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Full-Text Articles in Molecular Biology

Studies Of Sumoylation In Regulating Mif Stability And Rangap1 Nucleo-Cytoplasmic Shuttling In Controlling Its Sumo Modification, Progga Sen Jan 2017

Studies Of Sumoylation In Regulating Mif Stability And Rangap1 Nucleo-Cytoplasmic Shuttling In Controlling Its Sumo Modification, Progga Sen

Wayne State University Dissertations

SUMOylation is an essential post-translational modification that regulates a variety of critical cellular pathways ranging from nuclear transport to protein stability. Accumulating lines of evidence have shown that a perturbation of the SUMOylation pathway is associated with human diseases, especially various types of cancer. Our recent proteomic studies revealed a drastic increase in levels of SUMO2/3 modification on the proinflammatory cytokine MIF in the metastatic breast cancer cell line compared to the non-metastatic control cell line. Interestingly, the increase in levels of both MIF and global SUMO-2/3 modification in the metastatic cells are positively correlated to that of ...


Characterization Of The Yeast Cysteine Desulfurase Complex Within The Mitochondrial Fe-S Cluster Biogenesis, Dulmini Pabasara Barupala Jan 2016

Characterization Of The Yeast Cysteine Desulfurase Complex Within The Mitochondrial Fe-S Cluster Biogenesis, Dulmini Pabasara Barupala

Wayne State University Dissertations

Disrupted iron homeostasis within the human body materializes as various disorders. Pathophysiology of many of them relates to iron induced oxidative damage to key cellular components caused by iron accumulation within the tissues. Pertaining to the growing occurrence, cost of patient care and devastating burden associated with these diseases, the call for understanding the role of iron homeostasis within these disorders becomes inevitable. Being an abundant iron containing cofactor, the role of Fe-S clusters in cellular iron homeostasis is indisputable in the case of Friedreich’s ataxia, a disease caused by a deficiency in the protein frataxin that is indispensable ...


Insights Into De Novo Fes-Cluster Biogenesis Via The Eukaryotic Fes-Cluster (Isc) Pathway In Vitro, Stephen Paul Dzul Jan 2016

Insights Into De Novo Fes-Cluster Biogenesis Via The Eukaryotic Fes-Cluster (Isc) Pathway In Vitro, Stephen Paul Dzul

Wayne State University Dissertations

Fe-S clusters are iron-containing cofactors utilized by numerous proteins within several biological pathways essential to life. In eukaryotes, the primary pathway for Fe-S cluster production is the iron-sulfur cluster (ISC) pathway. The eukaryotic ISC pathway, localized primarily within the mitochondria, has been best characterized within Saccharomyces cerevisiae. In yeast, de novo Fe-S cluster formation is accomplished through coordinated assembly of the substrates iron and sulfur on the primary scaffold assembly protein “Isu1”. The sulfur used for cluster assembly is provided by the cysteine desulfurase “Nfs1”, a protein that works in union with its accessory protein “Isd11”. Frataxin “Yfh1” helps direct ...


Cardiolipin Is Required For Optimal Acetyl-Coa Metabolism, Vaishnavi Raja Jan 2016

Cardiolipin Is Required For Optimal Acetyl-Coa Metabolism, Vaishnavi Raja

Wayne State University Dissertations

The phospholipid cardiolipin (CL) is crucial for many cellular functions and signaling pathways, both inside and outside of mitochondria. My thesis focuses on the role of CL in energy metabolism. Many reactions of electron transport and oxidative phosphorylation, the transport of metabolites needed for these processes, and the stabilization of electron transport chain supercomplexes, require CL. Recent studies indicate that CL is required for the synthesis of iron-sulfur (Fe-S) co-factors, which are essential for numerous metabolic pathways. Activation of carnitine-acetylcarnitine translocase, which transports acetyl-CoA into the mitochondria, is CL dependent. The presence of substantial amounts of CL in the peroxisomal ...


Investigation Of Mutations In Nuclear Genes That Affect The Atp Synthase, Russell Dsouza Jan 2016

Investigation Of Mutations In Nuclear Genes That Affect The Atp Synthase, Russell Dsouza

Wayne State University Dissertations

The F1 domain is the catalytic subunit of the mitochondrial ATP synthase. Studies with respiratory-deficient yeast identified ATP1 and ATP2 as nuclear genes encoding the alpha and beta subunits, respectively, of the mitochondrial F1-ATPase. The mutations in the atp1 and atp2 genes were cloned and sequenced, and they appear to affect the ATP synthase. Most yeast strains with mutations in the or the subunit primarily show an F1 assembly defective phenotype. This feature is similar to the assembly-defective mutants missing the chaperones required for assembly of the F1 oligomer or either the alpha/beta subunits.

Some of the atp2 and ...


Development Of A Novel Class Of Chemicals For Labeling Abasic Sites In Cellular Dna And Killing Cancer Cells, Shanqiao Wei Jan 2016

Development Of A Novel Class Of Chemicals For Labeling Abasic Sites In Cellular Dna And Killing Cancer Cells, Shanqiao Wei

Wayne State University Dissertations

Abasic (AP) sites are the most common type of lesions in DNA. Numerous endogenous and exogenous agents and cellular processes can induce the formation of AP sites in DNA. If left unrepaired, the deleterious AP sites cause mutagenesis and cytotoxicity. Methoxyamine is known to react with AP sites and block base excision repair. Another alkoxyamine, aldehyde-reactive probe (ARP) tags AP sites with a biotin and has been widely used to quantify these sites. In this study, I have combined both these abilities into one alkoxyamine, AA3, which reacts toward AP sites with better reactivity than ARP at physiological pH. Additionally ...


Novel Regulatory Mechanisms Of Inositol Biosynthesis In Saccharomyces Cerevisiae And Mammalian Cells, And Implications For The Mechanism Underlying Vpa-Induced Glucose 6-Phosphate Depletion, Wenxi Yu Jan 2016

Novel Regulatory Mechanisms Of Inositol Biosynthesis In Saccharomyces Cerevisiae And Mammalian Cells, And Implications For The Mechanism Underlying Vpa-Induced Glucose 6-Phosphate Depletion, Wenxi Yu

Wayne State University Dissertations

Myo-inositol is the precursor of all inositol containing molecules, including inositol phosphates, phosphoinositides and glycosylphosphatidylinositols, which are signaling molecules involved in many critical cellular functions. Perturbation of inositol metabolism has been linked to neurological disorders. Although several widely-used anticonvulsants and mood-stabilizing drugs have been shown to exert inositol depletion effects, the mechanisms of action of the drugs and the role of inositol in these diseases are not understood. Elucidation of the molecular control of inositol synthesis will shed light on the pathologies of inositol related illnesses.

In Saccharomyces cerevisiae, deletion of the four glycogen synthase kinase-3 genes, MCK1, MRK1, MDS1 ...


A Critical Role Of Cxcr2 Pdz Motif-Mediated Interactions In Endothelial Progenitor Cell Homing And Angiogenesis, Yuning Hou Jan 2016

A Critical Role Of Cxcr2 Pdz Motif-Mediated Interactions In Endothelial Progenitor Cell Homing And Angiogenesis, Yuning Hou

Wayne State University Dissertations

Bone marrow-derived endothelial progenitor cells (EPCs) participate in postnatal vascularization in response to growth factors, cytokines, and chemokines. Chemokine receptor CXCR2 and its cognate ligands are reported to mediate EPC recruitment and angiogenesis. CXCR2 possesses a consensus PSD-95/DlgA/ZO-1 (PDZ) motif at its carboxyl terminus. The PDZ motif has been reported to regulate cellular signaling and functions. Here we investigated the potential role of the PDZ motif in CXCR2-mediated EPC motility and angiogenesis. We have found that introducing exogenous CXCR2 C-terminus significantly attenuated in vitro EPC migration and angiogenic activities in response to CXCR2 ligands, as well as in ...


Analyzing The Interactions Of Kdm5/Lid And Sin3 In Drosophila Melanogaster, Ambikai Gajan Jan 2015

Analyzing The Interactions Of Kdm5/Lid And Sin3 In Drosophila Melanogaster, Ambikai Gajan

Wayne State University Dissertations

SIN3, the scaffold protein of a histone modifying complex is conserved from yeast to mammals. Drosophila SIN3 associates with both a histone deactylase RPD3 and a histone demethylase dKDM5/LID. Immunopurification of dKDM5/LID verifies a previously observed interaction with SIN3 and RPD3. Furthermore, deficiency of dKDM5/LID phenocopies deficiency of SIN3 in many cellular and developmental processes. Knockdown of both Sin3A and lid hinder cell proliferation in Drosophila cultured cells and developing flies. Knockdown of these genes also results in a curved wing phenotype implicating a role in wing development. Analysis of underlying gene expression changes upon decreased expression ...


Biochemical, Structural, And Drug Design Studies Of Multi-Drug Resistant Hiv-1 Therapeutic Targets, Tamaria Grace Dewdney Jan 2013

Biochemical, Structural, And Drug Design Studies Of Multi-Drug Resistant Hiv-1 Therapeutic Targets, Tamaria Grace Dewdney

Wayne State University Dissertations

Protein point mutations acquired as a mechanism of survival against therapeutics cause structural changes that effect protein function and inhibitor binding. This work investigates the structural mechanisms that lead to multi-drug resistance to HIV-1 protease and integrase inhibitors.

Proper proteolytic processing of the HIV-1 Gag/Pol polyprotein is required for HIV infection and viral replication. This feature has made HIV-1 protease an attractive target for antiretroviral drug design for the treatment of HIV-1 infected patients, thus the development of drug resistance has arisen as a major therapeutic and drug design challenge. To understand the molecular mechanisms leading to drug resistance ...


The Role Of Cardiolipin In Iron Homeostasis And Glutathione Metabolism, Vinay A. Patil Jan 2013

The Role Of Cardiolipin In Iron Homeostasis And Glutathione Metabolism, Vinay A. Patil

Wayne State University Dissertations

Cardiolipin (CL) is the signature phospholipid of mitochondrial membranes, where it is synthesized locally and plays a critical role in mitochondrial bioenergetic functions. Inside the mitochondria, CL is a critical target of mitochondrial generated reactive oxygen species (ROS) and regulates signaling events related to apoptosis and aging. CL deficiency causes perturbation of signaling pathways outside the mitochondria, including the PKC-Slt2 cell integrity pathway and the high osmolarity glycerol (HOG) pathway, and is a key player in the cross-talk between the mitochondria and the vacuole. The importance of CL in human health is underscored by the observation that perturbation of CL ...


Genetic And Biochemical Studies Of Human Apobec Family Of Proteins, Priyanga Wijesinghe Jan 2012

Genetic And Biochemical Studies Of Human Apobec Family Of Proteins, Priyanga Wijesinghe

Wayne State University Dissertations

The AID/APOBEC family of proteins in higher vertebrates converts cytosines in DNA or RNA into uracil. These proteins have essential roles in either innate immunity or adaptive immunity. Recently, AID has also been implicated in DNA demethylation in the context of early embryogenesis in mammals. This is partly based on the reported ability of AID to deaminate 5-methyl cytosine to thymine (5mC to T). I reexamined this proposed new role of AID (5mC deamination) with two members of the APOBEC family in a novel Escherichia coli based genetic system. My results confirmed that while all three enzymes are strong ...


Prevalence And Physiological Significance Of Gene Looping In Saccharomyces Cerevisiae, Banupriya Mukundan Jan 2012

Prevalence And Physiological Significance Of Gene Looping In Saccharomyces Cerevisiae, Banupriya Mukundan

Wayne State University Dissertations

My Ph.D. dissertation work is focused on studying the role of promoter-bound transcription initiation factors involved in gene looping. In this study we showed that the RNAP II subunit Rpb4 has a significant effect on termination of transcription. Gene looping is disrupted in the absence of Rpb4. Rpb4 shows a strong physical interaction with the Mediator subunit Srb5. Mediator subunit Srb5 crosslinked to the 5' and 3' ends of INO1 and CHA1 genes and is required for proper termination of transcription of these genes. Srb5 affected termination of transcription through its interaction with the CF1 complex. Srb5 interaction with ...


Cardiac Calsequestrin Phosphorylation And Trafficking In The Mammalian Cardiomyocyte, Timothy Mcfarland Jan 2011

Cardiac Calsequestrin Phosphorylation And Trafficking In The Mammalian Cardiomyocyte, Timothy Mcfarland

Wayne State University Dissertations

Cardiac CSQ (CSQ2) is a multifaceted protein, capable of binding significant quantities of Ca2+ and altering ryanodine receptor activity at the junctional sarcoplasmic reticulum (SR). Little is known about the trafficking of CSQ2 from its unknown site of biosynthesis, which appears to be of importance as its structure changes in a trafficking-dependent manner in various types of heart failure. Through the use of multiple antibodies specific to classic rough ER markers, and with the creation of CSQ-DsRed tetramer fusion protein, we were able to establish a juxtanuclear localization of rough ER in cardiomyocytes. Using fluorescence confocal microscopy, the translocon complex ...


Molecular Details Of The Mitochondrial Iron Sulfur Cluster Assembly Pathway, Swati Rawat Jan 2011

Molecular Details Of The Mitochondrial Iron Sulfur Cluster Assembly Pathway, Swati Rawat

Wayne State University Dissertations

MOLECULAR DETAILS OF THE MITOCHONDRIAL IRON SULFUR CLUSTER ASSEMBLY PATHWAY

Iron-sulfur clusters are an important class of prosthetic group involved in electron transfer, enzyme catalysis, and regulation of gene expression. Their biosynthesis requires complex machinery located within the mitochondrion since free iron and sulfide are extremely toxic to the cell. Defects in this pathway results in several diseases such as Friedreich's Ataxia (FRDA), Sideroblastic Anemia and ISCU Myopathy. Therefore molecular details of the biogenesis pathway will provide deep insight in the pathway and treatment options for these diseases. FRDA is caused by deficiency of a single protein called as ...


Characterization Of Splicing Mechanisms By Single-Molecule Fluorescence, Krishanthi Sanjeewani Karunatilaka Jan 2011

Characterization Of Splicing Mechanisms By Single-Molecule Fluorescence, Krishanthi Sanjeewani Karunatilaka

Wayne State University Dissertations

Group II introns rank amongst the largest self-splicing ribozymes found in bacteria and organellar genomes of various eukaryotes. Despite the diversity in primary sequences, group II introns posses highly conserved secondary structures consisting of six domains (D1-D6). To perform its function, the large multidomain group II intron RNA must adopt the correctly folded structure. As a result, in vitro splicing of these introns requires high ionic strength and elevated temperatures. In vivo, this process is mainly assisted by protein cofactors. However, the exact mechanism of protein-mediated splicing of group II intron RNA is still not known.

In order to elucidate ...


Characterization Of Arsd: An Arsenic Chaperone For The Arsab As(Iii)-Translocating Atpase, Jianbo Yang Jan 2010

Characterization Of Arsd: An Arsenic Chaperone For The Arsab As(Iii)-Translocating Atpase, Jianbo Yang

Wayne State University Dissertations

Arsenic is a metalloid toxicant that is widely distributed throughout the earth's crust and causes a variety of health and environment problems. As an adaptation to arsenic-contaminated environments, organisms have developed resistance systems. In bacteria and archaea various ars operons encode ArsAB ATPases that pump the trivalent metalloids As(III) or Sb(III) out of cells. In these operons, an arsD gene is almost always adjacent to the arsA gene, suggesting a related function. ArsA is the catalytic subunit of the pump that hydrolyzes ATP in the presence of arsenite or antimonite. ArsB is a membrane protein which containing ...