Medical Molecular Biology Commons^™

Genetic Analysis Of Hereditary Gingival Fibromatosis Associated Sos1 Missense Variants Of Uncertain Significance In Caenorhabditis Elegans, Himani Patel

Theses

Hereditary gingival fibromatosis (HGF) is a disease that can present as benign overgrowth of gingival tissue in the mouth. The overgrowth can enclose the entire mouth and teeth in severe cases or present itself in a concentrated area. Researchers have identified that mutations in the SOS1 gene can be responsible for HGF. This disease can impair basic functions related to the mouth. Eating, smiling, speaking can all be affected. Additionally, excess inflammation can cause periodontal disease because of the difficulty in maintaining proper oral health. Periodontal disease can lead to severe bone loss which can lead to complete loss of …

Genetic Separation Of Brca1 Functions Reveal Mutation-Dependent Polθ Vulnerabilities, John J. Krais, David J. Glass, Ilse Chudoba, Yifan Wang, Wanjuan Feng, Dennis Simpson, Pooja Patel, Zemin Liu, Ryan Neumann-Domer, Robert G. Betsch, Andrea J. Bernhardy, Alice M. Bradbury, Jason Conger, Wei-Ting Yueh, Joseph Nacson, Richard T. Pomerantz, Gaorav P. Gupta, Joseph R. Testa, Neil Johnson

Department of Biochemistry and Molecular Biology Faculty Papers

Homologous recombination (HR)-deficiency induces a dependency on DNA polymerase theta (Polθ/Polq)-mediated end joining, and Polθ inhibitors (Polθi) are in development for cancer therapy. BRCA1 and BRCA2 deficient cells are thought to be synthetic lethal with Polθ, but whether distinct HR gene mutations give rise to equivalent Polθ-dependence, and the events that drive lethality, are unclear. In this study, we utilized mouse models with separate Brca1 functional defects to mechanistically define Brca1-Polθ synthetic lethality. Surprisingly, homozygous Brca1 mutant, Polq−/− cells were viable, but grew slowly and had chromosomal instability. Brca1 mutant cells proficient in DNA end resection were …

Conservation And Divergence In The Heterochronic Pathway Of C. Elegans And C. Briggsae, Maria Ivanova, Eric G. Moss

Rowan-Virtua Research Day

The heterochronic pathway of Caenorhabditis elegans is exemplary as a mechanism of developmental timing: mutations in genes of this pathway alter the relative timing of diverse developmental events independent of spatial or cell type specific regulation. It is the most thoroughly characterized developmental timing pathway known. Most of the heterochronic genes are conserved across great evolutionary time, and a few homologs seem to have developmental timing roles in certain contexts. The degree to which other organisms have explicit developmental timing mechanisms, and what factors comprise those mechanisms, isn’t generally known.

Developmental pathways evolve even if the resulting morphology remains the …

Replication Protein A (Rpa) Targeting Of Uracil Dna Glycosylase (Ung2), Derek Chen, Brian P Weiser

Rowan-Virtua Research Day

Replication Protein A (RPA) is a single stranded DNA binding protein which stabilizes ssDNA for replication and repair. One function of RPA is to bind the DNA repair enzyme uracil DNA glycosylase (UNG2) and direct its activity towards ssDNA dsDNA junctions.

UNG2 removes uracil bases from DNA which can appear through dUMP misincorporation or through cytosine deamination. If uracil is present instead of a cytosine, then the original GC pair becomes a GU pair. The uracil will then base pair to adenine in the replicated daughter strand. This results in a GC → AT mutation that could contribute to cancer …

Gasdermin Pores Permeabilize Mitochondria To Augment Caspase-3 Activation During Apoptosis And Inflammasome Activation., Corey Rogers, Dan A. Erkes, Alexandria Nardone, Andrew E. Aplin, Teresa Fernandes-Alnemri, Emad S. Alnemri

Department of Biochemistry and Molecular Biology Faculty Papers

Gasdermin E (GSDME/DFNA5) cleavage by caspase-3 liberates the GSDME-N domain, which mediates pyroptosis by forming pores in the plasma membrane. Here we show that GSDME-N also permeabilizes the mitochondrial membrane, releasing cytochrome c and activating the apoptosome. Cytochrome c release and caspase-3 activation in response to intrinsic and extrinsic apoptotic stimuli are significantly reduced in GSDME-deficient cells comparing with wild type cells. GSDME deficiency also accelerates cell growth in culture and in a mouse model of melanoma. Phosphomimetic mutation of the highly conserved phosphorylatable Thr6 residue of GSDME, inhibits its pore-forming activity, thus uncovering a potential mechanism by which GSDME …

Beta-Catenin Cleavage Enhances Transcriptional Activation, Tatiana Goretsky, Emily M. Bradford, Qing Ye, Olivia F. Lamping, Tomas Vanagunas, Mary Pat Moyer, Patrick C. Keller, Preetika Sinh, Josep M. Llovet, Tianyan Gao, Qing-Bai She, Linheng Li, Terrence A. Barrett

Internal Medicine Faculty Publications

Nuclear activation of Wnt/β-catenin signaling is required for cell proliferation in inflammation and cancer. Studies from our group indicate that β-catenin activation in colitis and colorectal cancer (CRC) correlates with increased nuclear levels of β-catenin phosphorylated at serine 552 (pβ-Cat⁵⁵²). Biochemical analysis of nuclear extracts from cancer biopsies revealed the existence of low molecular weight (LMW) pβ-Cat⁵⁵², increased to the exclusion of full size (FS) forms of β-catenin. LMW β-catenin lacks both termini, leaving residues in the armadillo repeat intact. Further experiments showed that TCF4 predominantly binds LMW pβ-Cat⁵⁵² in the nucleus of inflamed and …

Mutational Spectrum In A Worldwide Study Of 29,700 Families With Brca1 Or Brca2 Mutations, T. R. Rebbeck, T. M. Friebel, E. Friedman, U. Hamann, D. Huo, A. Kwong, E. Olah, O. I. Olopade, A. Lee, K. L. Nathanson, +231 Additional Authors

Journal Articles

No abstract provided.

Dysregulated Gpcr Signaling And Therapeutic Options In Uveal Melanoma., Vivian Chua, Dominic Lapadula, Clinita Randolph, Jeffrey L. Benovic, Philip B. Wedegaertner, Andrew E. Aplin

Department of Biochemistry and Molecular Biology Faculty Papers

Uveal melanoma is the most common primary intraocular malignant tumor in adults and arises from the transformation of melanocytes in the uveal tract. Even after treatment of the primary tumor, up to 50% of patients succumb to metastatic disease. The liver is the predominant organ of metastasis. There is an important need to provide effective treatment options for advanced stage uveal melanoma. To provide the preclinical basis for new treatments, it is important to understand the molecular underpinnings of the disease. Recent genomic studies have shown that mutations within components of G protein-coupled receptor (GPCR) signaling are early events associated …

Inhibition Of Rac1 Gtpase Sensitizes Pancreatic Cancer Cells To Γ-Irradiation, Y Yan, Ashley L. Hein, Asserewou Etekpo, Katrina M. Burchett, Chi Lin, Charles A. Enke, Surinder K. Batra, Kenneth Cowan, Michel M. Ouellette

Journal Articles: Biochemistry & Molecular Biology

Radiation therapy is a staple treatment for pancreatic cancer. However, owing to the intrinsic radioresistance of pancreatic cancer cells, radiation therapy often fails to increase survival of pancreatic cancer patients. Radiation impedes cancer cells by inducing DNA damage, which can activate cell cycle checkpoints. Normal cells possess both a G1 and G2 checkpoint. However, cancer cells are often defective in G1 checkpoint due to mutations/alterations in key regulators of this checkpoint. Accordingly, our results show that normal pancreatic ductal cells respond to ionizing radiation (IR) with activation of both checkpoints whereas pancreatic cancer cells respond to IR with G2/M arrest …

Testosterone Treatment Fails To Accelerate Disease In A Transgenic Mouse Model Of Spinal And Bulbar Muscular Atrophy., Erica S Chevalier-Larsen, Diane E Merry

Department of Biochemistry and Molecular Biology Faculty Papers

Evidence from multiple animal models demonstrates that testosterone plays a crucial role in the progression of symptoms in spinal and bulbar muscular atrophy (SBMA), a condition that results in neurodegeneration and muscle atrophy in affected men. Mice bearing a transgene encoding a human androgen receptor (AR) that contains a stretch of 112 glutamines (expanded polyglutamine tract; AR112Q mice) reproduce several aspects of the human disease. We treated transgenic male AR112Q mice with testosterone for 6 months. Surprisingly, testosterone treatment of AR112Q males did not exacerbate the disease. Although transgenic AR112Q males exhibited functional deficits when compared with non-transgenics, long-term testosterone …

Mediator Influences Telomeric Silencing And Cellular Life Span, Xuefeng Zhu, Beidong Liu, Jonas O. P. Carlsten, Jenny Beve, Thomas Nyström, Lawrence C. Myers, Claes M. Gustafsson

Dartmouth Scholarship

The Mediator complex is required for the regulated transcription of nearly all RNA polymerase II-dependent genes. Here we demonstrate a new role for Mediator which appears to be separate from its function as a transcriptional coactivator. Mediator associates directly with heterochromatin at telomeres and influences the exact boundary between active and inactive chromatin. Loss of the Mediator Med5 subunit or mutations in Med7 cause a depletion of the complex from regions located near subtelomeric X elements, which leads to a change in the balance between the Sir2 and Sas2 proteins. These changes in turn result in increased levels of H4K16 …

An Essential Role Of Human Ada3 In P53 Acetylation., Alo Nag, Aleksandra Germaniuk-Kurowska, Manjari Dimri, Michael A. Sassack, Channabasavaiah B. Gurumurthy, Qingshen Gao, Goberdhan Dimri, Hamid Band, Vimla Band

Journal Articles: Biochemistry & Molecular Biology

The p53 tumor suppressor protein functions as a critical component of genotoxic stress response by regulating the expression of effector gene products that control the fate of a cell following DNA damage. Unstressed cells maintain p53 at low levels through regulated degradation, and p53 levels and activity are rapidly elevated upon genotoxic stress. Biochemical mechanisms that control the levels and activity of p53 are therefore of great interest. We and others have recently identified hAda3 (human homologue of yeast alteration/deficiency in activation 3) as a p53-interacting protein and enhancer of p53 activity. Here, we show that endogenous levels of p53 …

Multiple Domains In Siz Sumo Ligases Contribute To Substrate Selectivity., Alison Reindle, Irina Belichenko, Gwendolyn R Bylebyl, Xiaole L Chen, Nishant Gandhi, Erica S Johnson

Department of Biochemistry and Molecular Biology Faculty Papers

Saccharomyces cerevisiae contains two Siz/PIAS SUMO E3 ligases, Siz1 and Siz2/Nfi1, and one other known ligase, Mms21. Although ubiquitin ligases are highly substrate-specific, the degree to which SUMO ligases target distinct sets of substrates is unknown. Here we show that although Siz1 and Siz2 each have unique substrates in vivo, sumoylation of many substrates can be stimulated by either protein. Furthermore, in the absence of both Siz proteins, many of the same substrates are still sumoylated at low levels. Some of this residual sumoylation depends on MMS21. Siz1 targets its unique substrates through at least two distinct domains. Sumoylation of …

Mitochondrial Dna Mutations, Apoptosis, And The Misfolded Protein Response., Justin L. Mott, Dekui Zhang, Hans Peter Zassenhaus

Journal Articles: Biochemistry & Molecular Biology

Studies of transgenic mice with accelerated accumulation of mtDNA mutations specifically in the heart lead us to propose that apoptotic signaling and cell death is central to the pathogenesis of mtDNA mutations in aging. It is the cellular response to that apoptotic signaling and the organ?s compensatory response to a loss of cells that specify the phenotype of an accumulation of mtDNA mutations. In the heart, cardiomyocytes induce a vigorous anti-apoptotic, pro-survival response to counteract mitochondrial apoptotic signaling. The heart up-regulates contractility of remaining myocytes in order to maintain cardiac output. We hypothesize that mutant mitochondrial proteins originate apoptotic signaling …

An Intramolecular Association Between Two Domains Of The Protein Kinase Fused Is Necessary For Hedgehog Signaling, Manuel Ascano Jr., David J. Robbins

Dartmouth Scholarship

The protein kinase Fused (Fu) is an integral member of the Hedgehog (Hh) signaling pathway. Although genetic studies demonstrate that Fu is required for the regulation of the Hh pathway, the mechanistic role that it plays remains largely unknown. Given our difficulty in developing an in vitro kinase assay for Fu, we reasoned that the catalytic activity of Fu might be highly regulated. Several mechanisms are known to regulate protein kinases, including self-association in either an intra- or an intermolecular fashion. Here, we provide evidence that Hh regulates Fu through intramolecular association between its kinase domain (ΔFu) and its carboxyl-terminal …

Nucleotide Excision Repair- And Polymerase Eta-Mediated Error-Prone Removal Of Mitomycin C Interstrand Cross-Links, H. Zheng, X. Wang, A. J. Warren, R. J. Legerski, Rodney S. Nairn, Joshua W. Hamilton, Lei Li

Dartmouth Scholarship

Interstrand cross-links (ICLs) make up a unique class of DNA lesions in which both strands of the double helix are covalently joined, precluding strand opening during replication and transcription. The repair of DNA ICLs has become a focus of study since ICLs are recognized as the main cytotoxic lesion inflicted by an array of alkylating compounds used in cancer treatment. As is the case for double-strand breaks, a damage-free homologous copy is essential for the removal of ICLs in an error-free manner. However, recombination-independent mechanisms may exist to remove ICLs in an error-prone fashion. We have developed an in vivo …

Purification Of Core-Binding Factor, A Protein That Binds The Conserved Core Site In Murine Leukemia Virus Enhancers., Shuwen W. Wang, Nancy A. Speck

Dartmouth Scholarship

The Moloney murine leukemia virus causes thymic leukemias when injected into newborn mice. A major genetic determinant of the thymic disease specificity of the Moloney virus genetically maps to two protein binding sites in the Moloney virus enhancer, the leukemia virus factor b site and the adjacent core site. Point mutations introduced into either of these sites significantly shifts the disease specificity of the Moloney virus from thymic leukemia to erythroleukemia (N. A. Speck, B. Renjifo, E. Golemis, T. Frederickson, J. Hartley, and N. Hopkins, Genes Dev. 4:233-242, 1990). We have purified several polypeptides that bind to the core site …