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Biomedical Sciences Seminar Series

Spring 2010 Poster Presentation
April 30, 2010 at 1 pm
Physical Sciences Lobby

Poster #1- Rebecca Borja

Characterization of Yeast Casein Kinase I by Gene Knockout
Approach in Candida Albicans

The identification of key virulence factors in the initiation of oral candidiasis is the prime aim of this project. One of the up-regulated genes in response to oral epithelial cell interaction is YCK2, which is Casein kinase 1 (CK1) homologue. To assess the function of YCK2 related to initiate Candida infection, a YCK2 knockout strain will be constructed. High Fidelity PCR was used to amplify the cassettes. Enzyme Digestion was a new technique that was used to cut the wanted gene from the plasmid cassette. Transformation was used to induce homozygous recombination of +HIS cassette to the second YCK2 gene. Genomic DNA and Whole Cell PCR were used for the verification of the mutant strains. From the previous repeated transformations of the second YCK2 allele with ARG4 KO cassette, results show that the second YCK2 allele seems essential for the viability of the heterozygous mutant. In order to study the effects of this mutation, this mutant will be studied and its phenotype will be checked. The whole purpose of this revised experiment is to ultimately transform +HIS, +ARG and +URA into the isogenic strain of the BWP17 heterozygous mutant."



Poster #2-Adrian Esqueda

The discovery of novel inhibitory compounds against human apurinic endonuclease 1 (APE1). Human apurinic endonuclease 1 (APE1)

belongs to endonuclease family involved in the base excision pathway. APE1 is an enzyme that repairs damaged or mismatched bases in DNA by removing the base, leaving a sugar phosphate residue. APE-1 has been shown to be over-expressed in specific cancers including colon making it an ideal target for the development of new anti-cancer drugs. In combination with alkylating and oxidizing agents, it has been shown that inhibition of APE1 in cancer cells leads to a high degree of DNA damage eventually leading to apoptosis. Using an enzymatic assay and gel electrophoresis, training sets of compounds designed with an abasic DNA fragment as the template are tested for inhibition of APE1 activity

Poster #3- Kevin Linares

Connecting Developmental Processes to Virtual Worlds: The Case of Second Life.

Research suggests that young people’s online activities may be related to the developmental issues in their lives, such as establishing intimate relationships and formulating their identity (Subrahmanyam, Smahel, & Greenfield, 2006). We explore this possibility among emerging adults (Arnett, 2004) in the context of a Second Life (SL), which is “a 3D virtual world, where users can socialize, connect, and create using voice and text chat.” SL users are represented by adjustable, motion-enabled graphical representations called avatars. Scholars have speculated that such online worlds may allow users to leave their physical bodies behind and assume new, virtual selves. Evidence is accumulating that for adolescents, online and offline worlds may instead be connected. Less is known about the extent of connectedness between emerging adults’ online and offline worlds, and the present paper explores this question for this age group. The data reported here are drawn from a convenience sample of SL participants (emerging and older adults); the present sample consisted of 190 emerging adults, ages 18- to 29 years (m = 23.3, SD = 3.53) and 63% females. Participants completed an online questionnaire about their SL activities, real life demographics, and the Identity Styles Inventory (ISI-6G). The ISI classifies individuals into one of three identity styles: Information, Normative, and Diffuse/Avoidant. Descriptive analysis revealed real life (RL) and SL gender reported correlated at .94. Multiple regression analysis indicated that Information identity style predicted length of residency on SL, RL gender and normative identity style predicted having an importance of their online appearance and negatively predicted having multiple online identities, diffuse/avoidant identity style predicted having multiple online identities, and information identity style negatively predicted having formed a romantic relationship. All findings were significant p<.05.These results suggest that like adolescents, emerging adults may use virtual contexts to play out offline concerns; furthermore, offline variables such as gender and identity styles may account for their online activities.  


Poster #4-Na'il Mitchell

Applications of nanoparticles of magnetite and magnetite derivatives to assays of biomarkers in Alzheimer’s disease

Magnetite nanoparticles have gathered considerable attention in an array of different research fields due to its low toxic effects, paramagnetic properties, and controllable size. These properties make magnetite particularly viable in medical and biomedical research. We wish to use magnetite nanoparticles to detect amyloid beta fibrils (amyloid protein aggregates in brains of patients of Alzheimer’s disease) and as a staining agent for imaging of amyloid fibrils in tissue samples. Oxidized magnetite (maghemite; contains only ferric iron; very similar in properties to magnetite) has already been shown to be able to bind to and remove amyloid fibrils from solution (Hadas Skaat et al) when coated with either Congo red or thioflavin dyes (both dyes are capable of binding to amyloid fibrils). We plan to coat the nanoparticles with antibodies that bind specifically to amyloid beta fibrils. We have already shown that magnetite nanoparticles are capable of binding to anti-bovine serum albumin (BSA is a large protein that is common in the blood of cows) and we have used the resulting complex to remove BSA. Our magnetite derivative’s solubility is a desirable factor when relating to medical applications. Even at low concentrations (0.16 g/L) our magnetite derivative in water causes the whole media to become a uniform light brown/golden color. We envision that similar to thioflavin T, the absorption spectra of magnetite will undergo either a blue or red shift when it binds to the amyloid fibrils but will not undergo this shift when bound to non-fibrils/monomers, etc, which has not been reported at all to our knowledge. We will verify that magnetite nanoparticles have complexed with amyloid beta fibrils by carrying out dynamic light scattering measurements and looking for notable increases in particle sizes. It is reasonable to believe that magnetite nanoparticles will be a multi-purpose material to aid in the detection and elimination of the primary component (amyloid fibrils) of Alzheimer’s disease.  


Poster #5-Cynthia Reyes

Optimizing parameters for successful analysis of genome-wide MyoDbinding +/- AKT1/2 activity by ChIP-seq

MyoD is a basic helix loop helix (bHLH) transcription factor that is capable of inducing skeletal muscle differentiation in a variety of cell types. Together with the other myogenic regulatory factors (MRFs), MRF4, Myf5 and myogenin, MyoD directs the process of myogenesis. All four of the myogenic regulatory factors bind to the E-box consensus sequence (CANNTG) to initiate expression muscle specific gene products. AKT 1/2 has been shown promote the association of MyoD, by acting as effectors of the IGF1/PI3K/AKT pathway. Through the use of chromatin immunoprecipitation (ChIP) assays, the sites of MyoD binding in a variety of expressed genes have been identified. The results of these ChIP experiments yielded a number of unexpected binding sites, including sites not consisting the E-box consensus sequence. The presence of these unexpected sites suggests that the known mechanisms that apply to MRFs binding at the E-box do not apply to all binding sites. Our objective is to investigate these alternative mechanisms by inhibiting binding through the known mechanisms with the use of LY, a PI3-kinase inhibitor. We hypothesize that MyoD will bind even when AKT1/2 is inhibited; therefore another differentiation mechanism must be functional for those sites versus others. Before initiating the ChIP assay, the optimal parameters must be set. First identification of the time point at which C2C12 cells begin to differentiate even in growth conditions was studied. Differentiation was monitored by the presence of myogenin, which was monitored by RT q-PCR and ICC, for mRNA and protein respectively. Myogenin mRNA appeared as early as 12 hours of growth but myogenin protein was not seen until cells were differentiated (for 12 hours after 72 hours of growth). Next differentiation conditions in the presence and absence of our LY-AKT 1/2 inhibitor was monitored. Growing cells to 60% confluency and differentiating for 24 hours gave the largest ratio of myogenin mRNA accumulation between +LY plates and –LY plates. The last parameter studied was the optimal concentration of LY needed stop cell proliferation and inhibit differentiation. Through treating cells with different concentrations of LY and monitoring the effect on cell proliferation through nuclei count, and affect on differentiation through ICC, we were able to conclude that 20nM of LY is sufficient to stop proliferation and inhibit differentiation. Having identified these conditions as optimal, we are now ready to proceed with our preliminary ChIP here at CSULA in anticipation of the ChIP-seq that will be performed at Caltech.  


Poster #6-Evelyn Barrios

Congruences, Modular Arithmetic and its applications
|PI: Dr Tamez

Abstract: Formally describe congruent relations and the notation of modular arithmetic, theorems and definitions relevant to the understanding of its structure and the applications it has in mathematics and other subject areas. Trindad Cisneros has abstract, title, and poster documents


Poster #7-Anthony Obisesan

Photochemical oxidation of Polycyclic aromatic hydrocarbons in aqueous solutions

Polycyclic aromatic hydrocarbons (PAHs) are a group of carcinogenic contaminants formed from the incomplete combustion of hydrocarbons in the environment. Pyrene, a four membrane ring compound has been classified by the Environmental Protection Agency as one of twelve most carcinogenic PAHs in the environment. During photodegradation, previous studies have shown the formation of secondary compounds known as polycyclic aromatic compounds (PAC’s) such as 1,6- and 1,8-pyrenequinones. These compounds are known to be hazardous due to their increased polarity relative to the parent compound. In our experiment, the photolysis of pyrene has been studied in methanol and mixtures of methanol and water solutions. Our data shows a kinetic rate of decomposition of -2 x 10 -5 molecules sec-1 in an apparent first-order mode in 100 % methanol.


Poster #8- Wayne Warner

In silico Identification of F.D.A. approved drugs that bind to MDM2

The integrity of the p53 pathway is compromised in almost every human cancer. In approximately half of these cancers the tumor suppressor p53 is mutated or deleted and in at least 20 percent of the remaining cancers, MDM2 the master regulator of p53 is over-expressed resulting in a reduction of the functional concentration of p53. Several proof-of- concept studies have shown that drugging the MDM2 hydrophobic cleft is a therapeutically viable mechanism for rescuing p53 function resulting in cell cycle arrest, apoptosis and ultimately tumor shrinkage. The high cost and lengthy development pipeline for new drugs has highlighted the need to look at the repurposing of existing approved drugs. We used a lead library of 3,200 Food and Drug Administration (F.D.A.) approved drugs and computationally assessed their interaction with the hydrophobic cleft of MDM2. This in silico docking procedure generated a hit list of three hundred drugs. Post screen decision-making involved an analysis of binding energies, chemical similarity to known inhibitors and the significance of the drug interactions with the active site. We report on the identity of these drugs here.


Poster #9- Michael Mendoza

Protein Purification of Wild Type p53 DNA Binding Domain

The ability of a cell to appropriately respond to stresses and maintain its redox potential is essential for life. Stressors that can alter the redox potential in the cell include UV light, reactive oxygen intermediates (ROI), heat shock, ionizing radiation, and DNA damage. Adverse changes in redox potential can alter the tertiary structure of proteins and affect their ability to perform their functions. It has been shown that the tumor suppressor protein p53 is subjected to a reversible form of modification called S-glutathionylation. (Velu et al. 2007) p53 is tetramer protein, containing two identical dimer subunits that together form a fully functional protein which can bind to DNA. Human p53 contains six solvent exposed cysteine residues: Cys 124, Cys 176, Cys 182, Cys 229, Cys 242, and Cys 277 within the DNA binding domain. Maintaining the reduced states of these cysteine residues is vital for p53 to remain functional. .


Poster #10-Deisy Contreras

Role of Protein Kinase-A and Casein Kinase Pathways in Candida albicans Resistance to Antimicrobial Peptides

Abstract: Candida albicans is a commensal organism of the human digestive system and vaginal tract as well as in other warm-blooded animals (Scherer and Magee, 1990). In imunocompromised patients, Candida is able to cause a variety of infections. C. albicans causes at least 80% of episodes of oropharyngeal candidiasis in patients with HIV/AIDS (Revankar, et al, 1998). Furthermore, C. albicans is the most common cause of candidiasis in other patients, such as those with Sjogren’s syndrome and diabetes mellitus, and who have received radiation therapy for head and neck cancers (Redding, S., et al, 1999, Willis AM et al, 1999). The predominance of C. albicans as a cause of candidiasis suggests that this organism possesses unique characteristics that enable it to colonize the oropharynx and, when host defense mechanisms are weakened, cause oropharyngeal candidiasis. Therefore, it is implemented that host immune system plays key role in controlling C. albicans outgrowth in human host. Previous gene knockout study on C. albicans have identified that two C. albicans mutant strains lack Casein kinase activities are hypersusceptible to protamine, which is one of the antimicrobial peptide commonly used in the laboratory (Chiang et al, 2007, Park et al, manuscript in preparation). Therefore, we hypothesized that YCK2 and CKA2, which encode the catalytic domain of Casein kinase I and II activities are involved in the signaling pathway triggering C. albicans resistance to antimicrobial peptide. Previous studies have isolated an antimicrobial peptide-resistant strain of C. albicans, which has altered gene expression of SSD1. It is also demonstrated that null mutation of SSD1 rendered C. albicans hypersusceptible to antimicrobial peptides (Gank et al, 2008). These results suggest that SSD1 governs C. albicans resistance to antimicrobial peptide activity. Therefore, we hypothesized that YCK2 and CKA2 encoding casein kinase activities are likely closely related to SSD1 in governing the response to antimicrobial peptide. .


Poster #11- Ana Ramirez

The Effects of N-acetylcysteine on Nicotine Self-Administration and Cue-Induced Reinstatement of Nicotine-Seeking in Rats

Cigarette smoking is the leading preventable cause of death in the United States. The addictive properties of tobacco are primarily attributed to nicotine. Chronic nicotine administration is hypothesized to disrupt the function of the main excitatory neurotransmitter in the brain, glutamate, and lead to a hypoglutamatergic state. N-acetylcysteine (NAC) is a compound that may reverse dysfunctions in glutamate transmission resulting from chronic nicotine exposure. NAC activates the cystine-glutamate exchanger, xc-, which exchanges intracellular glutamate for extracellular cystine resulting in increased glutamate levels in the extracellular space. Recently, NAC has been shown to reduce the number of cigarettes smoked daily by humans, when alcohol consumption was taken into consideration. In this study we assessed the effects of acute and repeated administration of N-acetylcysteine on nicotine self-administration and cue-induced reinstatement of nicotine-seeking behavior in rats. Intravenous nicotine self-administration allows for direct assessment of the reinforcing effects of nicotine and cue-induced relapse to nicotine-seeking is a putative animal model for relapse to drug-seeking in humans. Rats trained to self-administer nicotine intravenously were treated with one of four doses of NAC (0, 30, 60, and 90 mg/kg i.p.), 2.5 hours prior to the nicotine self-administration sessions. After the completion of nicotine self-administration testing, rats underwent extinction training and were tested in the cue-induced reinstatement procedure. In this study we found both acute and repeated NAC treatment significantly reduced nicotine self-administration, and NAC treatment significantly attenuated cue-induced reinstatement in rats. Reduction of nicotine self-administration and attenuation of cue-induced reinstatement by NAC indicate that NAC is a potential treatment for nicotine addiction and the prevention of relapse to tobacco smoking.



Poster #12- Henry Valle

Next Generation MRI Contrast Agents: Systems Bearing Multiple Gd(III) Ions Complexed to Adamantane Derivatives

Next Generation MRI Contrast Agents: Systems Bearing Multiple Gd(III) Ions Complexed to Adamantane Derivatives Henry U. Valle Abstract. In situations where insufficient image contrast adequately differentiates anatomy or pathology in magnetic resonance imaging (MRI), a contrast agent is administered that usually incorporates a paramagnetic metal (most commonly gadolinium). There is a need for improved contrast agents. Most of the MRI contrast agents currently in clinical use suffer from all or most of the following limitations: 1) they are able to complex and exchange only one water molecule at a time; 2) it is difficult to control the solubility of their complexes; 3) it is difficult to control their bio-distribution; 4) most are mononuclear, bearing each only one metal nucleus to assist in water relaxation. The work proposed here is the preparation of next-generation MRI contrast agents, small molecules that will bear 2, 3, And 4 metal centers, and bear functionality that will allow for control of bio-distribution and solubility in biological fluids. We also propose the synthesis of gold nano-particle-based MRI agents. The modular synthetic approach takes advantage of the 1,3,5,7-tetrakis(amino-methyl)adamantane building block developed by this laboratory. .


Poster #13- Vanessa Velazco

Regulation of Human Defensin 5 by Toll-like Receptor Ligands and Proinflammatory Cytokines in Female Genital Epithelial Cells

Epithelial cells lining the mucosal surfaces are part of the first line of defense against microbial infection. A crucial mechanism of defense against infectious diseases employed by epithelial cells is the production of antimicrobial peptides such as defensins. We have recently shown that human defensin 5 (HD5) is increased in the inflamed female genital tract and in the penile urethra in men with STIs. Thus, we hypothesize that HD5 is regulated by microbial products and pro-inflammatory cytokines in genital epithelial cells. Using an epithelial cell culture model with A2EN cell lines grown at air-liquid interface, we quantified the production of HD5 in the presence or absence of microbial products such as lipopolysaccharide (LPS) and peptidoglycan (PG) and proinflammatory cytokines such as IL-1?, IFN?, and IL-17A. The RNA was extracted from cell pellets and analyzed by quantitative real-time PCR (RT-PCR). Cell culture supernatants from apical and basolateral compartments were analyzed by Western immunoblotting for HD5. It was found that HD5 mRNA was up-regulated at least 100 fold by IL-17A and 20 fold by LPS. Moreover, HD5 peptide appeared to be about 2 fold increased upon IL17 stimulation reaching about 20 ng/mL concentrations in cell secretions.


Poster #14-Alon Roy Agua

Progress Towards the Chemistry of Phosphines with Singlet Oxygen

The chemistry of phosphines with singlet oxygen consists of several competing pathways: Intermolecular oxygen atom transfer , intramolecular hydroxylation , as well as rearrangment to a phosphinate ester has all been observed. We have systematically studied a biphenylphosphine (1) under different conditions, i.e. protoc vs. aprotic solvents, as well as different temperatures. By changing the amount of methanol-d4 added and temperature in the reaction of di-tert-butyl(2',4',6'-triisopropylbiphenyl-2-yl) phosphine (1) with singlet oxygen, different product ratios of di-tert-butyl(2',4',6'-triisopropylbiphenyl-2-yl)phosphine oxide (5), tert-butyl(2',4',6'-triisopropylbiphenyl-2-yl)phosphinate (4), and epoxides were obtained. We suggest that the initial intermediate, di-tert-butyl(2',4',6'-triisopropylbiphenyl-2-yl) phosphadioxirane (2), can undergo intramolecular rearrangment to form the phosphinate (4) and intermolecular oxygen transfer to make the phosphine oxide (5). However, under protic conditions, the phosphadioxirane (2) turns into di-tert-butyl(2',4',6'-triisopropylbiphenyl-2-yl) hydroperoxy phosphorane (3), which will undergo intermolecular oxygen transfer or an intramolecular epoxidation, but no rearrangment to a phosphinate ester (4). At larger volume of methanol-d4, less phosphine oxide (5) formed but more epoxide was observed. Increasing the temperature leads to greater formation of both epoxide and phosphinate (4). The greatest amount of epoxides was formed (56% of the product distribution) at 20% methanol-d4 and 60 °C.


Poster #15-Jennifer Uyere


Functional Analysis and Characterization of the Yeast Casein Kinase 3 Gene in Comparison to the Yeast Casein Kinase 2 Gene in C. albicans

Although pathogenic and severely life threatening, Candida albicans is a fungus that exists as a part of the human normal flora, specifically in the digestive system and vaginal tract. In individuals with compromised immune systems, Candida is able to cause a variety of infections. Most notably, C. albicans is responsible for causing candidiasis in the oral pharyngeal cavity. The ability of C. albicans to cause candidiasis in immunocompromised individuals implies that the pathogenic fungus attacks under extremely vulnerable conditions; therefore, it has been classified as an opportunistic pathogenic fungus. C. albicans causes approximately 80% of episodes of oropharyngeal candidiasis in patients with HIV/AIDS (Revankar, et al, 1998). The mechanisms by which C. albicans causes candidiasis are not fully understood, therefore, treatments are limited to fungistatic drugs that are not very effective in treating infections (Marrazzo, 2002). Several studies have revealed that the initial step to causing infection is that C. albicans invades the epithelium. Thus, the ability of C. albicans to damage epithelial cell lining is one of the key virulence factors involved in the initiation of candidiasis (Park et al, 2005). Previous studies in our laboratory have shown through transcriptional analysis that the C. albicans Yeast Casein Kinase 2 gene, YCK2, is remarkably upregulated and plays a key functional role in the interaction between C. albicans and oral epithelial cells. In addition to finding the YCK2 gene in C. ablicans and its function, the Yeast Casein Kinase 3 gene, YCK3, was discovered. At the genomic level, YCK3 shares over 80% homology to YCK2. Moreover, the function of casein kinase activities in C. albicans is not completely understood. In Saccharomyces cerevisiae, a non-pathogenic fungus that shares a great deal of homology to C. albicans, YCK2 is required for morphogenesis and cytokines (Robinson et al., 1992, 1993). Therefore, it has been hypothesized that YCK2 also plays an important role in morphogenesis in C. albicans, and thus influences virulence. The purpose of my research is to explore the specific phenotype present of the YCK3 gene to help in further understanding the fungal pathogenesis pathways, as well as elucidate the relationship between YCK2 and YCK3 to aid in understanding the functional redundancy, if any.


Caridad Wilson
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