The loss of organisms to cooling water intakes has been identified as having an impact on aquatic biota. Impingement and entrainment of organisms are unavoidable if natural waters are to be used by utilities for cooling water purposes. This study was initiated to determine the effects of operation of the intake of the Glen Lyn Power Station on the fish community of the New River, Glen Lyn, Virginia. Estimates of the numbers of fishes impinged by the Glen Lyn Power Station were made for the period mid-May 1976 through May 1977. The estimated total impingement was 6219 fish of 22 species. The alewife (Alosa pseudoharengus) accounted for 86.6% of the fish; most of these were impinged during the winter probably as a result of cold induced mortality in an upstream reservoir population. The estimated numbers of other species impinged ranged from 6 to 182. Most of the fish were dead prior to impingement. An estimated 30,200,000 larval fish of an estimated 172,000,000 that drifted by the Glen Lyn Power Station during June 1976 through May 1977 were entrained by the power station. Larvae of the carp (Cyprinus carpio) accounted for 94.8% of the ichthyoplankton drift. Estimates of 14.5 to 21.7% of the larvae of selected representative species that drifted by the power station were entrained. The criteria listed by the United States Environmental Protection Agency for assessing the impact of intake operation on aquatic communities were used in conjunction with collected data and information on the fish community of the New River available in the literature. It was concluded that the losses to the intake of the Glen Lyn Power Station were not significant enough to affect the fish community structure of the New River near Glen Lyn, Virginia in its present condition. / Master of Science

P68 is a prototypical member of the DEAD box RNA helicase family. Implicated in numerous functions such as cell proliferation, cancer metastasis, transcription regulation and pre-mRNA spllicing, p68 is a multifunctional protein whose roles are still not completely understood. In the studies presented, we found that p68 was an important regulator of numerous cancer related genes. This study focuses on the cancer related genes Snail and hTERT. We show that p68 binds to the promoter and a downstream region within each gene, suggesting that p68 operates via the same mechanism with both genes. We also show that tyrosine phosphorylated p68 is the major player in transcription regulation. p68 was also discovered to recruit CREB-binding protein to the promoters of these genes as well as aid in the removal of HDAC1 from the promoters; these findings are consistent with chromatin remodeling and active transcription. Also, we found that p68 phosphorylation level correlates with the expression level of these genes. Finally, we describe other genes that are potentially regulated by p68 in the same manner, through the use of ChiP-on-chip technology.

The purpose of this investigation is the application of the penetration model for gas absorption with chemical reaction in a stirred reactor and the evaluation of the parameters of the penetration model, viz. the gas-liquid interfacial area and gas-liquid particle contact time, for the different agitation intensities and gas rates. The values of these parameters for the model were determined by measuring the rates of absorption of carbon dioxide into caustic solutions and then forcing the model to give values of the parameter consistent with observed rates of absorption. The contact times and interfacial areas were determined for five agitation rates ranging from 150 revolutions of the agitator to 350 and for five gas input rates ranging from superficial gas velocities of 0.29 centimeter per second to about 1.2 centimeters per second. These parameters were evaluated for three different caustic strengths. The interfacial areas show discrepancies in values for the three different caustic strengths. These results, together with the work of an earlier investigator, seem to suggest that, for design and scale-up purposes for gas absorption in solutions, experiments be set up with the solutions of actual interest. The interfacial areas obtained correlate well] with the correlations given by Calderbank, which is based on the work of Hinze on bubble sizes in turbulent regimes. The gas-liquid particle contact times are again found to be dependent on the intensity of agitation and the gas rates. A correlation based on the theory of isotropic turbulence has been proposed and found to correlate the data well. / Ph. D.

Adults of Periplaneta americana, P. australasiae, P. brunnea, P. fuliginosa, and P. japonica are well known and have been well described. However, little systematic work has been conducted on the immature stages. There are very few descriptions of the nymphs of any Periplaneta species, and those that are available are rather brief and not completely accurate. This paper presents detailed descriptions and keys to the first-stage nymphs of the above species. Five types of setae, including unique mechanoreceptive setae, were found on the thoracic segments. P. australasiae, P. fuliginosa, and P. brunnea have very setose thoracic nota·and a banded color pattern, while thoracic nota of P. americana and P. japonica are less setose and solid in color. Great variability of setal number and color pattern probably exists among all the species in the genus. / Master of Science

The initiation of cancer metastasis usually requires Epithelial-Mesenchymal Transition (EMT), by which tumor cells lose cell-cell interactions and gain the ability of migration and invasion. Previous study demonstrated that p68 RNA helicase, a prototypical member of the DEAD-box RNA helicases, functions as a mediator to promote platelet-derived growth factor (PDGF)-induced EMT through facilitating nuclear translocation of β-catenin in colon cancer cells. In this context, p68 RNA helicase was found to be phosphorylated at the tyrosine 593 residue (referred as phosphor-p68) by c-Abl kinase, and this phosphorylation is required for the activation of β-catenin signaling and the consequent EMT. The phosphor-p68 RNA helicase-mediated EMT was characterized by the repression of an epithelial marker, E-cadherin, and the upregulation of a mesenchymal marker, Vimentin. E-cadherin, a major cell-cell adhesion molecule that is involved in the formation of adherens junctions, has been shown to sequester β-catenin at the cell membrane and thus inhibit its transcriptional activity. The functional loss of E-cadherin is the fundamental event of EMT. Despite the role of phosphor-p68 RNA helicase in regulating nuclear translocation of β-catenin, whether phosphor-p68 is involved in the regulation of E-cadherin remains unknown. Here, our data indicated that phosphor-p68 RNA helicase initiated EMT by transcriptional upregulation of Snail1, a master transcriptional repressor of E-cadherin. The data suggest that phosphor-p68 RNA helicase displaced HDAC1 from the chromatin remodeling MBD3:Mi-2/NuRD complex at the Snail1 promoter, thereby activating the transcription of Snail1. In the xenograft tumor model, abolishing the phosphorylation of p68 RNA helicase by the expression of Y593F mutant resulted in a significant reduction of metastatic potential in human colon cancer cells. Analyses in the colon cancer tissues also revealed that the tyrosine 593 phosphorylation level of p68 RNA helicase is substantially enhanced in the tumor tissues comparing to that in the corresponding normal counterparts, suggesting a correlation of phosphor-p68 and tumor progression. In conclusion, we showed that tyrosine phosphorylation of p68 RNA helicase positively correlated to the malignant status of colon cancer progression. The molecular basis behind this correlation could be partly through the transcriptional regulation of Snail1.

How tumor cells migrate and metastasize from primary sites requires four major steps: invasion, intravasation, extravasation and proliferation from micrometastases to malignant tumor. The initiation of tumor cell invasion requires Epithelial-Mesenchymal Transition (EMT), by which tumor cells lose cell-cell interactions and gain the ability of migration. The gene expression profile during the EMT process has been extensively investigated to study the initiation of EMT. In our studies, we indicated that tyrosine phosphorylation of human p68 RNA helicase positively associated with the malignant status of tumor tissue or cells. Studying of this relationship revealed that p68 RNA helicase played a critical role in EMT progression by repression of E-cadherin as an epithelial marker and upregulation of Vimentin as a mesenchymal marker. Insight into the mechanism of how p68 RNA helicase represses E-cadherin expression indicated that p68 RNA helicase initiated EMT by transcriptional upregulation of Snail. Human p68 RNA helicase has been documented as an RNA-dependent ATPase. The protein is an essential factor in the pre-mRNA splicing procedure. Some examples show that p68 RNA helicase functions as a transcriptional coactivator in ATPase dependent or independent manner. Here we indicated that p68 RNA helicase unwound protein complexes to modulate protein-protein interactions by using protein-dependent ATPase activity. The phosphorylated p68 RNA helicase displaced HDAC1 from the chromatin remodeling MBD3:Mi2/NuRD complex at the Snail promoter. Thus, our data demonstrated an example of protein-dependent ATPase which modulates protein-protein interactions within the chromatin remodeling machine.

Cancer cells take more glucose to provide energy and phosphoryl intermediates for cancer progression. Meanwhile, energy-provider function of mitochondria in cancer cells is disrupted. This phenomenon is so-called Warburg effect, which is discovered over eighty years ago. The detail mechanisms for Warburg effect are not well defined. How glycolytic enzymes contribute to cancer progression is not well known. PKM2 is a glycolytic enzyme dominantly localized in the cytosol, catalyzing the production of ATP from PEP. In this study, we discovered that there were more nuclear PKM2 expressed in highly proliferative cancer cells. The nuclear PKM2 levels are correlated with cell proliferation rates. According to our microarry analyses, MEK5 gene was upregulated in PKM2 overexpression cells. Our studies showed that PKM2 regulated MEK5 gene transcription to promote cell proliferation. Moreover, nuclear PKM2 phosphorylated Stat3 at Y705 site using PEP as a phosphoryl group donor to regulate MEK5 gene transcription. Our study also showed that double phosphorylated p68 RNA helicase at Y593/595 interacted with PKM2 at its FBP binding site. Under the stimulation of growth factors, p68 interacted with PKM2 to promote the conversion from tetrameraic to dimeric form so as to regulate its protein kinase activity. Overexpression PKM2 in less aggressive cancer cells induced the formation of multinuclei by regulating Cdc14A gene transcription. Overall, this study presents a step forward in understanding the Warburg effect.

P68 RNA helicase (p68), as a DEAD family protein, is a typical RNA helicase protein. P68 functions in many other biological processes, which include the regulations of the gene transcription, cell proliferation and cell differentiation. In our group, Y593 phosphorylated p68 was found to have a function in the epithelial mesynchymal transition, which is an important process for cancer metastasis. In the present study, we found that p68 is a nucleocytoplasmic shuttling protein. The protein carries two functional nuclear exporting signal sequences and two nuclear localization signal sequences. Calmodulin, a calcium sensor protein, is well known to play roles in cell migration by regulating the activities of its target proteins at the leading edge. Calmodulin interacts with p68 at the IQ motif of p68. However, the biological function of this interaction is not known. In this study, we found that the p68/calmodulin protein complex functions as a microtubule motor in migrating cells. The shuttling function of p68 along with the motor function of p68/calmodulin causes the leading edge distribution of calmodulin in migrating cells. Disruption the interaction between p68 and calmodulin inhibits cancer cell metastasis in an established mouse model. On the other hand, Y593-Y595 double phosphorylated p68 were found to interact with PKM2, an important tumor isoform of pyruvate kinase. The shuttling function of p68 is reasoned to promote the dimer formation of PKM2 and transport the PKM2 to the cell nucleus. The nuclear PKM2 was found to function as a protein kinase to promote cell proliferation. In specific, the nuclear PKM2 phosphorylates and activates Stat3, an important transcription factor functions in cell proliferation. Overall, p68 is found to have functions in both cell migration and cell proliferation, and these two functions depend on the nucleocytoplasmic shuttling activity and the post-translational modification of p68.

<div>G-quadruplexes (G4s) are non-canonical secondary structures formed in single-stranded guanine-rich nucleic acid sequences, such as those found in oncogene promoters and telomeres. <i>MYC</i>, one of the most critical oncogenes, has a DNA G4 (MycG4) in its proximal promoter region that functions as a transcriptional silencer. MycG4 is very stable and the pathological activation of <i>MYC</i> requires its active unfolding. However, it remains unclear what drives MycG4 unfolding in cancer cells. We have studied the interactions of DDX5 with the MycG4 at both molecular and cellular levels and discovered that DDX5 actively unfolds the MycG4 and is involved in the <i>MYC</i> gene transcriptional regulation, which is described in the first part of this dissertation. DDX5 is extremely proficient at unfolding the MycG4 and ATP hydrolysis is not directly coupled to the G4-unfolding of DDX5. In cancer cells, DDX5 is enriched at the <i>MYC</i> promoter and activates <i>MYC</i> transcription. G4-interactive small molecules inhibit the DDX5 interaction with the <i>MYC</i> promoter and DDX5-mediated <i>MYC</i> activation. The second part of this dissertation describes the study of interactions of indenoisoquinoline anticancer drugs with MycG4. The MycG4 transcriptional silencer is a very attractive therapeutic target. Compounds that bind and stabilize the MycG4 have been shown to repress <i>MYC</i> gene transcription and are antitumorigenic. Indenoisoquinolines are human topoisomerase I inhibitors in clinical testing. However, some indenoisoquinolines with potent anticancer activity do not exhibit strong topoisomerase I inhibition, suggesting a separate mechanism of action. Our studies show that indenoisoquinolines strongly bind and stabilize MycG4 and lower <i>MYC</i> levels in cancer cells. Moreover, the analysis of indenoisoquinoline analogues for their <i>MYC</i> inhibitory activity, topoisomerase I inhibitory activity, and anticancer activity reveals a synergistic effect of <i>MYC</i> inhibition and topoisomerase I inhibition on anticancer activity. Besides the MycG4, human telomeric G4s are also attractive targets for anticancer drugs due to their ability to inhibit telomere extension in cancer cells. The last part of this dissertation reviews two recent solution structural studies on small molecule complexes with the hybrid-2 telomeric G4 and the hybrid-1 telomeric G4. Structural information of those complexes can advance the design of telomeric G4-interactive small molecules in the cancer therapeutic areas.</div>

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