In this study, the active learning technique think-pair-share was tested in an independent high school non-honors chemistry classroom to see if it was more effective than teaching techniques already being used in the chemistry classes. Two classes of tenth grade chemistry students were combined as a test group and a third class of tenth grade chemistry students acted as the control. Learning gains for pre and post-tests from three different chemistry units were analyzed and no significant difference was found between the results of the control and test groups, indicating that think-pair-share was as effective as teaching methods already in place. Power analysis indicated that results had a low chance of showing a significant difference between the learning gains for the two groups. An attitude survey given after the study was over indicated that students felt more comfortable in the classroom when cooperative learning techniques were employed.

Initiation of DNA Replication is a highly regulated process and is characterized by the conversion of inactive Mcm2-7 double hexamer around dsDNA to an active helicase consisting of Mcm2-7 single hexamer which encircles ssDNA along with Cdc45 and GINS. This transition of inactive Mcm2-7 around dsDNA to the active helicase around ssDNA involves several unknown mechanisms. Many proteins like, Sld2, Sld3, Dpb11, Mcm10, have been reported to be essential for replication initiation. Dpb11 is one of these essential proteins that functions in the initiation of DNA replication. Dpb11 binds to S-CDK phosphorylated Sld2 and Sld3 to form a ternary complex during S phase. These interactions are essential for the CDK-dependent activation of DNA replication in budding yeast. Studies have also indicated that chromosomal replication initiates by a fundamentally similar process in all eukaryotes, with Dpb11 being an evolutionary conserved protein. Our studies using purified proteins from budding yeast show that Dpb11 alone binds to Mcm2-7, and Dpb11 also competes with GINS for binding to Mcm2-7. Furthermore, Dpb11 binds directly to single-stranded DNA (ssDNA), and ssDNA inhibits Dpb11 interaction with Mcm2-7. We also found that Dpb11 can recruit Cdc45 to Mcm2-7. We identified a mutant of the BRCT4 motif of Dpb11 that remains bound to Mcm2-7 in the presence of ssDNA (dpb11-m1,m2,m3,m5), and this mutant exhibits a DNA replication defect when expressed in budding yeast cells. Expression of this mutant results in increased interaction between Dpb11 and Mcm2-7 during S phase, impaired GINS interaction with Mcm2-7 during S phase, and decreased RPA interaction with origin DNA during S phase. We propose a model wherein Dpb11 first recruits Cdc45 to Mcm2-7. Dpb11, while bound to Cdc45-Mcm2-7, can block the interaction between GINS and Mcm2-7. Upon the extrusion of ssDNA from the central channel of Mcm2-7, Dpb11 dissociates from Mcm2-7 and Dpb11 binds to ssDNA, thereby allowing GINS to bind to Cdc45-Mcm2-7. Finally, we propose that Dpb11 functions with Sld2 and Sld3 to help control the assembly of the replication fork helicase.

Graphic organizers have been used in the classroom since the late 1960s. Several studies have been published on the effects of graphic organizers on cognitive learning and yet the traditional method of teaching in the science classroom still remains strongly focused on traditional lecture and note taking. This study shows that relative to traditional methods graphic organizers maintain equivalent learning gains, but increase excitement and engagement among the students. The purpose of this study was to assess traditional lecture vs. traditional lecture with the addition of concept maps to determine if concept maps affect learning gains in the high school science classroom. The study used 86 traditional Biology students. The students were given both a pre-test and post-test on two separate units in the Biology curriculum. The first unit covered Charles Darwin and the theory of natural selection, while the second unit covered the muscular and skeletal systems. Although there was a slight increase in learning gains when concept maps were used, statistical analysis determined that this small increase was not significant for this sample size. There was a statistically significant difference in learning gains between the two units however. An attitudinal survey given at the end of the course showed that students preferred using the concept maps in addition to traditional lecture over traditional lecture alone.

The integration of reproduction and metabolism is necessary for the survival and continuation of a species. While the neural circuits controlling energy homeostasis have been well-characterized, the signals controlling the relay of nutritional information to the reproductive axis are not conclusively defined. The cichlid fish Astatotilapia burtoni is ideal for studying the neural regulation of feeding and reproduction because during their parental care phase, females undergo a two-week period of forced starvation while holding developing young in their buccal cavity. To test the hypothesis that candidate neuropeptides known to be involved in feeding and energy homeostasis in mammals show conserved distribution patterns, we performed immunohistochemistry or in situ hybridization to localize appetite-stimulating (neuropeptide Y, NPY; agouti-related protein, AgRP) and appetite-inhibiting peptides (cocaine and amphetamine-regulated transcript, CART; pro-opiomelanocortin, POMC) in the cichlid fish brain. NPY, AgRP, CART, and pomc somata were localized to the lateral tuberal nucleus (NLT), the putative homolog of the arcuate nucleus, as well as other brain regions, and fiber distributions were similar to other teleosts as well as to mammals. To test whether conserved neuropeptide-containing neurons varied with reproductive state, we also quantified neuron somata size in the NLT as a proxy for their involvement in regulating changes in energy status and reproductive condition. Our results show that gravid females had larger NPY and AgRP neurons in the NLT compared to brooding females, but brooding females had larger POMC neurons compared to gravid females. CART neuron size did not differ between the two reproductive states. Thus, larger appetite-stimulating neurons (NPY, AgRP) likely promote feeding while females are gravid, while larger POMC neurons may act as a satiety signal to inhibit food intake during mouthbrooding. Hypothalamic mRNA levels for npy, agrp, pomc-α, cart 2 and cart 4 were also measured, and while AgRP mRNA levels were higher in gravid compared to brooding females, the remaining gene products did not differ between reproductive states. Collectively, however, our data suggest a potential role for NPY, AgRP, POMC and CART in regulating food intake in A. burtoni females during varying reproductive states.

Iron-sulfur proteins comprise one of the most ubiquitous and conserved classes of proteins in biology. The diverse functions of iron-sulfur clusters in proteins range from electron transfer to redox signaling. The assembly of iron-sulfur clusters in cells requires a complex protein system and acquisition of sulfur and iron. While the sulfur source for iron-sulfur cluster assembly is well-established, the iron source remains elusive. The first part of this work provides new evidence for the hypothesis that the conserved iron-sulfur cluster assembly protein IscA may act as the iron donor for iron-sulfur cluster assembly. Iron-sulfur clusters in proteins are also vulnerable to reactive oxygen and nitrogen species. Indeed, iron-sulfur clusters in proteins can be readily destroyed by nitric oxide (NO) forming protein-bound dinitrosyl iron complexes (DNICs). The second part of this work demonstrates that iron-sulfur proteins are the major source of protein-bound DNICs found in NO-exposed cells. The results reveal new aspects of the molecular mechanism underlying NO cytotoxicity. As dysfunction of iron-sulfur clusters has been implicated in several human diseases, two human iron-sulfur proteins have been chosen for functional investigation of their iron-sulfur clusters. The first example is Rtel1, a DNA helicase that regulates telomere length. Rtel1 was predicted to contain an iron-sulfur cluster, but this was not demonstrated. The third part of this work shows that the N-terminal domain of Rtel1 indeed contains a redox active [4Fe-4S] cluster. The second example is the mitochondrial outer membrane protein mitoNEET, a recently identified target of the type II diabetes drug pioglitazone. The studies show that mitoNEET [2Fe-2S] clusters can be readily reduced by biological thiols and human glutathione reductase, and is reversibly oxidized by hydrogen peroxide, suggesting that mitoNEET may undergo redox transitions to regulate mitochondrial energy metabolism in response to oxidative stress. In summary, the research presented in this dissertation advances our understanding of how iron-sulfur clusters may be assembled and how NO modification of iron-sulfur proteins may contribute to NO cytotoxicity. The human iron-sulfur proteins Rtel1 and mitoNEET further illustrate how iron-sulfur clusters may modulate protein functions via redox transition of their iron-sulfur clusters in response to oxidative signals.

Bariatric surgery is currently the only treatment for obesity that is effective in both the short and long term. Despite this, little is known about the mechanisms through which these surgeries act. While no single critical system has yet been identified, a number of major hypotheses have been proposed. One hypothesis is that adaptive hyperplasia of the intestines following Roux-en-Y gastric bypass (RYGB) surgery is required in order to receive the beneficial effects of the surgery. In this thesis I characterized the adaptive response of the intestines following RYGB and showed that significant hyperplasia occurs in the intestinal mucosa. In addition, I characterized the proliferation of enteroendocrine cells. I found that there is an increase in the number, but not density, of glucagon-like-peptide 1, neurotensin, and serotonin releasing cells, as well as an increase in both number and density of cholecystokinin releasing cells, in the Roux and common limbs following RYGB. This could link adaptive hyperplasia to increased circulating levels of these hormones, which has been implicated in a reduction of food intake and an improvement in glucose homeostasis. A second hypothesis that has been proposed states that RYGB and vertical sleeve gastrectomy (VSG), two similar but ultimately different surgeries, work through a common mechanism of action. In contrast to our observations in rats following RYGB, we found that adaptive hyperplasia is absent following VSG. This leads to the conclusion that either RYGB and VSG share a common mechanism that does not involve adaptive hyperplasia of the intestines, or they do not share a common mechanism.

Evaluating the conservation status of threatened species requires a sequential process of 1) taxonomic classification, 2) distributional mapping, and 3) evaluation of species according to criteria established by the IUCN Red List. Knowledge gaps in the first two phases of this process have been termed the Linnaean and Wallacean Shortfalls, respectively. The Indonesian archipelago is one of the most biologically diverse areas of the planet, as well as one of the most threatened and poorly studied, and the Linnaean and Wallacean Shortfalls pose substantial challenges for conservation in the region. Here I address each of the three stages of categorizing threatened species through a series of case studies on birds in two regions of Indonesia: Java and the Northern Moluccas. The first chapter addresses an example of the Linnaean Shortfall; using behavioural and vocal data, I provide insight on the taxonomic status of woodcock (genus: Scolopax) on Java. The display and vocalizations of these birds confirm that they are a distinct biological species and should be evaluated separately from woodcock in New Guinea. The second chapter focuses on the Wallacean Shortfall, and clarifies avian distributions in three Javan montane areas and on the North Moluccan island of Obi. On Obi, in particular, previous knowledge of bird distribution is woefully incomplete; I report nine species of resident birds previously unknown on the island. In the third and final section, I incorporate criteria from the IUCN Red List to re-evaluate the conservation status of two species on Obi, Chattering Lory Lorius garrulus and Moluccan Woodcock Scolopax rochussenii. One case results in good news for conservation; the Moluccan Woodcock is more common than previously believed, tolerates human habitat modification, and should be down-listed from Endangered to Vulnerable. The other is not; Chattering Lory is heavily trapped for the parrot trade and without conservation action, may likely become locally extinct. As such, it is more endangered than currently appreciated and should be considered Endangered rather than Vulnerable.

ABSTRACT Synapse formation is driven by precisely orchestrated intercellular communication between presynaptic and postsynaptic cells. Signals traverse a complex extracellular environment, where glycans attached to glycoproteins and proteoglycans modulate trans-synaptic signaling driving synapse formation, function and plasticity. To interrogate glycan effects on synapse structure and function, I performed a Drosophila transgenic RNAi screen targeting the glycan genome, including N/O/GAG-glycan biosynthesis/modifying enzymes and glycan-binding lectins. From the screen hits, I characterized two functionally paired genes to show unique synaptic effects. The first pair comprises the heparan sulfate (HS) 6-O-sulfotransferase (hs6st) and sulfatase (sulf1), which bidirectionally control HS proteoglycan (HSPG) sulfation. RNAi knockdown of hs6st and sulf1 causes opposite effects on functional synapse development, with neurotransmission strength and postsynaptic glutamate receptor machinery decreased in hs6st but elevated in sulf1 null mutants. Consistently, hs6st and sulf1 nulls differentially misregulate WNT (Wingless) and BMP (Glass Bottom Boat) ligands, their HSPG co-receptors Dally-like Protein and Syndecan, and downstream signaling. Genetic correction of altered WNT/BMP signaling restores normal synaptic development in both mutant conditions, proving that the altered trans-synaptic signaling causes the functional differentiation defects. The second screen-derived functional pair is two protein α-N-acetylgalactosaminyltransferases (pgant3 and pgant35A) that regulate synaptic O-linked glycosylation (GalNAcα1-O-S/T). Loss of either pgant alone elevates presynaptic/postsynaptic molecular assembly and evoked neurotransmission strength, but synapses appear restored to normal in double mutants. Likewise, activity-dependent facilitation, augmentation and post-tetanic potentiation are all suppressively impaired in pgant mutants. I show that Position Specific 2 (αPS2) integrin receptor and transmembrane tenascin ligand are both suppressively downregulated in pgant mutant synapses. Channelrhodopsin-driven electrical activity rapidly (<1 min) drives integrin signaling in wildtype synapses, but is suppressively abolished in pgant mutants. Optogenetic stimulation alters presynaptic vesicle trafficking and postsynaptic pocket size during perturbed integrin signaling underlying synaptic plasticity defects in pgant mutants. Critically, acute blockade of integrin signaling acts synergistically with pgant mutants to eliminate all activity-dependent synaptic plasticity. Thus, I identify two O- glycosylation synaptomatrix mechanisms that regulate trans-synaptic signaling underlying synaptic transmission and activity-dependent plasticity.

Zebrafish spontaneously regenerate their retinas after a variety of retinal insults. Key to regeneration are resident support cells called Müller glia (MG), which respond to injury by reverting to a stem cell-like state and generating progenitor cells that can proliferate, migrate and differentiate into any of the lost retinal cell types. Although studies have identified and characterized a number of genes and signaling pathways that control distinct steps of the regeneration process, very little is known about how expression of these genes is regulated. Small non-coding RNAs called microRNAs (miRNAs) regulate regeneration of various tissues in lower vertebrates. However, the functions and overall requirement of miRNAs for zebrafish retina regeneration are poorly understood. Using in vivo loss-of-function studies, we demonstrate that miRNAs are required for normal initiation and progression of retina regeneration. Small RNA high throughput sequencing revealed that miRNAs expression is dynamic during regeneration, but returned to its basal expression levels once regeneration was completed. We characterize the functions of two miRNAs that were downregulated during regeneration, miR-203 and miR-216, and discover that they regulate progenitor cell proliferation and MG dedifferentiation, respectively, during adult zebrafish retina regeneration.

Oyster reefs in the Gulf of Mexico provide water quality enhancement, shoreline stabilization, carbon sequestration, and facilitate spat recruitment. They are also essential refuges for numerous resident fish and invertebrates, in turn supporting commercial fisheries. Oyster reefs are however in danger worldwide as oyster fisheries increase and pollution from oil spills, such as the Deepwater Horizon spill further degrade reefs. The development of artificial reefs has therefore become a necessity. This study assesses both the long-term and acute response of oyster reef commensal communities to hydrocarbon contamination, as well as comparing the efficacy of artificial reef substrates for restoring these faunal assemblages. Long-term effects were analyzed by quantifying commensal abundance, taxa richness, and diversity from cultch-filled bags deployed at two sites in Barataria Bay, Louisiana, that experienced oiling from Deepwater Horizon, and two control sites. Bags were deployed seasonally in both 2012 and 2013, and the results indicated that while commensal abundance was generally greater at oiled sites, the effects of hydrocarbon contamination several years post spill were neither large nor consistent. To observe the acute colonization response, oil-soaked and control bags were retrieved 1, 2, 4, and 8 weeks after deployment at Grand Isle, LA, an area in Barataria Bay where no oil contamination was documented, in both June and September 2013. Oil effects on commensal communities were inconsistent and minimal by week 8, perhaps due to biodegradation of the hydrocarbons. Commensal communities were also sampled from bags containing either disarticulated oyster shell, limestone rubble or a composite material known as OysterCrete. While OysterCrete had the greatest abundance of commensal organisms, the experiments indicated that seasonal variation was more influential for commensal community dynamics, as well as new spat recruitment and growth, than the presence of hydrocarbons or various substrates. In areas in close proximity to major oil operations, such as the northern Gulf of Mexico, any restoration efforts that provide a hard substrate will be beneficial for the recruitment of commensal organisms if natural oyster reefs are impacted by anthropogenic disturbances.