Mr. Alon Agua
Mentor: Dr. Ohyun Kwon
Title: Systematic Structural Variation of a
Novel Class of Chiral Phosphines
(Christopher Henry, Alon Agua, Ohyun Kwon)
Alon Agua is a 1 st year MARC Trainee and a 4 th year undergraduate student majoring in Chemistry. He joined Dr. Ohyun Kwon’s lab group in Spring quarter of his 2 nd year at UCLA and is currently under the mentorship of graduate student Christopher Henry.
Recent efforts in organophosphine catalysis have resulted in the discovery of numerous useful heterocycle forming reactions. Formation of asymmetric variants of the reactions remains a challenge; therefore, synthesis of new chiral phosphine ligands is important. Stereoselective reactions are also useful in drug development because the resultant compounds need to be enantiomerically pure compounds. This project hopes to study the effects of systematic structural modifications on the ability of a chiral phosphine to catalyze annulations with high yield and stereoselectivity. In particular, Alon is trying to functionalize the amino position of a rigid azaphospha [2,2,1] bicyclic chiral phosphine catalyst. He has carried out the necessary synthetic route in the synthesis of the phosphine ligands. From trans-4-hydroxy proline, the route begins with N-sulfonylation or N-acylation of the amine followed by reduction of the acid. Di-O-sulfonylation prepares the compound for a ring closure with dilithium phenyl phosphine to synthesize the desired bicyclic chiral phosphine ligands. Systematic variation of the N-substituent will allow the synthesis of a collection of structurally related chiral phosphines. These phosphines will be evaluated in various annulations to determine the effects of catalyst structure on asymmetric induction.
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Ms. Cecil Benitez
Mentor: Dr. Ellen Carpenter and Dr. Patricia Phelps
Title: Migration Defects in Identified Cell Populations
in reeler Mutant Spinal Cord
(Cecil Benitez, Ellen Carpenter)
Cecil Benitez is a fourth year student majoring in Physiological Science and a second year MARC U*Star participant. She is currently researching the role of Reelin in the migration of spinal cord neurons. This is her second year in this lab.
Reelin is a large extracellular glycoprotein that is involved in neuronal migration and lamination of the cerebral and cerebellar cortexes. However, little is known about the role of reelin in the spinal cord. Reeler mice do not produce Reelin and as a result have ataxia, tremors, and show disrupted cerebral and cerebellar lamination. To elucidate the positioning effects of Reelin on various spinal cell populations, Cecil compared wildtype and reeler mutant transverse spinal cord sections using immunohistochemistry. She found that in heterozygous mice at embryonic age 12.5, evenly dispersed HB9-expressing motor neurons progressively split to form medial and lateral motor clusters whereas in reeler mutants these motor neurons failed to segregate and tightly aggregated medially. Anti-neurofilaments immunohistochemisty also revealed disorganization of axonal trajectories in reeler mice. In wildtype mice, axons labeled with anti-neurofilament antibodies are tightly bundled as they travel from the dorsolateral to the ventromedial regions of the spinal cord. Axons are much less tightly bundled in reeler mutant mice and also made aberrant medial projections. These positioning defects support Reelin’s role in neuronal migration in the spinal cord.
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Ms. Thelma Escobar
Alexander van der Bliek
Title: Determining FIS-1 and FIS-2 Function in Mitochondrial Dynamics
(Alexander van der Bliek, Brian Head, Thelma Escobar )
Thelma Escobar is a fourth year student majoring in Molecular Cellular Developmental Biology. She is a second year MARC student and is currently working in a Biological Chemistry laboratory where Mitochondrial Morphology and Dynamics research is being conducted. She entered this lab in the fall quarter of her sophomore year and is under the supervision of Alexander van der Bliek and her lab mentor Brian Head. Thelma will be graduating this June and she will be conducting research for a year before she begins her graduate studies.
Responsible for the cells' metabolic processes, the mitochondria are essential organelles whose division and fusion events are important for cell growth and survival. With Caenorhabditis elegans as a model organism, Alexander van der Bliek’s laboratory is investigating different proteins that are involved in mitochondrial division and fusion. They are interested in the factors that influence mitochondrial division and its determination of mitochondrial shape. Thelma has been analyzing the function and localization of two proteins (Fis-1 and Fis-2) in the mitochondria of C. elegans muscle. Previous research on Fis-1 has elucidated the function of Fis-1 in both S. cerevisiae and mammalian cells. Fis-1 function in S. cerevisiae and mammalian cells is to recruit Drp-1, a dynamin related protein, to the outer mitochondrial membrane where mitochondrial division will occur. Loss of function in Fis-1, results in a connected net like mitochondrial morphology due to the absence of an essential mitochondrial division protein. Thelma has acquired data that suggests that both Fis-1 and Fis-2 are outer mitochondrial membrane proteins and they both have an affect in mitochondrial morphology. However, her data revealed that there is no division defect in C. elegans muscle cells when Fis-1 and Fis-2 are both knocked-out. With these results, she is currently conducting experiments to find the outer membrane protein that recruits Drp-1 to a division complex on the outer mitochondrial membrane.
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Ms. Lizette Flores
Dr. Stephanie White
Title: Understanding the Role of FoxP2 in Learned Vocalization
Lizette Flores is a third year student majoring in Psychology-Biology, with a minor in Spanish. As a first year MARC student she is currently working in a Physiological Science laboratory where they are interested in how social interactions influence neural plasticity. Lizette joined this lab summer of 2006.
FOXP2 protein is a transcriptional regulator that is known to be the only single molecule directed linked to human speech and language. FoxP2 therefore provides an entry point to underlying neural mechanisms in humans. Though language is a unique human characteristic, there are some species who share characteristics required for language , such as vocal learning. Vocal learning requires the ability to modify innate vocalization, and Zebra Finches (a songbird species) share this trait with humans. The Zebra finch song circuitry has been well characterized, which makes it a model organism for study. Zebra finches learn their songs analogous to how humans learn speech, through sensory acquisition (listening to others) and sensory motor (practicing) stages. Lizette’s project involves analyzing the regulation of FoxP2 protein in Zebra finches during different social contexts, either undirected singing (practicing) or directed singing (singing to a female).
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Ms. Denise F. Gonzalez
Mentor: Dr. David A. Campbell and Dr. Nancy R. Sturm
Title: Characterization of Two Distinct Tandem
Arrays of Spliced Leader RNA Genes in Leishmania tarentolae
(David Campbell, Denise Gonzalez, Nancy Sturm, Robert Hitchcock )
Denise Gonzalez is a fourth year student majoring in Biology. She is a second year MARC student and is currently working in a microbiology laboratory specializing in parasitology. She began working in the Campbell lab fall quarter of her second year and is under the supervision of Dr. David Campbell and Dr. Nancy Sturm, in addition to her graduate student mentor Robert Hitchcock.
Trypanosomes are parasitic protests that infect millions of humans and animals worldwide, usually in developing countries. The Spliced Leader (SL) RNA is a small nuclear RNA that plays a critical role in the expression of nuclear protein-coding genes in trypanosomes. The 96-nucleotide SL RNA of Leishmania tarentolae donates its 5’ 39-nucleotide exon to all protein-coding polycistronic mRNAs through trans-splicing to complete the maturation of the mRNA molecules.
Previous studies have shown that there are two types of SL RNA gene arrays, MINA and MINB in L. tarentolae. These two different tandem arrays differ in the length, unit copy numbers, and the sequence of their intergenic regions. MINA and MINB co-localize on a 320-kb band on Southern blots of chromosomal DNA resolved by Pulsed-Field Gel Electrophoresis, however, the organization of these two gene arrays relative to the chromosome(s) is not yet known. The goal of Denise’s project is to locate the region(s) on the 320-kb chromosome band where the two arrays are found. There are two scenarios that can explain the current observations: the genes are at different positions on the same chromosome; or, the genes are on different chromosomes with similar electrophoretic mobilities. Denise has separated the chromosomes of L. tarentolae using Pulsed-Field Gel Electrophoresis. She is currently using polymerase chain reaction to construct probes for the two gene arrays. By using the technique of fiber- FISH, these probes will allow the visualization of the arrangement of MINA and MINB arrays relative to strands of DNA that comprise the chromosome bands. Learning more about SL RNA will put us one step closer to understanding trypanosomes’ cellular processes that aid in their infections.
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Mr. Rodrigo Gonzalez
Mentor: Dr. Giovanni Zocchi
Title:Stability of Polynucleotide Hairpin
Rodrigo Gonzalez is a fourth year student majoring in Biophysics and is in his first year of the MARC program. He is participating in a lab run by Giovanni Zocchi in the physics department. The lab is focused on the study of conformational changes of biological macromolecule. Rodrigo began in the lab fall quarter of his junior year.
Rodrigo’s project consists of looking at the conformational changes of polynucleotide chains. The self-complementary ssDNA and ssRNA sequences can fold into either a hairpin or random coil conformations. The hairpin conformation is composed of two separate parts, the stem and loop. The loop exerts stress on the stem which is proportional to the length. The loop is composed of paired nucleotides, the longer the stem the more stable the molecule. When the hairpin becomes unstable it changes into the random coil conformation. The change of the conformation is temperature sensitive, where the hairpin conformation is stable at lower temperatures. The temperature at which the nucleotide sequence acquires the hairpin conformation and the other half the hairpin conformation is known as the melting temperature (Tm). Both DNA and RNA have a maximum absorbance of approximately 260nm wavelengths. The paired polynucleotide has a lower absorption than unpaired polynucleotide. A polynucleotide in the hairpin conformation will therefore have a lower absorbance than the random coil. A use of UV Spectrophotometer with temperature control allows nucleotide absorbance measurements at different temperatures. Stress induced on the stem by the loop will decrease hairpin stability, decreasing its melting temperature. The goal of these experiments is to quantify this mechanical stress, in order to probe the mechanical response of DNA and RNA to sharp bends. Studies are now underway to examine the effect of varying stem lengths on the stability of DNA and RNA hairpins.
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Mr. Kristopher Kennedy
Mentor: Nicholas C. Brecha, Ph.D. and
Salvatore L. Stella, Ph.D.
Title: Development of thy-1.2 YFP positive mouse cone bipolar neurons in vitro
(Salvatore Stella, Kristopher Kennedy, Nicholas Brecha)
Kristopher is a junior majoring in Neuroscience. He recently transferred from Los Angeles Pierce College to UCLA during the Fall/2006 quarter. Kris received an excellent introduction to UCLA’s research through CARE’s Bridge Summer Research Program for community college students. While in Bridge he chose to study the vertebrate retina in Dr. Nicholas Brecha’s neurobiology lab, under the guidance of Dr. Salvatore Stella. Shortly after completing the Bridge Program, Kris was accepted in the MARC Fellowship Program. As a first year MARC trainee, he is now continuing his summer project in Dr. Brecha’s lab on cone bipolar cell development in vitro.
The Brecha lab is interested in characterizing the functional anatomy of the vertebrate retina, the neurosensory tissue of the eye involved with vision. Kris’s project is concerned with how a subset of mouse retinal bipolar cells develops in culture. The development of retinal neurons in culture provides a critical step in understanding their differentiation in culture, but is poorly understood. Insights into retinal cell development in culture may provide a way to isolate and harvest cultured retinal progenitor cells. The goal of Kris’s project is to characterize the morphological development of yellow fluorescent protein (YFP) expressive mouse cone bipolar cells using a novel culture technique. These transgenic mice express YFP in a subset of neurons in the retina, which includes bipolar cells. This provides a clear fluorescent reporter for easy cell identification in culture. Kris is involved in each step of this study. Once he finishes collecting and analyzing data, he hopes to embark on the rigorous challenge of preparing a publishable manuscript under the guidance of his lab mentors.
As a MARC trainee, Kristopher encourages his peers, especially transfer students, to engage in CARE’s research programs at UCLA. He deeply appreciates the support he has received from everybody involved with CARE and MARC including faculty, staff and fellow students. Kris’s academic goal is to take advantage of all the opportunities here at UCLA like the MARC Program to develop a skill set that will make him an outstanding applicant to Ph.D. programs. From there, he hopes to fulfill his career goal of uncovering mysteries of the human brain and becoming an influential and contributing member of the biomedical research community.
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Mr. Daniel Krauth
Mentor:Dr. Marie-Francoise Chesselet
Title:The Effects of Small Peptides Derived from Amyloids on Induced Cell Dysfunction
(Dr. Marie-Francoise Chesselet and Daniel Krauth)
Daniel Krauth is a 4th year student majoring in Neuroscience. As a first year MARC student in the lab of Dr. Marie-Francoise Chesselet, in the Department of Neurology, Daniel is examining the effects of various small peptides derived from amyloids, specifically, alpha-synuclein and amylin, along with their effect on induced cytotoxicity.
Prior investigations have described the presence of proteinaceous aggregates in several diseases such as Huntington’s, Alzheimer’s, Parkinson’s and other prion-related diseases. The role of these aggregates in cell dysfunction is the subject to much debate especially in diseases such as Huntington’s disease. These aggregates are amyloid in structure and recently the laboratory of Dr. Eisenberg (UCLA) has described the smallest sequences of these disease related proteins that are capable of forming amyloid fibers. In collaboration with the Eisenberg Lab, Daniel looks to examine the cytoxicity or cell dysfunction induced by these disease related peptides. To do this he will continue to examine LDH release, MTT formazan production, and Alamar Blue reduction following administration of these peptides in differentiated PC12 cell lines. Through these experiments, Daniel's work will shed light on how these disease-related proteins which form amyloid can cause cell dysfunction. His experiments will also help his lab further understand whether the cell dysfunction induced by these disparate peptides are related.
Daniel would like to thank Dr. Elma Gonzalez, Dr. Miriam Hickey, and Dr. Marie-Francoise Chesselet in their encouragement towards his ideas and research.
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Mr. Edwin A. Paz
Mentor: Dr. John Colicelli
Title: Creating Signal Transduction Probes to Profile Activated Receptor Tyrosine Kinases in Epithelial Tumor Cells
(Edwin Paz and John Colicelli)
Edwin Paz is a fourth year Microbiology, Immunology & Molecular Genetics student at UCLA. His undergraduate research career began under the direction of Dr. Benjamin Bonavida where he assisted in studying the effects of the antibody Rituximab, and its sensitizing effects on Non-Hodgkin’s Lymphoma Cells. He later joined Dr. Christina Jamieson’s lab where he conducted studies to characterize and detect the levels of the DNA damage sensor, Ataxia Telegactasia Mutated (ATM), to quantify the threshold level at which T cells abandon DNA repair and undergo apoptosis in Dexamethasone treated cells. He is currently working under the guidance of Dr. John Colicelli helping to delineate the specific receptor tyrosine kinases responsible for cancer development in epithelial tumor cells. The objective of this study is to create and purify SH2 domain constructs as probes for evaluating change in RTK activity in tumors and to elucidate the specific receptor tyrosine kinases responsible for cancer development in epithelial tumor cells. By using nanotechnology techniques including Shape Encoded Particles (SEPs) together with different SH2 domain proteins labeled with fluorescent Quantum Dots (Q-dots) it should provide a new level of tumor characterization and may help elucidate specific RTKs responsible for tumorigenesis. Edwin aspires to earn a Ph.D. and to conduct research in the biomedical field while providing future students from inner-city schools with the tools required to excel in the sciences.
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Ms. Lena Pernas
Mentor: Dr. Kent L. Hill
Title: Characterizing the subcellular localization of TbCMF 63 and TbCMF 70 in Trypanosoma brucei
(Kent Hill and Lena Pernas)
A first year MARC student, Lena Pernas is a third year Microbiology, Immunology, and Molecular Genetics major. She began her undergraduate research career working in Dr. Hill’s lab in Fall 2006 under the mentorship of 5th year Katharine S. Ralston, and has since decided to pursue a PhD in a field that fascinates her: parasitology.
Her current research is on flagellar motility in the protozoan parasite Trypansoma brucei, the causative agent of the African Sleeping Sickness. Little is known about the T brucei flagellum on a molecular level, but motility is hypothesized to play a major role in disease pathogenesis by allowing the parasite to spread into vital host tissues. In order to further characterize the role of the flagellum, the lab has used a comparative genomics approach to identify a core group of 50 genes that are conserved in motile flagella, referred to as the T. brucei components of motile flagella (TbCMF). To validate this approach, 41 of these genes have been targeted for RNAi knockdown in T. brucei, and the majority of these knockdown mutants had defective motility. The lab will now further characterize several of these genes by raising antibodies to allow the examination of protein localization and to identify interacting proteins, and by performing site-directed mutagenesis to identify key residues. To begin this process, Lena has selected two putative flagellar genes, TbCMF 63 and 70. TbCMF 63 is of particular interest because it contains a calcium-binding EF hand, and calcium concentration has been shown to affect motility in several organisms. To raise antibodies to these proteins, Lena has cloned these genes into a plasmid with a 6x His marker for protein purification by nickel affinity chromatography. These proteins were expressed in Escherichia coli, purified, and used to immunize mice. Subsequent steps will involve immunofluorescence to determine the subcellular localization of the recombinant protein, and co-immunoprecipitation to identify interacting proteins. To better characterize the role of the calcium binding EF-hand in TbCMF 63, Lena will be probing the function of this putative domain. This will be done by mutagenizing key amino acids and examining mutant phenotypes in collaboration wtih Dr. Springer's lab at Mount Holyoke College.
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Ms. Angelica Riestra
Mentor: Dr. Sherie Morrison
Title: Construction of Mouse-Human Chimeric Antibodies against Cryptococcus neoformans
(Angelica Riestra and Sherie Morrison)
Angelica Riestra is a fourth year student majoring in Microbiology, Immunology, and Molecular Genetics (MIMG). She is a second year MARC student and is currently conducting research in the laboratory of Dr. Sherie Morrison under the mentorship of Kileen Mershon and Ryan Trinh. Angelica started working in the Morrison lab in the summer following her sophomore year. One of the lab’s main areas of research is to develop a better understanding of antibody structure and function, and the application of this knowledge to construct recombinant mouse–human chimeric antibodies with possible therapeutic use.
Cryptococcus neoformans is an opportunistic fungus that poses a life-threatening problem to immunocompromised persons especially those with HIV infection. Although treatment is available, this anti-fungal therapy is not completely effective since 10-20% of treated individuals die from the disease. Furthermore, this treatment does not eradicate the infection in AIDS patients. An alternate treatment that is currently being researched is passive antibody therapy, which has shown promise in murine models. These studies have found that the murine immunoglobulin IgG1 extends the life-span of mice infected with Cryptococcus neoformans. However, preliminary studies in the Morrison laboratory have shown that mouse-human chimeric IgG3 also extended the survival of BALB/C infected mice when using repeated lower antibody doses rather than the usual single dose administered in other studies. The Morrison lab is, therefore, interested in studying the properties of this antibody that may help for the treatment of cryptococcosis. In order to do this, Angelica’s project involves the production, expression, and purification of two mouse-human IgG3 chimeric antibodies against the glucuronoxylomannan polysaccharide component of Cryptococcus neoforman’s capsule. She has already constructed a mouse –human recombinant antibody that contains an N297Q mutation in the CH2 region. This mutation should prevent glycosylation of the antibody. She transfected NSO/1 cells with the construct and has obtained successful expression of the protein. Enzyme-Linked Immunosorbent Assays and incorporation of radioactively-labeled methionine are being used to compare antibody production in order to identify the best protein producers. She is also working on making a second construct in which the IgG3 hinge region will be swapped for an IgG1 hinge. IgG3 is distinct from the other IgG subclasses due to its extended hinge region, which is composed of 62 amino acids and is about four times as long as the IgG1 hinge. The hinge helps determine the flexibility of the molecule which may play a role in triggering certain effector functions like complement activation and Fc gamma receptor binding. Protein A affinity chromatography will be used to purify the proteins, and after a sufficient amount has been collected, their effects against Cryptococcus infection will be tested in-vivo. The study of these antibodies should provide insight about the role that glycosylation and the hinge region play in activating effector functions and cryptococcosis.
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Mr. Jose Rodriguez
Mentor: Gustavo Helguera, and Manuel Penichet
Title: Immunotherapy of Cancer
(Gustavo Helguera, Jose Rodriguez, and Manuel Penichet)
Jose Rodriguez is a fourth year senior majoring in Biophysics, and a second year MARC student. Jose previously worked in fungal genetics, acoustics, and imaging projects in the departments of Biochemistry and Physics. He currently works with Dr. Penichet in the departments of MIMG and Surgical Oncology, and has worked there for two years. Jose has participated in the PEERS, URFP, and EXROP programs, is now a MARC trainee, and will begin graduate studies next year with an HHMI Gilliam fellowship.
Jose uses antibody fusion proteins targeting tumor associated antigens to either directly destroy tumor cells or awaken an immune response against the tumor. His first project involved the use of antibody-cytokine fusion proteins targeting the extracellular domain of the human HER2/neu protein (a common marker of poor prognosis in breast, colon, ovarian, and other cancers). Previous investigations in the laboratory have demonstrated that combinations of these fusion proteins could be used as adjuvant vaccines against tumors expressing HER2/neu. Lately the lab has used antibody-cytokine fusion protein combinations to treat tumor-bearing mice as a test of their in vivo efficacy in a therapeutic setting. Studies in the laboratory also focus on a second type of antibody fusion protein designed as a cytotoxic delivery vehicle for biotynilated agents into cancer cells. The antibody-avidin fusion protein is a mouse/human chimeric IgG3 that targets the transferrin receptor, and is genetically fused to chicken avidin. Researchers in the laboratory previously demonstrated that it is intrinsically cytotoxic to multiple myeloma cancer cells expressing high levels of human Transferrin Receptor. Jose’s work in this project involves characterization of its binding properties and its interactions with the receptor. His latest work focuses on finding the affinity of this molecule for its receptor, tracking its rate of endocytosis in different cancer cell lines, and identifying the mechanism for its induction of lethal iron starvation.
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