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«Strengthening the Nation through Diversity, Innovation & Leadership in STEM San Antonio,Texas · October 3-6, 2013 Get Connected! Connect with the ...»

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In the cardiovascular system, the aorta serves as the passageway between the heart and body. Critical to its development are bone morphogenic protein (BMP) and transforming growth factor B (TGF-B) signaling pathways.

Within these pathways, inhibitory Smad6 and Smad7 (iSmads) function as downstream signal attenuators that fine tune the cells’ response to extracellular receptor activation. Previous experiments show that global knockout of Smad6 and Smad7 results in outflow tract malformations and other cardiac defects. What remains unknown is the role iSmads play in cardiac progenitor survival, migration, and differentiation. We hypothesized that phenotypic defects linked to iSmad knockout are associated with the role these molecules play in the progenitor cells of the second heart field (SHF), which give rise to the outflow tract, among other parts of the heart. Assessment of Smad6 knockout mice revealed previously unobserved defects, including double outlet right ventricle. Additionally, in situ hybridization of Smad6 and SHF marker Isl1 in E.10.5 embryos revealed that these molecules were expressed in the same anatomic regions, suggesting that iSmads may be expressed in the SHF. Conclusions from this project will lead to a better understanding of the role Smad6 and Smad7 play in aortic development. Given that developmental gene programs are often reactivated during disease, our studies may give insight into the progression of congenital heart defects, specifically aortic diseases.




Adrian Diaz, Seana Davidson.

University of Washington, Seattle, WA.

A symbiotic system has been discovered between the lumbricid earthworm Eiseniae fetida and the bacterial species Verminephrobacter eiseniae, which colonizes the excretory organs of the earthworm. V. eiseniae has been successfully cultivated and its genome sequenced. The earthworms do not acquire bacteria from the soil after hatching but must be colonized during development by bacteria deposited in the egg capsule. Past research has shown that both bacterial flagella and type IV pili are required for motility used in early colonization of embryonic earthworms. During these studies of bacterial migration into the embryo, it appeared the bacteria are directed into the embryo. In our study, we are exploring the requirement for bacterial chemotaxis during successful colonization of the host worm. The methyl-accepting chemotaxis protein (MCP), a transmembrane sensor protein in bacteria, 2 of which have been discovered in the V. eisenia genome, detect specific molecules in the extracellular matrix and this ligand binding produces a signal transduction that allows the flagella to commence tumbling and flagellar mechanisms. In this experiment, we develop methods to produce a viable chemotaxis assay to study possible molecular attractants necessary for localized colonization of the earthworm host by bacterial symbionts.



Francisco Mendez, Michael Dores, JoAnn Trejo.

University of California, San Diego, La Jolla, CA.

As part of the G-protein coupled receptor (GPCR) family, the purinergic P2Y1 receptor transmits signals from extracellular stimuli and activates signal transduction inside the cell. Degradation of GPCRs by endocytosis and sorting to lysosomes following agonist stimulation is important for regulating receptor signaling. Most GPCRs are modified with ubiquitin, a protein that acts as a targeting signal for the lysosome. A mutant of P2Y1 that cannot be ubiquitinated is degraded with the same kinetics as a wild-type receptor. We discovered a novel lysosomal sorting motif, YPXnL within the second intracellular loop of P2Y1, where X is any amino acid. We predict that mutation of the YPXnL motif will block ubiquitin-independent degradation of P2Y1. To disrupt the YPXnL motif, we generated a P2Y1


Y155A mutant using a site-directed PCR mutagenesis. Interestingly, we found out that this mutation causes a block in agonist-induced degradation. As a control, we analyzed the internalization of P2Y1 WT, and P2Y1 Y155A mutant.

The P2Y1 Y155A mutant does not affect internalization, suggesting that the mutation inhibits trafficking within the endosomal system. YPXnL motifs are binding sites for the endocytic adaptor protein ALIX, which is known to facilitate ubiquitin-independent lysosomal sorting of another GPCR, protease-activated receptor 1. We predict that ALIX and its binding partner ARRDC3 are required for the lysosomal degradation of the P2Y1 receptor. Experimentation on the purinergic P2Y1 receptor has extended our knowledge of endosomal sorting pathways and the roles of various proteins within those pathways in mammalian cells.



Carolina Zamora1, Needhi Bhalla2.

Hartnell College, King City, CA, 2University of California, Santa Cruz, Santa Cruz, CA.

1 The gene mdf-1 is a component of the spindle assembly checkpoint in mitosis and has been identified as a component of a meiotic prophase checkpoint that monitors chromosome interactions, suggesting it plays multiple roles to maintain genomic integrity. Another gene, pch-2, is also a component of the meiotic prophase checkpoint, and we want to test if pch-2 is also required for the spindle assembly checkpoint. The spindle assembly checkpoint is activated by a mutated zyg-1 gene in which the cells arrest in metaphase of mitosis. This arrest is dependent on genes such as mdf-1. The testing of zyg-1;pch-2 double mutants will help determine if pch-2 is required for the spindle assembly checkpoint. If the double mutant arrests in metaphase, unlike a zyg-1;mdf-1 double mutant, it is not required for the spindle assembly checkpoint. If it does not arrest in metaphase, like a zyg-1;mdf-1 double mutant, pch-2 is required for the spindle assembly checkpoint. Thus, these experiments will help determine if pch-2 is a newly identified component of the spindle assembly checkpoint.




DeAnna Ayupova, Reyna Valdez, Bhumi Patel, Rachna Sadana.

University of Houston-Downtown, Houston, TX.

Cancer is a collection of diseases hallmarked by uncontrolled cell division. Cancer treatment involves varying combinations of surgery, radiation, chemotherapy, and hormone therapy. Chemotherapy employs the use of drugs that kill the rapidly dividing cells (characteristic of cancer cells). Various chemotherapeutic drugs such as paclitaxel and vinblastine interrupt cell division by binding to tubulin, a protein responsible for spindle formation which is a critical step in cell division. For more than 50 years, tubulin-binding drugs have been used to treat cancer. Scientists are still in search of novel anti-cancer compounds targeting tubulin for 2 reasons: 1) patients develop resistance to existing drugs, and 2) tubulin is one of the most validated targets for cancer treatment. A series of synthetic analogues of marine natural products were evaluated for their cytotoxic effects on 3 different cancer cell lines. Our initial screen identified three compounds (BA-2, BA-3, and BH-6) that caused 50% cell death at concentrations less than 10 µM.

BA-2, BA-3, and BH-6 cause cell death via apoptotic mechanism when investigated for the mechanism of cell death using DNA fragmentation as marker. The compounds are currently being analyzed for their IC50 on various cancer cell lines.



Anthony Bortolazzo, Jodie Wu, Theresa Logan-Garbisch, Audrey Ford, Hilal Jarrar, David Do, Rachael French.

San Jose State University, San Jose, CA.

We use Drosophila melanogaster as a genetic model to study the relationship between ethanol exposure and developmental defects such as developmental delay, lowered survival, and behavioral changes including increased resistance to ethanol sedation and reduced development of tolerance. Recent studies have linked oxidative stress to the onset of ethanol-induced developmental abnormalities. Nearly all species experience oxidative stress, caused by an unbalance of the cell’s redox state. We have shown that developmental exposure to either paraquat or hydrogen peroxide, two well-established inducers of oxidative stress, phenocopies ethanol-induced phenotypes; in addition, hydrogen peroxide and ethanol act synergistically to cause developmental defects. We therefore hypothesize that oxidative stress resulting from developmental ethanol exposure explains a subset of the developmental defects we

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Hannah Collins1, Akela Kuwahara1, Robin Lerner2, Valeri Vasioukhin3, Claudia Petritsch2, Amy Sprowles1.

Humboldt State University, Arcata, CA, 2University of California, San Francisco, San Francisco, CA, 3Fred Hutchinson 1 Cancer Research Center, Seattle, WA.

Gliomas are the most common primary brain tumors. Their aggressive and highly invasive nature makes them a leading cause of cancer-related deaths by central nervous system tumors. Increased self-renewal and impaired differentiation capacities characterize both human tumor propagating cells and murine cells of origin in glioma.

Recent evidence suggests the WD40 protein lethal giant larvae (LGL) and C-Jun N-terminal kinases (JNK) may play an important role in regulating these properties. We hypothesize that LGL represses JNK activity such that loss of LGL results in JNK activation and phenotypes associated with malignancy such as increased self-renewal in glioma precursors. To test this model, neuroblasts isolated from adult Lgl loxp/loxp mice are treated with AD-CRE GFP and assayed for neurosphere formation, measurable increase in JNK activity, and increased anchorage-independent growth in soft agar. Preliminary evidence shows increased neuropshere formation in lgl -/- cells that is suppressed by the addition of the pharmacological JNK inhibiter SP600125. Understanding the relationship between JNK, LGL, and tumorigenicity in gliomas will provide valuable insight into the properties and treatment of these tumors.



Vanessa Cota, Jennifer Gilbert, Dana Byrd, Diana Chu.

San Francisco State University, San Francisco, CA.

During spermatogenesis, DNA is compacted into a highly condensed structure for delivery to the oocyte. In most animals, sperm-specific small nuclear basic proteins, called protamines, replace histones and are responsible for sperm DNA compaction. Problems occurring with the packaging of DNA, including differences in protamine expression, have been correlated with male infertility in humans. However, the precise function of protamines remains unclear. Through proteomic analysis, three nearly identical, small, highly basic, sperm chromatin-enriched proteins (SPCH-1, 2, and 3) were identified in C. elegans. Based on molecular characteristics and localization data, we hypothesize that SPCH proteins function as protamines. To determine SPCH function, we observed mutant populations of each gene individually by looking at their phenotypic expression. However, populations with the same spch-1 deletion allele display different phenotypes. One population is showing a previously unobserved “twitching” while the other population shows no apparent defect. To determine whether the “twitching” is related to the spchmutation or, if it was from a background mutation, we are backcrossing each mutant strain to the wild type. If unrelated to the mutation, we would expect to see both strains “un-twitch” after the backcross. Then, by doing progeny counts, we will determine the effect of spch-1 on fertility. We will then use cytology to view the localization of the two additional SPCH proteins in spch-1. The results of these studies of SPCH proteins in C. elegans may give insight into the function of protamines and their specific role in fertility.



AND DISEASE Grober Baltazar1, Peter Walentek2, Richard Harland2.

University of California, San Diego, La Jolla, CA, 2University of California, Berkeley, Berkeley, CA.

1 Mucociliary epithelia consist of mucus-secreting goblet cells, ciliated cells, and transmembrane proton/ion transporters that regulate homeostasis and gene expression. Certain respiratory diseases such as cystic fibrosis are caused by

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the accumulation of thick viscous mucus in the airway epithelium due to a recessive mutation of the cystic fibrosis transmembrane conductance regulator gene family (CFTRs). These genes regulate the concentrations of chloride and sodium ions across the airway epithelium, leading to infections of the carriers. Recently, it has been discovered that the ciliated epidermis of Xenopus laevis embryos can be used as a model system to study and understand the mechanisms of mucociliary epithelial homeostasis and development. Thus, this model organism can be used for research to find novel therapeutic treatments for diseases such as cystic fibrosis. Transmembrane proton regulators, e.g., ATP4 and ATP6, interact with the Wnt signaling pathway, which is required for development and function of mucociliary epithelia. In this study, we are cloning ion pumps and transporters (namely Duox1, Hvcn1, CFTR7, NHE1, NHE2, NHE3, and NHE4) for the first time in the frog Xenopus laevis. We want to analyze their expression in Xenopus laevis embryos and investigate their potential role in pH homeostasis of the ciliated skin epithelium and/or interaction with the Wnt signaling pathway. The long-term goal of these experiments is to gain a better understanding of proton/ ion secretion across ciliated epithelia and the mechanisms involved in human airway diseases.

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