr Home school: St. Peter’s College Loyola mentor: Dr. Dennis Lanning This summer I had the opportunity to work with Dr. Dennis Lanning in the Knight lab. The lab studies B-cell development and antibody repertoire diversification in rabbits. My project for the summer focused on how B-cells and T-cells develop in the appendix of rabbits and what might induce proliferation of B-cells and diversification of the antibody repertoire. Two color in situ hybridization allowed us to examine the expression of the two major chemokines that establish B-cell and T-cell areas in the developing appendix of neonatal rabbits. Our results suggested that interaction between B-cell and T-cell areas might be required to induce B-cell proliferation and antibody repertoire diversification.

Summer 2005
 

	Erin Boileve Home school: Augustana College Loyola mentor:  Dr. Dennis Lanning This summer I have had the unique opportunity to work with Dr. Dennis Lanning in the Knight lab. This lab studies rabbit B cell development and has discovered that two specific bacteria, B. subtilis and B. fragilis, together, can induce gut-associated lymphoid tissue (GALT) formation. My project involved Toll-like receptors (TLRs), which recognize molecular patterns associated with microorganisms. My goal this summer was to study TLR expression in rabbit GALT through the method of in situ hybridization in order to investigate the possible involvement of TLRs in bacteria-induced GALT development.
	Jason Cascio Home school: Knox College Loyola mentor:  Dr. Katherine Knight I am working in the lab of Dr. Katherine Knight which studies the role of commensal bacteria in the development of the gut-associated lymphoid tissue (GALT) of rabbits. My project involves the creation of a soluble single chain Fv/immunologlobulin G fusion protein and using it to detect interactions that may stimulate B cell development in GALT. We know that the commensal bacteria are required for GALT development, but the use of this fusion protein can help elucidate exactly how the bacteria are having an effect.
	Agnese Collino Home school: U. Perugia-Italy Loyola mentor:  Dr. Adam Driks This summer I had the opportunity of joining Dr. Adam Driks’ lab, where they study bacterial spore coat assembly. The coat is essential for many spore resistance properties and its assembly depends on a subset of proteins called morphogenetic coat proteins, one of which is CotH. A newly identified coat protein candidate, YisJ, is a homolog of CotH and may also have a morphogenetic role. The goal of my summer research is to analyze the spore coat of a yisJ strain of Bacillus subtilis: this required me to delete yisJ and examine the resulting spores for the loss of coat proteins, spore resistance properties, and for changes in coat ultrastructure. I will then compare the yisJ and cotH phenotypes to determine if the proteins play similar roles in spore coat assembly.
	Alex Hagmeyer Home school: Miami U. of Ohio Loyola mentor: Dr. Alan Wolfe This summer I worked in the laboratory of Dr. Alan Wolfe, who studies a small phosphorylated molecule called acetyl phosphate. Recently, he found this molecule to influence two-component signaling transducton pathways involved in the ability of E. coli to form biofilms, communities of bacteria that impact many aspects of our daily lives. Since acetyl phosphate influences the formation of biofilms in E. coli, it is thought that there might be other important cell functions that this small molecule influences as well. The focus of my project was identifying these other functions. To do this, I looked for signaling pathways that are required for cells that make too much or too little acetyl phosphate.  Using P1 mediated transduction, I generated strains of E. coli that lacked genes responsible for specific two-component signaling pathways as well as genes that regulate levels of acetyl phosphate. By characterizing the growth or death of these double mutants compared to a wild type control, I determined signaling pathways required for the presence or absence of acetyl phosphate. Furthermore, I optimized the protocol for P1-mediated transduction for future transductions in the laboratory.
	Jamie Hofmeister Home school: U. Illinois Urbana-Champaign Loyola mentor: Dr. Alan Wolfe This summer, I have had the unique opportunity to work in two microbiology labs.  In Dr. Alan Wolfe’s lab, the research focuses around small molecules and signal transduction pathways.  Most of the lab studies the small molecule, acetyl-phosphate.  However, I am involved in preliminary research on another small molecule, cyclic-di-GMP, and its effect on motility in E. coli.  I worked with an E. coli strain transformed with a plasmid that overexpresses a diguanylate cyclase (DGC), an enzyme that converts two GTP molecules into the second messenger cyclic-di-GMP. Thus, this transformant produces excess cyclic-di-GMP; it also produces very few flagella.  In an attempt to understand the relationship between high levels of cyclic-di-GMP and the lack of flagella, I sought mutants that regained the ability to assemble flagella.  I am now attempting to locate and identify these mutations.  During the second half of the 10 weeks, I also have been working in Dr. Karen Visick’s lab, which studies the symbiosis of V. fischeri, a bacterium that colonizes in Euprymna scolopes, a Hawaiian squid.  V. fischeri contains two putative DGCs (MifA and MifB) that appear to be involved in inhibiting motility of V. fischeri in the absence of Mg2+.  I am investigating the consequences of overexpression of one of these DGCs, MifA, on motility of V. fischeri.
	Samantha Michalski Home school: Milwaukee School of Engineering Loyola mentor: Dr. Alan Wolfe Dr. Wolfe's lab is interested in the role small molecules play in signaling pathways. My project focuses on one small molecule (acetyl phosphate) and one two-component signaling pathway (the EnvZ-OmpR pathway in E. coli). I have been using strains of E. coli that have fluorescent protein transcription fusions at the loci of two genes whose expression can be affected by this pathway. By monitoring the levels of fluorescence of cultures grown in media with varying osmolytes and carbon sources, I am indirectly studying acetyl phosphate levels under these conditions.
	Brittany Mortensen Home school: Augustana College Loyola mentor: Dr. David Keating I worked in Dr. Keating's lab, whose research focuses on the symbiotic relationship between a legume, alfalfa, and a bacterium, Sinorhizobium meliloti. My project focused on understanding how a gene, syrA, could upregulate the transcription of the lpsS gene and exo genes. These genes are important in the symbiosis. SyrA does not look like any other known transcriptional regulator, but rather like a membrane-associated protein. To then understand how it may work as a transcriptional regulator, we isolated mutations that affect SyrA function. We were able to select for mutated plasmids and then analyzed them using agarose and SDS gels, assays, phenotype analysis and gene sequencing. I have truly enjoyed the experience and the concepts and techniques I have learned are invaluable.

 
 

Summer 2004
 
 

	Ali Chinwalla Home school: U. Illinois Urbana Champaign Loyola mentor:  Dr. Tom Gallagher This summer, I have had the opportunity to work in Dr. Tom Gallagher's lab, where the focus lies primarily in the study of coronaviruses and their entry into cells.  With the recent emergence of the potentially fatal SARS Coronavirus, the need exists for the study of various mechanisms and requirments of SARS viral entry into human cells.  In order to acheive this, I aimed to make pseudotyped versions of the SARS virus which I could use to infect 293T human kidney cells.  After producing SARS pseudotypes, I attempted to transduce 293T cells that had been transfected to express the SARS-specific ACE2 receptor and 293T cells that had not been transfected.  Transduction efficiency was checked by measuring luciferase activity and associated production of light.  Via the methods of transfection, transduction, and luciferase assays, I successfully produced pseudotyped SARS viruses, an accomplishment that could have clinical applications in the near future.
	Elaine Cruz Home school: Loyola University Chicago Loyola mentor:  Dr. Dennis Lanning I worked with Dr. Dennis Lanning in the Knight lab, which studies rabbit B-cell development. As a summer student, my objective was to study the positive feedback loop that occurs between B-cells and stromal cells in the rabbit gut-associated lymphoid tissue (GALT). To do this, I cloned and looked at the expression of TNF, LT  and BLC by performing radioactive in situ hybridization with appendix tissues exposed to follicle and non-follicle inducing bacteria. By looking at this interaction, we can learn how the positive feedback loop contributes to the development of rabbit GALT.
	Paul Duffin Home school: Grinnell College Loyola mentor:  Dr. Adam Driks My summer research was conducted in the lab of Dr. Adam Driks, whose primary research goal is elucidating mechanisms involved in spore coat assembly in Bacillus species. My project involved identifying spore surface proteins in the novel spore-forming bacterium Bacillus odysseyi, which was isolated from the Mars Odyssey spacecraft prior to launch. B. odysseyi spores can survive NASA's rigorous decontamination protocols and are highly adhesive to spacecraft surfaces. We believe the tight adhesion is due to novel spore proteins found in the exosporium, the outermost surface. These proteins share homology to outer membrane proteins. Using degenerate primer PCR based on peptide sequences, I have amplified and cloned these putative genes for sequencing. Additionally, using Western Blotting, I have found a putative CotE protein B. odysseyi. CotE is a major morphogenic protein required for the proper spore coat assembly. Furthermore, using degenerated PCR based on other Bacillus species I have attempted to amplify cotE in B. odysseyi.
	Erin Marciniec Home school: Augustana College Loyola mentor: Dr. Katherine Knight My goal this summer was to examine Th1 and Th2 cytokine expression during bacterially induced follicle development in rabbit gut associated lymphoid tissue (GALT). We used in situ hybridization of radiolabeled RNA probes to determine if Interleukin-4 (a signature Th2 cytokine) would be more highly expressed in germ-free appendix exposed to follicle-inducing bacteria than in germ-free appendix tissue exposed to non-follicle-inducing bacteria. I worked in the lab of Dr. Katherine Knight with an incredibly fun and diverse group of hard-working scientists. Despite the unchecked hilarity of our daily chitchat, the members of the Knight lab achieve excellent experimental results and always understand the concepts and "big picture" behind every procedure.
	Emily Richter Home school: Illinois Wesleyan University Loyola mentor: Dr. Susan Baker I worked in Dr. Baker's lab, where we research coronaviruses, such as SARS coronavirus, and their replication. One of our focuses is on the papain-like proteases. Plpro, found in SARS coronavirus, is involved in cleaving several sites in the viral replicase polyprotein. My project centered on the nsp1/nsp2 SARS replicase cleavage site. If Plpro is unable to cleave this site, viral replication ceases. Through site-directed mutagenesis I altered the amino acids around this cleavage site so we can better understand how Plpro works, forming a foundation on which therapeutic drugs can be developed.
	Theresa Wenzel Home school: Augustana Loyola mentor: Dr. Adam Driks This summer I have had the opportunity of working in the lab of Dr. Adam Driks.  This lab studies the proteins that make up the spore of several Bacillus species.  Some Bacillus spores, including Bacillus anthracis spores, possess an exosporium, which is an outer layer surrounding the spore coat.  My project focused on determining if CotE, CotB and ExsF are exosporial-specific proteins in Bacillus anthracis.  Either French Press or acid wash treatments were used to strip exosporial proteins from the spore.  Then I used Western blotting to detect the presence of these proteins in different spore fractions.  Knowledge of the localization of these proteins could lead to methods of detection and treatment of anthrax.

 

Summer 2003
 
 

	Michael Durkin Home school: Illinois Wesleyan Loyola mentor:  Dr. Katherine Knight The Knight lab recently discovered that B cell development arrests in adult rabbits. This summer, I studied with Susan Kalis in the Knight lab in an attempt to determine what causes the arrest of B cell development. To do this, I cloned and examined the expression levels of various genes required for B cell lymphopoeisis using Northern blot analysis. These genes include: Stem Cell Factor, Flt3 ligand (which are both environmental factors) and E2a (which is an intrinsic factor). If we find that one of these factors is expressed in the bone marrow of young rabbits, but not adult rabbits, then it might help us explain what causes this arrest of B cell development.
	Christine Zacharia Home school: Illinois Wesleyan Loyola mentor:  Dr. Katherine Knight I am working in Dr. Katherine Knight'slab this summer, studying B-cell development in the rabbit. My objective as a summer student is to clone rabbit interleukin-5, a cytokine secreted by Th2 cells that aids in B-cell proliferation.  Additionally, I will be assisting Dr. Dennis Lanning with Real Time PCR to detect the relative expression of different cytokine genes in the appendix of the rabbit.  By looking at their expression levels, we will be able to identify whether the Th1 or Th2 response is occurring in the rabbit appendix in the presence of follicle or non-follicle forming bacteria.  Thus, the IL-5 gene I clone this summer, along with other cytokines Dennis has already cloned in the lab over the past year will serve to assist us in addressing this concern effectively.
	Catherine Smith Home school: St. Xavier University Loyola mentor:  Dr. Susan Baker  I am working in Dr. Susan Baker's lab, which specializes in the replication of Coronaviruses.  The lab is currently working on SARS, which is a newly discovered coronavirus.  My goal for the summer is to generate a specific antibody against the PLP2 region of the SARS replicase polyprotein.  The antibody is generated by first cloning the PLP2 region of SARS, translating the cloned product into protein, and injecting the protein into rabbits.  The rabbits will make the antibody.  This antibody will aid in the characterization of the SARS PLP2 domain.
	Jonathan Moreira Home school: U. Illinois Urbana Champaign Loyola mentor: Dr. Susan Baker I am working in Dr. Susan Baker's molecular virology lab.  Dr. Baker is researching the replication of coronaviruses, and with the recent discovery of a coronavirus as the microbial agent responsible for SARS, her work and research is vital to understanding how to treat the disease.  My specific project this summer involves mutating the cysteine residues of the zinc-binding domain of PLP2, an important protease in murine hepatitis virus (MHV).  Research has shown that by mutating the cysteine residues surrounding the zinc-binding domain, the enzyme undergoes a conformational change that directly affects its ability to bind to its substrate and cleave protein.  Thus, Dr. Baker postulates that if the same is done to SARS PLP2-cen, it will no longer be able to cleave protein.
	Alex Slack Home school: Harvard University Loyola mentor: Dr. Adam Driks Adam Driks’ lab studies the formation and structure of bacterial spore coats in different bacteria, including Bacillus subtilis and Bacillus anthracis. In the Driks lab, I worked to make GFP-fusions to four novel proteins in B. anthracis. Using fluroescence microscopy on these GFP-fusion strains of B. anthracis would allow us to pin down where each protein localizes in the spore. Any coat proteins identified through these means could then be targeted with antibodies or chemicals to aid in decontamination and detection of anthrax.
	Byron Burnette Home school: University of Wisconsin-Whitewater Loyola mentor: Dr. Adam Driks My summer research project was carried out in the laboratory of Dr. Adam Driks  whose primary research objective is elucidating the mechanism of assembly of proteins in the spore coat of spore-forming bacteria, as well as identifying those components which are as yet unknown. I was fortunate enough to be assigned to a very appealing project involving a novel spore-forming Bacillus species just discovered within the last year following its isolation from the outer surface of the Mars Odyssey spacecraft pre-launch. My project was to characterize the outermost structure of this novel Bacillus species known as the “exosporium” with the intention of developing decontamination protocols to avoid the translocation of the bacterium into our extraterrestrial environment.
	Erin McGonigle Home school:  Augustana Loyola mentor: Dr. Adam Driks This summer I have had the privilege of working in the lab of Dr. Adam Driks. The Driks lab is primarily interested in studying the proteins associated with the spore coat. My project has entailed working with the genes that encode for two of the proteins present in the Bacillus subtilis spore coat: CotQ and YgaK. I have overproduced the 5' portion of the CotQ protein in E. coli, and have also created a ygaK disruption strain in E. coli, which I am in the process of transforming into B. subtilis. The ultimate goal of the research I have begun is to provide evidence for the hypothesis that CotQ and YgaK are enzymes. This would support the current theory that the coat plays an active role in the maintenance of the spore.
	Alexandra Asrow Home school: Brown University Loyola mentor:  Dr. Karen Visick I am working in the lab of Dr. Karen Visick. This lab studies the symbiotic relationship between the marine bacterium Vibrio fischeri and the Hawaiian squid Euprymna scolopes.  V. fischeri colonizes the symbiotic light organ of the squid after it hatches and provides bioluminescence. My particular project involves constructing and characterizing mutants defective for a subset of the 33 candidate response regulator proteins encoded by the V. fischeri genome. Response regulators comprise one-half of a two-component signal transduction system (the other half being sensor kinases),  regulatory systems in bacteria responsible for recognizing and responding to environmental signals. We anticipate that this approach will help us identify the putative response regulator, RscR, that is proposed to function with RscS, a sensor kinase important for colonization.  Identifying and characterizing rscR will shed new light on the process of symbiotic colonization.

 

Summer 2002
 
 

	Patrick Kennedy Home school: Amherst College  Loyola mentor:  Dennis Lanning  I worked with Dr. Dennis Lanning in the laboratory of  Dr. Katherine Knight.  This summer Dennis and I worked on B-cell development in rabbits with both normal and ligated appendixes.  More specifically, the interaction that takes place between intestinal bacteria, B-cells and follicular- dendritic cells (FDCs) and how this drives gut associated lymphoid tissue (GALT) development and diversification of the primary repertoire in the appendix.  We hypothesized that B-cell development is a direct result of stimulating effects caused by the FDCs interacting with specific intestinal bacteria.  Once they are stimulated they will in turn release specific molecules (LTƒa and TNF) which will allow the FDCs to develop as well.
	Heather Mears Home school:  Loyola University Chicago  Loyola mentor:  Dr. David Keating  The Keating lab studies the symbiotic relationship between the bacteria Sinorhizobium meliloti and the legume Medicago satina.  My research involves the identification of genes required for symbiosis.  In particular, my project involves the use of a promoter trap plasmid for the identification of genes that are transcriptionally active on or around the plant, but transcriptionally inactive on laboratory media.  I will then use arbitrary PCR to identify the regions of DNA responsible for the transcription activity.
	Kristin Tracy Home school:  Illinois Wesleyan  Loyola mentor:  Dr. Susan Baker  This summer I am working in Dr. Susan Baker's lab, which investigates the mechanism by which Mouse Hepatitis Virus (MHV), a representative coronavirus, replicates itself.  MHV is a positive strand RNA virus that uses an RNA-dependent RNA polymerase to replicate its genetic material.  The protein encoded by the MHV gene region PLP2-MP1 has been shown to form double-membrane vesicles in infected cells, and my project involves infecting cells that express a tagged PLP2-MP1 protein with wild-type MHV to determine whether the wild-type replicase will associate with the double-membrane vesicles induced by the recombinant PLP2-MP1.  In order to deliver the recombinant gene to the cells, I am optimizing a Sindbis viruse expression system, which packages recombinant RNA into pseudovirions that can infect cells and cause them to express recombinant protein.  I am also cloning the PLP2-MP1 and MP1 regions for use with the Sindbis system.
	Patricia Valassis Home school:  University of Illinois  Loyola mentor: Dr. Karen Visick  I worked with Kristine Barney in Dr. Karen Visick's lab.  The Visick lab studies the symbiotic relationship between the Hawaiian squid Euprymna scolopes and the luminescent bacteria Vibrio fischeri.  The lab discovered a bacterial gene, rscS, which is essential for colonization by V. fischeri.  My project was to determine how this gene is controlled at the level of transcription. By fusing the rscS gene with the lacZ gene, we were able to monitor the amount of transcription of the gene through b-galactosidase assays.  We discovered that transcription of rscS is dependent on the cAMP-receptor protein, Crp.

Summer 2001
 
 

	Joy Campbell Home school:  North Park University  Loyola mentor:  Karen Visick  Research in the laboratory of Dr. Karen Visick focuses on the symbiotic relationship of the luminescent marine bacterium Vibrio fischeri and its host, the Hawaiian squid Euprymna scolopes.  Recently, the  Vibrio fischeri gene rscS has been identified as a component necessary  for colonization.  My project  has focused on investigating the regulation of transcription of this gene.  To do this, I am working on  constructing a fusion of rscS to the reporter gene lacZ.  I will determine the level of transcription of the  rscS gene under various laboratory conditions by assaying b--galactosidase activity resulting from the rscS-lacZ fusion.
	Nana Sarkoah Fenny Home school:  Beloit  Loyola mentor:  Dr. Dennis Lanning  I am working with Dr. Dennis Lanning in the laboratory of  Dr. Katherine Knight.  The primary objective of the lab is to determine the mechanism of VDJ gene diversification in rabbits.  My research project is to verify that the number of a- genes being found in the tissues of a  ligated appendix of a rabbit after introduction of different bacteria is representative of the actual numbers present as opposed to a skewing of results due to  PCR amplification techniques.  I have conducted experiments in which DNA was PCR-amplified instead of mRNA.  Other experiments I am conducting are PCR amplifications of different ratios of known a- and a+ genes to verify if the same ratios are obtained after amplification.
	Sarah Knopf Home school:  Benedictine University  Loyola mentor:  Kathy Cho   I am working with Kathy Cho, a graduate student in the laboratory of  Dr. Katherine Knight.  Our overall project is to determine the Vgamma  repertoire in the immune system of the rabbit.  Specifically, I am working on mapping the V gamma locus in the rabbit.  By doing this, I will be able to determine the relative location and the number of V gamma genes.
	Rishad Motlani Home school:  Elmhurst College  Loyola mentor: Dr. Gayatri Vedantam  In the Hecht lab, our studies focus on understanding how antibiotic resistance occurs in the bacterium Bacteroides fragilis.  It has been seen that as a result of bacterial conjugation, antibiotic resistance genes can be transferred.  This transfer is greatly enhanced when the bacteria are exposed to low doses of tetracycline (induction). Furthermore, the Hecht lab has captured all the genes of B. fragilis on a conjugation plasmid.  My project is centered on trying to understand what proteins are expressed or encoded by these captured genes during conjugation.
	Kari Roettger Home school:  Beloit College  Loyola mentor: Dr. Susan Baker   This summer I am working in Dr. Susan Baker’s lab, which studies the pathogenesis of coronaviruses.  The lab has successfully generated 10 different antisera to distinct regions of the coronavirus replicase polyprotein.  I have been given the task of generating polyclonal antibodies specific for PLP2, the papain-like cysteine proteinase 2 region of the coronavirus replicase polyprotein.  The antiserum will then be used for further characterization of the coronavirus PLP2 domain.
	Taylor Tooley Home school:  U. Illinois Urbana Champaign  Loyola mentor:  Dr. Kim Foreman  I am spending the summer in Dr. Kim Foreman's lab, which focuses on the pathogenesis of Karposi's Sarcoma (KS), the most common skin cancer in HIV-1 infected patients.  Several factors have been linked to the development of KS, such as infection by human herpesvirus 8 (HHV8).  This virus has been detected in virtually all KS tumors and encodes many proteins that are cellular homologues of regulatory cell cycle proteins.  My project has been to characterize the expression of the HHV8 protein LANA (latency associated nuclear antigen) in endothelial cells.  LANA has been shown to inhibit p53 activation of cell death. In addition, we can also determine LANA's effect on other proteins upregulated in KS that may be involved with HHV8 infection.
	Aaron Ufferman Home school:  Valparaiso University  Loyola mentor:  Dr. Veronica Volgina   This summer I am working in the laboratory of Dr. Katherine Knight.  Research conducted by the Knight lab is focused on factors influencing the expression and diversification of the B-cell antibody repertoire.  Under the direction of Dr. Veronica Volgina, I am studying regulatory elements associated with the rabbit immunoglobulin locus, specifically, the enhancers located 3' of cd genes.  Hopefully, by the end of the summer, I will have combined two distinct enhancers (hs3 and hs1,2) into a single construct, alowing me to test whether or not these two elements have a synergistic relationship.

Summer 2000

	Cynthia Bugajsky Home school: Elmhurst college Loyola mentor: Pam Witte  This summer I am working in Dr. Pam Witte’s lab, where the overall goal is to study the regulation of B-cell lymphopoiesis. During development in the bone marrow, B cell precursors are in close contact with a supporting population of cells termed stromal cells. Limited information is available on factors that regulate the interaction between B cell precursors and stromal cells. My project is focused on characterizing the protein CD28, which is found on the surface of stromal cells and may regulate stromal-dependent B lymphopoiesis. Specifically, I am studying the pattern of CD28 expression on stromal cells both in vitro and in vivo using immunofluorescence, flow cytometry, and confocal microscopy.
	Kimberly Dittmar Home school: Bendictine University Loyola mentor: Phong Le  This summer, I am working in the lab of Dr. Phong Le who studies the thymus and aging.  One main focus of the lab is the mechanism of thymic involution, a physiological condition associated with aging.  In humans, it has been shown that this involution is associated with decreased thymic output.  During T cell receptor rearrangement, an important step in T cell development, circular excision products (TRECs) are formed.  I have been measuring the TREC levels in young vs. old mice by PCR.  The data confirms that there are more TRECs in the young mice than in old mice, indicating that the old mice have decreased thymic function (i.e. old mice are producing fewer T cells).
	Stephanie Miller Home school: Illinois Wesleyan University Loyola mentor: Karen Visick  The Visick lab is studying genetic requirements for the bacterium, Vibrio fischeri, to colonize its symbiotic host, the Hawaiian squid Euprymna scolopes.  One gene, rscS, has been identified as being necessary for V. fischeri to successfully colonize its host.  The rscS gene encodes a protein similar to the sensor component of hybrid two-component regulatory systems.  It is hypothesized that the role of RscS is to regulate the transcription of genes important in symbiotic colonization.  By making a rscS deletion mutant, randomly inserting the lacZ reporter gene, and using complementation analysis with the rscS gene, a blue/white screen will allow me to identify possible genes under the control of RscS.
	Rishad Motlani Home school: Elmhurst College Loyola mentor: Alan Wolfe  Dr. Alan Wolfe's lab studies the machinery that regulates the transcription of acs, the gene that encodes acetyl-CoA synthetase (Acs). Acs, an enzyme that scavenges for small amounts of environmental acetate, helps cells survive starvation. This summer, I have been examining the interaction between the cyclic AMP receptor protein (CRP) and the a-CTD (the C-terminal domain of the RNA polymerase subunit a). We hypothesize that this protein-protein interaction can either activate or repress acs transcription, depending upon signals that emanate from the environment. To test this hypothesis, I am scoring the effect exerted upon promoter activity by a library of mutations in the a-CTD. If we are correct, then a-CTD mutations that alter amino acids involved in the interaction with CRP should equally affect activation and repression.
	Ana Mrejeru Home school: Loyola University Chicago Loyola mentor: Karen Visick  In Dr. Karen Visick's lab, I have been exploring the role of chemotaxis in the symbiosis between Vibrio fischeri bacterium and its Hawaiian squid host.  We hypothesize that chemotaxis, or the ability of bacteria to recognize and respond to certain chemical attractants and repellents present in the environment, may be vital for successful colonization of the squid. I am taking two approaches to investigate chemotaxis of V. fischeri. In collaboration with Dr. Alan Wolfe's lab, I have been characterizing the environmental conditions necessary for a bacterial cell to produce and utilize flagella, its mode of transportation from the sea water into the squid's symbiotic organ. Further, I am working on mutating the cheR gene responsible for chemotaxis, which will allow us to determine what role this gene plays in colonization. Together, these approaches will allow us to get a better understanding of how V. fischeri cells chemotax in nature
	Laurie L. Smith Home school: North Park University Loyola mentor: Jody Brewer  I worked in Dr. Jody Brewer's lab, where the research objective is to better understand the mechanisms that regulate the homeostasis of the endoplasmic reticulum (ER).  Proteins destined for secretion or insertion into the cell membrane are properly folded and assembled in the ER.  However, conditions such as the depletion of intracellular calcium can cause ER “stress”—a buildup of unfolded proteins—and activate the unfolded protein response (UPR).  The UPR induces an overall decrease in cellular protein synthesis while leading to increased expression of molecular chaperones, such as BiP, that facilitate normal protein folding in the ER.  One of my projects was to determine if the UPR is activated in response to clotrimazole, an anti-tumor and anti-fungal drug known to deplete calcium from the ER. Our data indicates that expression of BiP and CHOP, another gene activated by the UPR, are induced by clotrimazole. A second project was to use reverse transcriptase and PCR to generate a cDNA encoding the cytoplasmic sections of Ire1a, an ER transmembrane protein considered to be a critical component of the UPR signaling pathway.

The 2000 summer student T-shirt.

Alumni notes