Life Sciences Commons^™

Structural Characterization Of Black Widow Spider Dragline Silk Proteins Crp1 And Crp4, Mikayla Shanafelt

University of the Pacific Theses and Dissertations

Spider dragline silk is a biomaterial with outstanding material properties, possessing high-tensile strength and toughness. In nature, dragline silk serves a central role during spider locomotion and web construction. Today, scientists are racing to elucidate the molecular machinery governing silk extrusion, attempting to translate this knowledge into a mimicry process in the laboratory to create synthetic fibers for a wide range of different applications. During extrusion, it has been established that biochemical and mechanical forces govern spidroin folding, aggregation, and assembly. In black widow spiders, at least 7 different proteins have been identified as constituents of dragline silk fibers. These …

Transcriptome Analysis For Novel Peptide In Breeding Gland Of Hymenochirus Boettgeri, Minjin Ko

University of the Pacific Theses and Dissertations

The primary goal of the project was to find nucleotide sequences potentially encoding a pheromone from the breeding gland of Hymenochirus boettgeri. The reasons in searching for the sequence of a pheromone were to better understand the organism and to use the information for application in reproduction of other species. Due to climate change and rampant deforestation, such as in Africa’s Congo Basin, many amphibian species are being threatened. With these increasing threats, a viable option for the future may be breeding in captivity for the amphibian species. Pheromone characterization from the breeding gland of Hymenochirus boettgeri may help with …

A Trio Of Sigma Factors Control Hormogonium Development In Nostoc Punctiforme, Alfonso Gonzalez Jr.

University of the Pacific Theses and Dissertations

Cyanobacteria are prokaryotes capable of oxygenic photosynthesis, and for many species, nitrogen fixation, giving cyanobacteria an important role in global carbon and nitrogen cycles. Furthermore, multicellular filamentous cyanobacteria are developmentally complex, capable of differentiation into different cell types, including cells capable of nitrogen fixation and cells for motility, making them an ideal platform for studying development, as well as for practical use in biotechnology. Understanding how developmental programmes are activated require an understanding of the role of alternative sigma factors, which are required for transcriptional activation in bacteria. In order to investigate the gene regulatory network and to determine the …

Characterization And Analysis Of Proteins Secreted By The Mutant Pichia Pastoris Strain, Bgs13, Christopher Alan Naranjo

University of the Pacific Theses and Dissertations

The methylotrophic yeast Pichia pastoris has been utilized for heterologous protein expression for research, clinical, and industrial purposes to produce thousands of recombinant protein products. Because P. pastoris secretes very few of its own proteins, the exported recombinant protein is the major polypeptide in the extracellular medium, making purification relatively easy. Unfortunately, a disadvantage to the programmed export is that some recombinant proteins intended for secretion are retained within the cell and may be subsequently degraded. A mutant strain isolated in our lab, containing a pREMI-derived disruption of the BGS13 gene, has displayed elevated levels of secretion for a variety …

Phylogenomic Analyses Clarify Butterfly Species Within The Genus Speyeria Despite Evidence Of A Recent Adaptive Radiation, Erin Thompson

University of the Pacific Theses and Dissertations

The North American genus Speyeria is an especially challenging radiation of butterflies due to ongoing hybridization, incomplete lineage sorting, and similar morphological characters among species. Adaptive radiations often require considerable evidence in order to resolve the evolutionary relationships of closely related individuals. Previous studies of this genus have found paraphyly among species and have been unable to disentangle these taxa due to a lack of data and/or incomplete sampling of the genus. As a result, the interspecific relationships among Speyeria remain unresolved. In an attempt to achieve phylogenetic resolution of the genus, we conducted population genomic and phylogenomic analyses of …

Development Of A Biomarker Panel For Identifying Stressed Marine Mammals, Laura Pujade

University of the Pacific Theses and Dissertations

Increasing anthropogenic disturbance in marine ecosystems such as fishing, oil-drilling, and noise pollution can have detrimental effects on the reproduction and survival of apex predators such as marine mammals. Stress activates the hypothalamic-pituitary-adrenal (HPA) axis, resulting in increased circulating glucocorticoid (GCs) hormones, which alter expression of target genes encoding metabolic enzymes and other mediators of stress. Prolonged HPA axis stimulation may increase catabolism of nutrient stores and suppress immune and reproductive functions, impacting the fitness of marine mammals. GCs measurements are used to identify wild animals experiencing stress. However, these measurements may not be sensitive enough to distinguish between an …

Expression, Purification, And Characterization Of Recombinant Basic Fibroblast Growth Factor In Pichia Pastoris, Henry Hieu Minh Le

University of the Pacific Theses and Dissertations

Wounds in the mouth, occurring after oral surgery, take time to heal. No ointment can be added to help with the healing process because mouth saliva will constantly wash it away. In order to combat this problem, we propose engineering a normal flora microbe to grow at the site of injury and secrete a recombinant growth factor to promote healing of the damaged tissue. Our goal is to have the yeast Pichia pastoris produce human basic fibroblast growth factor (bFGF), which aids in cellular proliferation. P. pastoris is a good choice for this application because not only is it considered …

Knob-Socket Investigation Of Stability And Specificity In Alpha-Helical Secondary And Quaternary Packing Structure, Taylor Renee Rabara

University of the Pacific Theses and Dissertations

The novel knob-socket (KS) model provides a construct to interpret and analyze the direct contributions of amino acid residues to the stability in α-helical protein structures. Based on residue preferences derived from a set of protein structures, the KS construct characterizes intra- and inter-helical packing into regular patterns of simple motifs. The KS model was used in the de novo design of an α-helical homodimer, KSα1.1. Using site-directed mutagenesis, KSα1.1 point mutants were designed to selectively increase and decrease stability by relating KS propensities with changes to α-helical structure. This study suggests that the sockets from the KS Model can …

Investigating The Effect Of Rutaecarpine On The Benzo[A]Pyrene-Induced Dna Damage In Vitro, You Li

University of the Pacific Theses and Dissertations

Benzo[a]pyrene (BaP), is one of the most potent mutagens and carcinogens known. It requires metabolic activation through cytochrome P450 (CYP)1A1 to yield the ultimate carcinogenic metabolite, benzo[a]pyrene-7,8-dihydrodiol-9,10-epoxide (BPDE). BPDE can bind to DNA and form predominantly covalent (+) trans adducts at the N2 position of guanine causing DNA damage. Rutaecarpine (RTC) is an herbal medicine that has been used to treat several diseases such as headache, hypertension, gastrointestinal disorders, amenorrhea, and anti-inflammation. It has also been reported as a potent inducer of CYP enzymes, including CYP1A1, and CYP1A2. The mechanisms underlying up-regulation of CYP1A1 by RTC is dependent on aryl …

Blubber Transcriptome And Proteome Responses To Repeated Adrenocorticotropic Hormone Administration In A Marine Mammal, Jared Deyarmin

University of the Pacific Theses and Dissertations

Chronic physiological stress impacts animal fitness by catabolizing metabolic stores and suppressing reproduction and immunity. This can be especially deleterious for capital breeding carnivores, such as marine mammals, which rely on lipid stores accrued during intensive foraging to sustain prolonged periods of fasting associated with reproduction. Therefore, chronic stress may cause a decrease in fitness in these animals, leading to population declines and potentially detrimental shifts in food web dynamics as a result. However, the impacts and indicators of chronic stress in animals are currently poorly understood. To identify downstream mediators of repeated stress responses in marine mammals, adrenocorticotropic hormone …

Characterization Of The Roles Of Ppp1r15a (Gadd34) And Ppp1r15b (Crep) In Er Stress-Induced Apoptosis In Vivo, Danielle Hicks

University of the Pacific Theses and Dissertations

Failure to balance synthesis, folding and degradation of secreted proteins results in the accumulation of unfolded proteins in the endoplasmic reticulum, termed ER stress. Cells respond to ER stress by activating a signaling pathway known as the unfolded protein response (UPR). The UPR induces phosphorylation of eIF2α to attenuate global protein translation, allowing the ER to clear misfolded proteins. Opposing this function, eIF2α phosphatases contain a catalytic subunit, Protein Phosphatase 1, and either of two homologous scaffolding subunits, GADD34 and CReP. Inhibition of eIF2α phosphatases has been shown to prolong UPR signaling and promote survival in many cells types and …

City Limits: Heat Tolerance Is Influenced By Body Size And Hydration State In An Urban Ant Community, Dustin Jerald Johnson

University of the Pacific Theses and Dissertations

Cities are rapidly expanding, and global warming is intensified in urban environments due to the urban heat island effect. Therefore, urban animals may be particularly susceptible to warming associated with ongoing climate change. Thus, I used a comparative and manipulative approach to test three related hypotheses about the determinants of heat tolerance or critical thermal maximum (CTmax) in urban ants—specifically, that (1) body size, (2) hydration status, and (3) preferred micro-environments influence CTmax. I further tested a fourth hypothesis that native species are particularly physiologically vulnerable in urban environments. I manipulated water access and determined CTmax for 11 species common …

Evaluating Methods Of Obtaining Male Pheromone From Hymenochirus Sp. Using Analytical Chemistry, Vincent Wing-Kun Leung

University of the Pacific Theses and Dissertations

Male Hymenochirus sp. frogs are known to release pheromone that attracts females of the same species. Four methods for collecting secretions containing pheromone in Hymenochirus sp. were tested: norepinephrine injection, gonadotropin-releasing hormone injection, homogenization of gland tissue, and electrostimulation of the skin over the breeding gland area. The samples collected were analyzed using high-performance liquid chromatography (HPLC) and mass spectrometry. The HPLC chromatograph for the male norepinephrine sample contained a peak at 6.4 min that was not in the female norepinephrine sample HPLC chromatograph. The male norepinephrine sample mass spectrum had a peak of m/z 292.0 not in the female …

Design And In Vitro Characterization Of Lipids With A Ph-Sensitive Conformational Switch And Their Liposomes For Anticancer Drug Delivery, Shen Zhao

University of the Pacific Theses and Dissertations

The traditional anticancer drugs are distributed in vivo through systemic blood circulation with a very small portion reaching the tumor site. Targeted drug delivery systems are developed in efforts to concentrate the drug molecules in the tissue of interest while reducing the drug distribution to healthy tissues to reduce the side effects. Liposomes are colloidal systems composed of amphiphilic molecules that assemble into vesicle structures in aqueous media. They are common carriers for targeted drug delivery with the advantages of low toxicity, low immunogenicity and the ability of encapsulating both lipophilic and hydrophilic drugs.

Prior research indicated the advantages of …

Spatial And Temporal Dynamics Of Batesian Mimicry Between Adelpha Californica And Limenitis Lorquini, Louis Albert Prusa

University of the Pacific Theses and Dissertations

Conspicuous coloration is one of the main ways that animals communicate. The use of eye-catching color patterns to warn predators of an unprofitable trait is referred to as aposematism. Once predators learn to recognize the color pattern, a new signaling niche becomes available where other species can share the same signal. This mimicry niche can involve a “hide in plain sight” strategy by mimicking or parasitizing this signal, with mimics lacking the defense and associated costs that make them unprofitable. This is termed Batesian mimicry, and it decreases predation by taking advantage of the memory and learning of the predator …

Unknown And Cryptic Diversity In The Adelpha Serpa-Group, Cassidi Rush

University of the Pacific Theses and Dissertations

The tropics are host to incredible insect diversity. One of the most charismatic groups that exhibit this diversity are the butterflies. Despite the tropical butterfly fauna being heavily researched, there remains much hidden diversity in the form of undescribed life histories and cryptic species. This is especially true among the species rich Nymphalidae, the brush-footed butterflies. Species in the genus Adelpha are known to be “the most trying taxonomically of all nymphalids” DeVries (1987), and as such are fruitful ground for uncovering unknown diversity. About half of the species within Adelpha have undescribed life histories, while A. serpa stands out …

Should I Stay Or Should I Go? Complex Environments Drive The Developmental Plasticity Of Flight Capacity And Flight-Related Tradeoffs, Jordan R. Glass

University of the Pacific Theses and Dissertations

Animals must balance multiple, fitness-related traits in environments that are complex and characterized by co-varying factors, such as co-variation in temperature and food availability. Thus, experiments manipulating multiple environmental factors provide valuable insight into the role of the environment in shaping not only important traits (e.g., dispersal capacity or reproduction), but also trait-trait interactions (e.g., trade-offs between traits). We employed a multi-factorial design to manipulate variation in temperature (constant 28°C vs. 28±5°C daily cycle) and food availability (unlimited vs. intermittent access) throughout development in the sand field cricket, Gryllus firmus. We found that fitness-related, life-history traits and trait trade-offs can …

Influence Of The Estrous Cycle And Female Sex Hormones On Ghb Toxicokinetics, Hao Wei

University of the Pacific Theses and Dissertations

Gamma-Hydroxybutyrate (GHB) is an endogenous short-chain fatty acid formed from Gamma-aminobutyric acid (GABA). Clinically, GHB is marketed in the United States as Xyrem to treat narcolepsy with cataplexy and in Europe for the treatment of alcohol withdrawal and narcolepsy. However, the illicit use and abuse of GHB occurs due to its sedative/hypnotic and euphoric effects. Monocarboxylate transporters (MCTs and SMCTs) are integral membrane proteins that control the bidirectional transport of endogenous substrates including lactate, acetate and pyruvate. They have also been found to transport and mediate the clearance and distribution of GHB. MCTs demonstrate a wide tissue distribution, including brain, …

Production Of Synthetic Spider Silk, Ryan Matthew Hekman

University of the Pacific Theses and Dissertations

Spider silk is a material that both has impressive mechanical properties and is also environmentally friendly. Though there are limitless potential engineering applications for such materials, industrial production of spider silk has proven to be challenging. Farming silk from spiders, as is done with silkworms, is not a viable option for large-scale production of spider silk due to the venomous and predatory nature of spiders. Here, an attempt is made to express synthetic spider silk minifibroins heterologously in Escherichia coli, to purify the recombinant spidroins from cell lysate, and to spin them into artificial fibers through a biomimetic process. Silk …

Vector Competence Of Northern California Mosquitoes For Dirofilaria Immitis (Spirurida: Onchocercidae), Rebekah Lynn Dian

University of the Pacific Theses and Dissertations

Dirofilaria immitis, commonly known as dog heartworm, is a dangerous filarial nematode that is transmitted by a mosquito vector. Although this parasite is historically localized to regions closer to the equator, the last few decades have experienced an increase in the amount of dog heartworm cases in Northern California, most notably in Lake and San Joaquin Counties. The primary vector of D. immitis in Northern California is Aedes sierrensis, but it is not prevalent enough in some counties to explain the increase in infections. Because of this, more prevalent and abundant species should be assessed for their ability to transmit …

Behavioral Patterns Of Blinded Vs. Mock-Blinded Male Individuals In The Presence Of Females: Analysis Of The Role Of Eyes In Euphilomedes Carcharodonta Mating, Alexis Arenz

University of the Pacific Theses and Dissertations

Sexual dimorphism is exhibited throughout various organisms and takes an extensive variety of forms, although rarely does it take the form of eye-reduction within a population. This poses the question of whether this dimorphism arises due to sexual selection in order to increase reproductive success or rather from ecological selection as a result of sex-specific niche partitioning. In this case, we study an organism that displays a drastic form of sexual dimorphism, in that the males have large complex lateral eyes whereas the females lack eyes and merely have rudimentary lateral eye-spots. Euphilomedes carcharodonta is a small crustacean native to …

Evidence That A Partner-Switching Regulatory System Modulates Hormogonium Motility In The Filamentous Cyanobacterium Nostoc Punctiforme, Kelsey Wynne Riley

University of the Pacific Theses and Dissertations

Partner-switching regulatory systems (PSRSs) are utilized by many different bacteria to regulate a wide array of cellular responses, from stress response to expression of virulence factors. The filamentous cyanobacterium Nostoc punctiforme can transiently differentiate motile filaments, called hormogonia, in response to various changes in the environment. Hormogonia utilize a Type IV pilus (T4P) complex in conjunction with a secreted polysaccharide for gliding motility along solid surfaces. This study identified three genes, designated hmpU, hmpW, and hmpV, encoding the protein components of a PSRS involved in regulation of hormogonium motility in N. punctiforme. Although mutant strains with in-frame deletions in hmpU, …

Endocrine Responses To Repeated Adrenocorticotropic Hormone Administration In Free-Ranging Elephant, Molly Mccormley

University of the Pacific Theses and Dissertations

Understanding the physiological response of marine mammals to anthropogenic stressors can inform marine ecosystem conservation strategies. Stress stimulates release of glucocorticoid (GC) hormones, which increase energy substrate availability while suppressing energy-intensive processes. Exposure to repeated stressors can potentially affect an animal’s ability to respond to and recover from subsequent challenges. To assess the endocrine response of a marine mammal to repeated stressors, we administered adrenocorticotropic hormone (ACTH) to free-ranging juvenile northern elephant seals (Mirounga angustirostris; n=7) once daily for four days. ACTH administration induced significant, but transient (<24 h) elevation in circulating cortisol levels (p < 0.0001). These increases did not vary in magnitude between the first ACTH challenge on day 1 and the last challenge on day 4. In contrast, aldosterone levels remained elevated above baseline for at least 24 hours after each ACTH injection (p < 0.001), and responses were greater on day 4 than day 1 (p < 0.01). Total triiodothyronine (tT3) levels were decreased on day 4 relative to day 1 (p < 0.01), while reverse triiodothyronine (rT3) concentrations increased relative to baseline on days 1 and 4 (p < 0.001) in response to ACTH, indicating a suppression of thyroid hormone secretion. There was no effect of ACTH on the sex steroid dehydroepiandrosterone (DHEA). These results suggest that elephant seals are able to mount adrenal responses to multiple ACTH challenges. However, repeated stress results in facilitation of aldosterone secretion and suppression of tT3, which may impact osmoregulation and metabolism. We propose that aldosterone and tT3 are informative additional indicators of repeated stress in marine mammals.

Peptide-Drug Conjugate For Her2-Targeted Drug Delivery, Yan Wang

University of the Pacific Theses and Dissertations

Recent strategies for anticancer drug design have been focused on utilizing antibody as a drug or targeted moiety for targeted drug delivery. Antibody−drug conjugates (ADCs) have become a promising new class of targeted therapeutic agents for treatment of cancer. ADCs are designed to preferentially direct a cytotoxic drug to a cell-surface antigen recognized by an antibody. However, there are some challenges in developing ADCs, such as limited solid tumor penetration, high manufacturing costs and antibody-drug stoichiometry. Smaller molecules such as peptides have been shown to specifically bind to cancer related targets. These peptides can be used to form peptide-drug conjugates …

Dna Lesions Produced From The Reaction Of Diols And 5-Formylcytosine And Their Effects On Dna Replication, Brock Allen

University of the Pacific Theses and Dissertations

Nucleic acids are complex macromolecules that are susceptible to both endogenous and exogenous damage. This study explored damage resulting from interactions with environmental nucleophilic toxins, such as a variety of diols and amines found in industry. These nucleophiles can react with electrophilic groups, such as 5-formylcytosine. 5-formylcytosine is an oxidation product of the epigenetic base 5-methylcytosine. It is typically removed by thymine DNA glycosylase (TDG) but is known to accumulate in the genome, making the formyl group susceptible to attack. In this study we used GC/MS and ESI-MS to show that DNA lesions from the nucleophilic addition reaction of toxins …

Investigations On Cancer Cell Biological Effects Of Cdk8 Inhibitor Q-12, Zhixin Lu

University of the Pacific Theses and Dissertations

Over the past two decades, protein kinases have been intensively investigated as targets to treat neoplastic diseases. Many protein kinase inhibitors not only have therapeutic potential but are becoming invaluable reagents for the study of cell signaling.

We aspired to use our Cyclin-Dependent Kinase 8 inhibitor, Q-12, as a probe for biomarker discovery for CDK8 inhibitor sensitive tumor types. Q-12 shows potent inhibition of cell viability and induction of apoptosis process in some triple-negative breast cancer and colorectal cancer cell lines in vitro. Western blot results indicate that the reduction of STAT1 phosphorylation could be a robust indicator of CDK8 …

Determining The Role Played By Aryl Hydrocarbon Receptor (Ahr) In The Colon Carcinoma Tumor Model, Poonam Yakkundi

University of the Pacific Theses and Dissertations

Aryl hydrocarbon receptor (AHR), commonly known as an environmental sensor involved in the metabolism and elimination of xenobiotic substances, is also an important modulator in the development and functioning of the immune system. AHR expression is varied in the T cell subsets with the highest expression in T-helper 17 and T regulatory cells. Work from many researchers has suggested that AHR can act as a tumor promoter or a tumor suppressor depending on the tumor type. Our goal is to understand the role played by AHR in MC38 syngeneic colon carcinoma tumor model. In the absence of AHR, MC38 tumor …

Neuromuscular Control Of The Calling Apparatus In The Túngara Frog (Engystomops Pustulosus), Kiran Kaur Grewal

University of the Pacific Theses and Dissertations

Male túngara frogs can add a distinctive note ("chuck”) to their mating call. Production of the chuck involves vibrating a pair of laryngeal fibrous masses that is attached to the vocal cords. The muscular control of this mechanism remains unknown. Recent studies revealed a split in the laryngeal dilator muscle, which unveiled the deep dilator as a novel laryngeal muscle with unique attachments, innervation, and (likely) function. The deep dilator may position the fibrous masses for chuck production. The goals of this study were 1) to confirm the innervation of the novel muscle through electrophysiology; and 2) to determine the …

Studies On Development In Euphilomedes Ostracods: Embryology, Nervous System Development, And The Genetics Of Sexually Dimorphic Eye Development, Kristina Koyama

University of the Pacific Theses and Dissertations

Model organism studies have been fundamental in understanding evolutionary and developmental biology. However, non-model organisms present opportunities to study unique characteristics and as comparisons to model organisms, leading us toward broader and more relevant perspectives on diversity. The Euphilomedes genus of ostracods is an example of a non-model group with potential for evolutionary and developmental studies.

Ostracoda is an ancient, basally branching lineage of Crustaceans with a complete and prodigious fossil record. Despite the group’s promise for evolutionary studies, much remains unknown about the basic biology of this clade. There are a limited number of embryogenesis studies in Ostracoda; here, …

Isolating The Targets Of Six Transcription Factor In Ephydatia Muelleri And Identifying The Role Of The Superoxide Dismutase 6 In Host Immune Response To Trichomonas Vaginalis, Gurbir Kaur Gudial

University of the Pacific Theses and Dissertations

Sponges are the descendants of the oldest members of the metazoan phylogenetic lineage and their genome contains animal specific genes but lack true tissues, organ systems, and neurons. Thus, the sponge model system can be used to elucidate origin of developmental processes. The PSED (RDGN) network (Pax/Six/Eya/Dac) is important in development of eyes, muscles, and other structures in Bilaterians. Similarly, sponges contain a precursor Pax-Six gene network. The Ephydatia muelleri (Em) PaxB protein binds to a

Pax2/5/8 consensus sequence site and two cis-regulatory elements upstream and one intron sequence of EmSix1/2 (Rivera et al., 2013).

This study aimed to determine …