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Articles 31 - 60 of 69
Full-Text Articles in Molecular Biology
Post-Transcriptional Methylation Of Mitochondrial-Trna Differentially Contributes To Mitochondrial Pathology, Sunita Maharjan, Howard Gamper, Yuka Yamaki, Thomas W. Christian, Robert Y. Henley, Nan-Sheng Li, Takeo Suzuki, Tsutomu Suzuki, Joseph A. Piccirilli, Meni Wanunu, Erin L. Seifert, Douglas C. Wallace, Ya-Ming Hou
Post-Transcriptional Methylation Of Mitochondrial-Trna Differentially Contributes To Mitochondrial Pathology, Sunita Maharjan, Howard Gamper, Yuka Yamaki, Thomas W. Christian, Robert Y. Henley, Nan-Sheng Li, Takeo Suzuki, Tsutomu Suzuki, Joseph A. Piccirilli, Meni Wanunu, Erin L. Seifert, Douglas C. Wallace, Ya-Ming Hou
Department of Biochemistry and Molecular Biology Faculty Papers
Human mitochondrial tRNAs (mt-tRNAs), critical for mitochondrial biogenesis, are frequently associated with pathogenic mutations. These mt-tRNAs have unusual sequence motifs and require post-transcriptional modifications to stabilize their fragile structures. However, whether a modification that stabilizes a wild-type (WT) mt-tRNA would also stabilize its pathogenic variants is unknown. Here we show that the N1-methylation of guanosine at position 9 (m1G9) of mt-Leu(UAA), while stabilizing the WT tRNA, has a destabilizing effect on variants associated with MELAS (mitochondrial myopathy, encephalopathy, lactic acidosis, and stroke-like episodes). This differential effect is further demonstrated, as removal of the m1G9 …
Cryo-Em Analysis Of Pseudomonas Phage Pa193 Structural Components, Stephano M. Iglesias, Chun-Feng Hou, Johnny Reid, Evan Schauer, Renae Geier, Angela Soriaga, Lucy Sim, Lucy Gao, Julian Whitelegge, Pierre Kyme, Deborah Birx, Sebastien Lemire, Gino Cingolani
Cryo-Em Analysis Of Pseudomonas Phage Pa193 Structural Components, Stephano M. Iglesias, Chun-Feng Hou, Johnny Reid, Evan Schauer, Renae Geier, Angela Soriaga, Lucy Sim, Lucy Gao, Julian Whitelegge, Pierre Kyme, Deborah Birx, Sebastien Lemire, Gino Cingolani
Department of Biochemistry and Molecular Biology Faculty Papers
The World Health Organization has designated Pseudomonas aeruginosa as a critical pathogen for the development of new antimicrobials. Bacterial viruses, or bacteriophages, have been used in various clinical settings, commonly called phage therapy, to address this growing public health crisis. Here, we describe a high-resolution structural atlas of a therapeutic, contractile-tailed Pseudomonas phage, Pa193. We used bioinformatics, proteomics, and cryogenic electron microscopy single particle analysis to identify, annotate, and build atomic models for 21 distinct structural polypeptide chains forming the icosahedral capsid, neck, contractile tail, and baseplate. We identified a putative scaffolding protein stabilizing the interior of the capsid 5-fold …
Nanopore Signal Deviations From Pseudouridine Modifications In Rna Are Sequence-Specific: Quantification Requires Dedicated Synthetic Controls, Amr Makhamreh, Sepideh Tavakoli, Ali Fallahi, Xinqi Kang, Howard Gamper, Mohammad Nabizadehmashhadtoroghi, Miten Jain, Ya-Ming Hou, Sara H Rouhanifard, Meni Wanunu
Nanopore Signal Deviations From Pseudouridine Modifications In Rna Are Sequence-Specific: Quantification Requires Dedicated Synthetic Controls, Amr Makhamreh, Sepideh Tavakoli, Ali Fallahi, Xinqi Kang, Howard Gamper, Mohammad Nabizadehmashhadtoroghi, Miten Jain, Ya-Ming Hou, Sara H Rouhanifard, Meni Wanunu
Department of Biochemistry and Molecular Biology Faculty Papers
Chemical modifications to mRNA respond dynamically to environmental cues and are important modulators of gene expression. Nanopore direct RNA sequencing has been applied for assessing the presence of pseudouridine (ψ) modifications through basecalling errors and signal analysis. These approaches strongly depend on the sequence context around the modification, and the occupancies derived from these measurements are not quantitative. In this work, we combine direct RNA sequencing of synthetic RNAs bearing site-specific modifications and supervised machine learning models (ModQuant) to achieve near-analytical, site-specific ψ quantification. Our models demonstrate that the ionic current signal features important for accurate ψ classification are sequence …
Inorganic Pyrophosphate Plasma Levels In Patients With Ggcx-Associated Pxe-Like Phenotypes, Qiaoli Li, Catherine Troutman, Mary Peckiconis, Tamara Wurst, Sharon Terry
Inorganic Pyrophosphate Plasma Levels In Patients With Ggcx-Associated Pxe-Like Phenotypes, Qiaoli Li, Catherine Troutman, Mary Peckiconis, Tamara Wurst, Sharon Terry
Department of Biochemistry and Molecular Biology Faculty Papers
ntroduction: Pseudoxanthoma elasticum (PXE) is an autosomal recessive ectopic calcification disorder clinically affecting the skin, eyes, and vascular system. Most cases of PXE are caused by inactivating pathogenic variants in the ABCC6 gene encoding a hepatic transmembrane efflux transporter, which facilitates the extracellular release of ATP, the precursor of inorganic pyrophosphate (PPi), a potent endogenous inhibitor of calcification. Pathogenic variants in GGCX, encoding γ-glutamyl carboxylase required for activation of vitamin K-dependent coagulation factors as well as matrix Gla protein (MGP) and Gla-rich protein (GRP), two inhibitors of ectopic calcification, have also been reported to cause cutaneous changes like those seen …
The C-Terminal 4cxxc-Type Zinc Finger Domain Of Cdca7 Recognizes Hemimethylated Dna And Modulates Activities Of Chromatin Remodeling Enzyme Hells, Akeo Shinkai, Hideharu Hashimoto, Chikako Shimura, Hiroaki Fujimoto, Kei Fukuda, Naoki Horikoshi, Masaki Okano, Hitoshi Niwa, Erik W Debler, Hitoshi Kurumizaka, Yoichi Shinkai
The C-Terminal 4cxxc-Type Zinc Finger Domain Of Cdca7 Recognizes Hemimethylated Dna And Modulates Activities Of Chromatin Remodeling Enzyme Hells, Akeo Shinkai, Hideharu Hashimoto, Chikako Shimura, Hiroaki Fujimoto, Kei Fukuda, Naoki Horikoshi, Masaki Okano, Hitoshi Niwa, Erik W Debler, Hitoshi Kurumizaka, Yoichi Shinkai
Department of Biochemistry and Molecular Biology Faculty Papers
The chromatin-remodeling enzyme helicase lymphoid-specific (HELLS) interacts with cell division cycle-associated 7 (CDCA7) on nucleosomes and is involved in the regulation of DNA methylation in higher organisms. Mutations in these genes cause immunodeficiency, centromeric instability, and facial anomalies (ICF) syndrome, which also results in DNA hypomethylation of satellite repeat regions. We investigated the functional domains of human CDCA7 in HELLS using several mutant CDCA7 proteins. The central region is critical for binding to HELLS, activation of ATPase, and nucleosome sliding activities of HELLS-CDCA7. The N-terminal region tends to inhibit ATPase activity. The C-terminal 4CXXC-type zinc finger domain contributes to CpG …
Non-Redundant Roles For The Human Mrna Decapping Cofactor Paralogs Dcp1a And Dcp1b, Ivana Vukovic, Samantha M. Barnada, Jonathan W. Ruffin, Jon Karlin, Ravi Kumar Lokareddy, Gino Cingolani, Steven B. Mcmahon
Non-Redundant Roles For The Human Mrna Decapping Cofactor Paralogs Dcp1a And Dcp1b, Ivana Vukovic, Samantha M. Barnada, Jonathan W. Ruffin, Jon Karlin, Ravi Kumar Lokareddy, Gino Cingolani, Steven B. Mcmahon
Department of Medicine Faculty Papers
Eukaryotic gene expression is regulated at the transcriptional and post-transcriptional levels, with disruption of regulation contributing significantly to human diseases. The 5' m7G mRNA cap is a central node in post-transcriptional regulation, participating in both mRNA stabilization and translation efficiency. In mammals, DCP1a and DCP1b are paralogous cofactor proteins of the mRNA cap hydrolase DCP2. As lower eukaryotes have a single DCP1 cofactor, the functional advantages gained by this evolutionary divergence remain unclear. We report the first functional dissection of DCP1a and DCP1b, demonstrating that they are non-redundant cofactors of DCP2 with unique roles in decapping complex integrity and specificity. …
Engineered Nls-Chimera Downregulates Expression Of Aggregation-Prone Endogenous Fus, Miyuki Hayashi, Amandeep Girdhar, Ying-Hui Ko, Kevin Kim, Jacquelyn Depierro, Joseph R. Buchler, Nikhita Arunprakash, Aditya Bajaj, Gino Cingolani, Lin Guo
Engineered Nls-Chimera Downregulates Expression Of Aggregation-Prone Endogenous Fus, Miyuki Hayashi, Amandeep Girdhar, Ying-Hui Ko, Kevin Kim, Jacquelyn Depierro, Joseph R. Buchler, Nikhita Arunprakash, Aditya Bajaj, Gino Cingolani, Lin Guo
Department of Biochemistry and Molecular Biology Faculty Papers
Importin β-superfamily nuclear import receptors (NIRs) mitigate mislocalization and aggregation of RNA-binding proteins (RBPs), like FUS and TDP-43, which are implicated in neurodegenerative diseases. NIRs potently disaggregate RBPs by recognizing their nuclear localization signal (NLS). However, disease-causing mutations in NLS compromise NIR binding and activity. Here, we define features that characterize the anti-aggregation activity of NIR and NLS. We find that high binding affinity between NIR and NLS, and optimal NLS location relative to the aggregating domain plays a role in determining NIR disaggregation activity. A designed FUS chimera (FUSIBB), carrying the importin β binding (IBB) domain, is …
Structural Basis For Substrate Binding And Selection By Human Mitochondrial Rna Polymerase, Karl Herbine, Ashok Nayak, Dmitry Temiakov
Structural Basis For Substrate Binding And Selection By Human Mitochondrial Rna Polymerase, Karl Herbine, Ashok Nayak, Dmitry Temiakov
Department of Biochemistry and Molecular Biology Faculty Papers
The mechanism by which RNAP selects cognate substrates and discriminates between deoxy and ribonucleotides is of fundamental importance to the fidelity of transcription. Here, we present cryo-EM structures of human mitochondrial transcription elongation complexes that reveal substrate ATP bound in Entry and Insertion Sites. In the Entry Site, the substrate binds along the O helix of the fingers domain of mtRNAP but does not interact with the templating DNA base. Interactions between RNAP and the triphosphate moiety of the NTP in the Entry Site ensure discrimination against nucleosides and their diphosphate and monophosphate derivatives but not against non-cognate rNTPs and …
Enpp1 Enzyme Replacement Therapy Improves Ectopic Calcification But Does Not Rescue Skeletal Phenotype In A Mouse Model For Craniometaphyseal Dysplasia, Ernst Reichenberger, Kevin O'Brien, Ayano Hatori, Thomas Carpenter, Koen Van De Wetering, Lisa Flaman, Jennifer Howe, Daniel Ortiz, Yves Sabbagh, I-Ping Chen
Enpp1 Enzyme Replacement Therapy Improves Ectopic Calcification But Does Not Rescue Skeletal Phenotype In A Mouse Model For Craniometaphyseal Dysplasia, Ernst Reichenberger, Kevin O'Brien, Ayano Hatori, Thomas Carpenter, Koen Van De Wetering, Lisa Flaman, Jennifer Howe, Daniel Ortiz, Yves Sabbagh, I-Ping Chen
Jefferson Institute of Molecular Medicine Papers and Presentations
Craniometaphyseal dysplasia (CMD) is a rare genetic bone disorder, characterized by progressive thickening of craniofacial bones and flared metaphyses of long bones. Craniofacial hyperostosis leads to the obstruction of neural foramina and neurological symptoms such as facial palsy, blindness, deafness, or severe headache. Mutations in ANKH (mouse ortholog ANK), a transporter of small molecules such as citrate and ATP, are responsible for autosomal dominant CMD. Knock-in (KI) mice carrying an ANKF377del mutation (AnkKI/KI) replicate many features of human CMD. Pyrophosphate (PPi) levels in plasma are significantly reduced in AnkKI/KI mice. PPi is a potent inhibitor of …
Stem-Loop And Circle-Loop Tads Generated By Directional Pairing Of Boundary Elements Have Distinct Physical And Regulatory Properties, Wenfan Ke, Miki Fujioka, Paul Schedl, James Jaynes
Stem-Loop And Circle-Loop Tads Generated By Directional Pairing Of Boundary Elements Have Distinct Physical And Regulatory Properties, Wenfan Ke, Miki Fujioka, Paul Schedl, James Jaynes
Department of Biochemistry and Molecular Biology Faculty Papers
The chromosomes in multicellular eukaryotes are organized into a series of topologically independent loops called TADs. In flies, TADs are formed by physical interactions between neighboring boundaries. Fly boundaries exhibit distinct partner preferences, and pairing interactions between boundaries are typically orientation-dependent. Pairing can be head-to-tail or head-to-head. The former generates a stem-loop TAD, while the latter gives a circle-loop TAD. The TAD that encompasses the Drosophila even skipped (eve) gene is formed by the head-to-tail pairing of the nhomie and homie boundaries. To explore the relationship between loop topology and the physical and regulatory landscape, we flanked the nhomie boundary …
G12/13 Signaling In Asthma, Elizabeth L. Mcduffie, Reynold A Panettieri, Charles P. Scott
G12/13 Signaling In Asthma, Elizabeth L. Mcduffie, Reynold A Panettieri, Charles P. Scott
Department of Biochemistry and Molecular Biology Faculty Papers
Shortening of airway smooth muscle and bronchoconstriction are pathognomonic for asthma. Airway shortening occurs through calcium-dependent activation of myosin light chain kinase, and RhoA-dependent calcium sensitization, which inhibits myosin light chain phosphatase. The mechanism through which pro-contractile stimuli activate calcium sensitization is poorly understood. Our review of the literature suggests that pro-contractile G protein coupled receptors likely signal through G12/13 to activate RhoA and mediate calcium sensitization. This hypothesis is consistent with the effects of pro-contractile agonists on RhoA and Rho kinase activation, actin polymerization and myosin light chain phosphorylation. Recognizing the likely role of G12/13 signaling in the pathophysiology …
Mutant Androgen Receptor Induces Neurite Loss And Senescence Independently Of Are Binding In A Neuronal Model Of Sbma, Jordyn Karliner, Y Liu, Diane Merry
Mutant Androgen Receptor Induces Neurite Loss And Senescence Independently Of Are Binding In A Neuronal Model Of Sbma, Jordyn Karliner, Y Liu, Diane Merry
Department of Biochemistry and Molecular Biology Faculty Papers
Spinal and bulbar muscular atrophy (SBMA) is a slowly progressing neuromuscular disease caused by a polyglutamine (polyQ)-encoding CAG trinucleotide repeat expansion in the androgen receptor (AR) gene, leading to AR aggregation, lower motor neuron death, and muscle atrophy. AR is a ligand-activated transcription factor that regulates neuronal architecture and promotes axon regeneration; however, whether AR transcriptional functions contribute to disease pathogenesis is not fully understood. Using a differentiated PC12 cell model of SBMA, we identified dysfunction of polyQ-expanded AR in its regulation of neurite growth and maintenance. Specifically, we found that in the presence of androgens, polyQ-expanded AR inhibited neurite …
Conformational Alterations Of The Cell Surface Of Monomeric And Dimeric Β2m-Free Hla-I (Proto-Hla) May Enable Novel Immune Functions In Health And Disease, Mepur Ravindranath, Narendranath Ravindranath, Carly Amato-Menker, Fatiha El Hilali, Edward Filippone
Conformational Alterations Of The Cell Surface Of Monomeric And Dimeric Β2m-Free Hla-I (Proto-Hla) May Enable Novel Immune Functions In Health And Disease, Mepur Ravindranath, Narendranath Ravindranath, Carly Amato-Menker, Fatiha El Hilali, Edward Filippone
Division of Nephrology Faculty Papers
Human leukocyte antigens (HLAs) are polymorphic glycoproteins expressed on the cell surface of nucleated cells and consist of two classes, HLA class I and HLA class II. In contrast, in mice, these molecules, known as H-2, are expressed on both nucleated cells and erythrocytes. HLA-I molecules (Face-1) are heterodimers consisting of a polypeptide heavy chain (HC) and a light chain, B2-microglobulin (B2m). The heterodimers bind to antigenic peptides and present them to the T-cell receptors of CD8+ cytotoxic T lymphocytes. The HCs can also independently emerge on the cell surface as B2m-free HC monomers without peptides (Face-2). Early investigators suggested …
Frontotemporal Dementia-Like Disease Progression Elicited By Seeded Aggregation And Spread Of Fus, Sonia Vazquez-Sanchez, Britt Tilkin, Fatima Gasset-Rosa, Sitao Zhang, Diana Piol, Melissa Mcalonis-Downes, Jonathan Artates, Noe Govea-Perez, Yana Verresen, Lin Guo, Don Cleveland, James Shorter, Sandrine Da Cruz
Frontotemporal Dementia-Like Disease Progression Elicited By Seeded Aggregation And Spread Of Fus, Sonia Vazquez-Sanchez, Britt Tilkin, Fatima Gasset-Rosa, Sitao Zhang, Diana Piol, Melissa Mcalonis-Downes, Jonathan Artates, Noe Govea-Perez, Yana Verresen, Lin Guo, Don Cleveland, James Shorter, Sandrine Da Cruz
Department of Biochemistry and Molecular Biology Faculty Papers
RNA binding proteins have emerged as central players in the mechanisms of many neurodegenerative diseases. In particular, a proteinopathy of fused in sarcoma (FUS) is present in some instances of familial Amyotrophic lateral sclerosis (ALS) and about 10% of sporadic Frontotemporal lobar degeneration (FTLD). Here we establish that focal injection of sonicated human FUS fibrils into brains of mice in which ALS-linked mutant or wild-type human FUS replaces endogenous mouse FUS is sufficient to induce focal cytoplasmic mislocalization and aggregation of mutant and wild-type FUS which with time spreads to distal regions of the brain. Human FUS fibril-induced FUS aggregation …
Neuronal Lrp4 Directs The Development, Maturation And Cytoskeletal Organization Of Drosophila Peripheral Synapses, Alison Depew, Joseph Bruckner, Kate O'Connor-Giles, Timothy Mosca
Neuronal Lrp4 Directs The Development, Maturation And Cytoskeletal Organization Of Drosophila Peripheral Synapses, Alison Depew, Joseph Bruckner, Kate O'Connor-Giles, Timothy Mosca
Student Papers, Posters & Projects
Synaptic development requires multiple signaling pathways to ensure successful connections. Transmembrane receptors are optimally positioned to connect the synapse and the rest of the neuron, often acting as synaptic organizers to synchronize downstream events. One such organizer, the LDL receptor-related protein LRP4, is a cell surface receptor that has been most well-studied postsynaptically at mammalian neuromuscular junctions. Recent work, however, identified emerging roles, but how LRP4 acts as a presynaptic organizer and the downstream mechanisms of LRP4 are not well understood. Here, we show that LRP4 functions presynaptically at Drosophila neuromuscular synapses, acting in motoneurons to instruct pre- and postsynaptic …
Predictive And Prognostic Biomarkers And Tumor Antigens For Targeted Therapy In Urothelial Carcinoma, Aditya Eturi, Amman Bhasin, Kevin Zarrabi, William Tester
Predictive And Prognostic Biomarkers And Tumor Antigens For Targeted Therapy In Urothelial Carcinoma, Aditya Eturi, Amman Bhasin, Kevin Zarrabi, William Tester
Department of Medical Oncology Faculty Papers
Urothelial carcinoma (UC) is the fourth most prevalent cancer amongst males worldwide. While patients with non-muscle-invasive disease have a favorable prognosis, 25% of UC patients present with locally advanced disease which is associated with a 10-15% 5-year survival rate and poor overall prognosis. Muscle-invasive bladder cancer (MIBC) is associated with about 50% 5 year survival when treated by radical cystectomy or trimodality therapy; stage IV disease is associated with 10-15% 5 year survival. Current therapeutic modalities for MIBC include neoadjuvant chemotherapy, surgery and/or chemoradiation, although patients with relapsed or refractory disease have a poor prognosis. However, the rapid success of …
Discovery Of A Small-Molecule Inhibitor That Traps Polθ On Dna And Synergizes With Parp Inhibitors, William Fried, Mrityunjay Tyagi, Leonid Minakhin, Gurushankar Chandramouly, Taylor Tredinnick, Mercy Ramanjulu, William Auerbacher, Marissa L Calbert, Timur Rusanov, Trung Hoang, Nikita Borisonnik, Robert Betsch, John Krais, Yifan Wang, Umeshkumar Vekariya, John Gordon, George Morton, Tatiana Kent, Tomasz Skorski, Neil Johnson, Wayne Childers, Xiaojiang Chen, Richard Pomerantz
Discovery Of A Small-Molecule Inhibitor That Traps Polθ On Dna And Synergizes With Parp Inhibitors, William Fried, Mrityunjay Tyagi, Leonid Minakhin, Gurushankar Chandramouly, Taylor Tredinnick, Mercy Ramanjulu, William Auerbacher, Marissa L Calbert, Timur Rusanov, Trung Hoang, Nikita Borisonnik, Robert Betsch, John Krais, Yifan Wang, Umeshkumar Vekariya, John Gordon, George Morton, Tatiana Kent, Tomasz Skorski, Neil Johnson, Wayne Childers, Xiaojiang Chen, Richard Pomerantz
Department of Biochemistry and Molecular Biology Faculty Papers
The DNA damage response (DDR) protein DNA Polymerase θ (Polθ) is synthetic lethal with homologous recombination (HR) factors and is therefore a promising drug target in BRCA1/2 mutant cancers. We discover an allosteric Polθ inhibitor (Polθi) class with 4-6 nM IC50 that selectively kills HR-deficient cells and acts synergistically with PARP inhibitors (PARPi) in multiple genetic backgrounds. X-ray crystallography and biochemistry reveal that Polθi selectively inhibits Polθ polymerase (Polθ-pol) in the closed conformation on B-form DNA/DNA via an induced fit mechanism. In contrast, Polθi fails to inhibit Polθ-pol catalytic activity on A-form DNA/RNA in which the enzyme binds in …
Parp2 Promotes Break Induced Replication-Mediated Telomere Fragility In Response To Replication Stress, Daniela Muoio, Natalie Laspata, Rachel L Dannenberg, Caroline Curry, Simone Darkoa-Larbi, Mark Hedglin, Shikhar Uttam, Elise Fouquerel
Parp2 Promotes Break Induced Replication-Mediated Telomere Fragility In Response To Replication Stress, Daniela Muoio, Natalie Laspata, Rachel L Dannenberg, Caroline Curry, Simone Darkoa-Larbi, Mark Hedglin, Shikhar Uttam, Elise Fouquerel
Department of Biochemistry and Molecular Biology Faculty Papers
PARP2 is a DNA-dependent ADP-ribosyl transferase (ARTs) enzyme with Poly(ADP-ribosyl)ation activity that is triggered by DNA breaks. It plays a role in the Base Excision Repair pathway, where it has overlapping functions with PARP1. However, additional roles for PARP2 have emerged in the response of cells to replication stress. In this study, we demonstrate that PARP2 promotes replication stress-induced telomere fragility and prevents telomere loss following chronic induction of oxidative DNA lesions and BLM helicase depletion. Telomere fragility results from the activity of the break-induced replication pathway (BIR). During this process, PARP2 promotes DNA end resection, strand invasion and BIR-dependent …
Blocking The Dimerization Of Polyglutamine-Expanded Androgen Receptor Protects Cells From Dht-Induced Toxicity By Increasing Ar Turnover, Allison Lisberg, Yuhong Liu, Diane E. Merry
Blocking The Dimerization Of Polyglutamine-Expanded Androgen Receptor Protects Cells From Dht-Induced Toxicity By Increasing Ar Turnover, Allison Lisberg, Yuhong Liu, Diane E. Merry
Department of Biochemistry and Molecular Biology Faculty Papers
Spinal and bulbar muscular atrophy (SBMA) is a neuromuscular degenerative disease caused by a polyglutamine expansion in the androgen receptor (AR). This mutation causes AR to misfold and aggregate, contributing to toxicity in and degeneration of motor neurons and skeletal muscle. There is currently no effective treatment or cure for this disease. The role of an interdomain interaction between the amino- and carboxyl-termini of AR, termed the N/C interaction, has been previously identified as a component of androgen receptor-induced toxicity in cell and mouse models of SBMA. However, the mechanism by which this interaction contributes to disease pathology is unclear. …
Biomarkers For Managing Neurodegenerative Diseases, Lara Cheslow, Adam E. Snook, Scott A. Waldman
Biomarkers For Managing Neurodegenerative Diseases, Lara Cheslow, Adam E. Snook, Scott A. Waldman
Department of Pharmacology, Physiology, and Cancer Biology Faculty Papers
Neurological disorders are the leading cause of cognitive and physical disability worldwide, affecting 15% of the global population. Due to the demographics of aging, the prevalence of neurological disorders, including neurodegenerative diseases, will double over the next two decades. Unfortunately, while available therapies provide symptomatic relief for cognitive and motor impairment, there is an urgent unmet need to develop disease-modifying therapies that slow the rate of pathological progression. In that context, biomarkers could identify at-risk and prodromal patients, monitor disease progression, track responses to therapy, and parse the causality of molecular events to identify novel targets for further clinical investigation. …
Two Dot1 Enzymes Cooperatively Mediate Efficient Ubiquitin-Independent Histone H3 Lysine 76 Tri-Methylation In Kinetoplastids, Victoria Frisbie, Hideharu Hashimoto, Yixuan Xie, Francisca De Luna Vitorino, Josue Baeza, Tam Nguyen, Zhangerjiao Yuan, Janna Kiselar, Benjamin Garcia, Erik Debler
Two Dot1 Enzymes Cooperatively Mediate Efficient Ubiquitin-Independent Histone H3 Lysine 76 Tri-Methylation In Kinetoplastids, Victoria Frisbie, Hideharu Hashimoto, Yixuan Xie, Francisca De Luna Vitorino, Josue Baeza, Tam Nguyen, Zhangerjiao Yuan, Janna Kiselar, Benjamin Garcia, Erik Debler
Department of Biochemistry and Molecular Biology Faculty Papers
In higher eukaryotes, a single DOT1 histone H3 lysine 79 (H3K79) methyltransferase processively produces H3K79me2/me3 through histone H2B mono-ubiquitin interaction, while the kinetoplastid Trypanosoma brucei di-methyltransferase DOT1A and tri-methyltransferase DOT1B efficiently methylate the homologous H3K76 without H2B mono-ubiquitination. Based on structural and biochemical analyses of DOT1A, we identify key residues in the methyltransferase motifs VI and X for efficient ubiquitin-independent H3K76 methylation in kinetoplastids. Substitution of a basic to an acidic residue within motif VI (Gx6K) is essential to stabilize the DOT1A enzyme-substrate complex, while substitution of the motif X sequence VYGE by CAKS renders a rigid active-site …
Differentially Disrupted Spinal Cord And Muscle Energy Metabolism In Spinal And Bulbar Muscular Atrophy, Danielle Debartolo, Frederick Arnold, Y Liu, Elana Molotsky, Hsin-Yao Tang, Diane Merry
Differentially Disrupted Spinal Cord And Muscle Energy Metabolism In Spinal And Bulbar Muscular Atrophy, Danielle Debartolo, Frederick Arnold, Y Liu, Elana Molotsky, Hsin-Yao Tang, Diane Merry
Department of Biochemistry and Molecular Biology Faculty Papers
Prior studies showed that polyglutamine-expanded androgen receptor (AR) is aberrantly acetylated and that deacetylation of the mutant AR by overexpression of nicotinamide adenine dinucleotide-dependent (NAD+-dependent) sirtuin 1 is protective in cell models of spinal and bulbar muscular atrophy (SBMA). Based on these observations and reduced NAD+ in muscles of SBMA mouse models, we tested the therapeutic potential of NAD+ restoration in vivo by treating postsymptomatic transgenic SBMA mice with the NAD+ precursor nicotinamide riboside (NR). NR supplementation failed to alter disease progression and had no effect on increasing NAD+ or ATP content in muscle, despite producing a modest increase of …
Tail-Tape-Fused Virion And Non-Virion Rna Polymerases Of A Thermophilic Virus With An Extremely Long Tail, Anastasiia Chaban, Leonid Minakhin, Ekaterina Goldobina, Brain Bae, Yue Hao, Sergei Borukhov, Leena Putzeys, Maarten Boon, Florian Kabinger, Rob Lavigne, Kira Makarova, Eugene V Koonin, Satish Nair, Shunsuke Tagami, Konstantin Severinov, Maria Sokolova
Tail-Tape-Fused Virion And Non-Virion Rna Polymerases Of A Thermophilic Virus With An Extremely Long Tail, Anastasiia Chaban, Leonid Minakhin, Ekaterina Goldobina, Brain Bae, Yue Hao, Sergei Borukhov, Leena Putzeys, Maarten Boon, Florian Kabinger, Rob Lavigne, Kira Makarova, Eugene V Koonin, Satish Nair, Shunsuke Tagami, Konstantin Severinov, Maria Sokolova
Department of Biochemistry and Molecular Biology Faculty Papers
Thermus thermophilus bacteriophage P23-45 encodes a giant 5,002-residue tail tape measure protein (TMP) that defines the length of its extraordinarily long tail. Here, we show that the N-terminal portion of P23-45 TMP is an unusual RNA polymerase (RNAP) homologous to cellular RNAPs. The TMP-fused virion RNAP transcribes pre-early phage genes, including a gene that encodes another, non-virion RNAP, that transcribes early and some middle phage genes. We report the crystal structures of both P23-45 RNAPs. The non-virion RNAP has a crab-claw-like architecture. By contrast, the virion RNAP adopts a unique flat structure without a clamp. Structure and sequence comparisons of …
Structural Basis For Dna Proofreading, Gina Buchel, Ashok Nayak, Karl Herbine, Azadeh Sarfallah, Viktoriia Sokolova, Angelica Zamudio-Ochoa, Dmitry Temiakov
Structural Basis For Dna Proofreading, Gina Buchel, Ashok Nayak, Karl Herbine, Azadeh Sarfallah, Viktoriia Sokolova, Angelica Zamudio-Ochoa, Dmitry Temiakov
Department of Biochemistry and Molecular Biology Faculty Papers
DNA polymerase (DNAP) can correct errors in DNA during replication by proofreading, a process critical for cell viability. However, the mechanism by which an erroneously incorporated base translocates from the polymerase to the exonuclease site and the corrected DNA terminus returns has remained elusive. Here, we present an ensemble of nine high-resolution structures representing human mitochondrial DNA polymerase Gamma, Polγ, captured during consecutive proofreading steps. The structures reveal key events, including mismatched base recognition, its dissociation from the polymerase site, forward translocation of DNAP, alterations in DNA trajectory, repositioning and refolding of elements for primer separation, DNAP backtracking, and displacement …
Novel Treatments For Pxe: Targeting The Systemic And Local Drivers Of Ectopic Calcification, Ida Joely Jacobs, Qiaoli Li
Novel Treatments For Pxe: Targeting The Systemic And Local Drivers Of Ectopic Calcification, Ida Joely Jacobs, Qiaoli Li
Department of Biochemistry and Molecular Biology Faculty Papers
Pseudoxanthoma elasticum (PXE) is a heritable multisystem ectopic calcification disorder. The gene responsible for PXE, ABCC6, encodes ABCC6, a hepatic efflux transporter regulating extracellular inorganic pyrophosphate (PPi), a potent endogenous calcification inhibitor. Recent studies demonstrated that in addition to the deficiency of plasma PPi, the activated DDR/PARP signaling in calcified tissues provides an additional possible mechanism of ectopic calcification in PXE. This study examined the effects of etidronate (ETD), a stable PPi analog, and its combination with minocycline (Mino), a potent inhibitor of DDR/PARP, on ectopic calcification in an Abcc6-/- mouse model of PXE. Abcc6-/- mice, at 4 weeks of …
Loss Of Pml Nuclear Bodies In Familial Amyotrophic Lateral Sclerosis-Frontotemporal Dementia, Francesco Antoniani, Marco Cimino, Laura Mediani, Jonathan Vinet, Enza M. Verde, Valentina Secco, Alfred Yamoah, Priyanka Tripathi, Eleonora Aronica, Maria Elena Cicardi, Davide Trotti, Jared Sterneckert, Anand Goswami, Serena Carra
Loss Of Pml Nuclear Bodies In Familial Amyotrophic Lateral Sclerosis-Frontotemporal Dementia, Francesco Antoniani, Marco Cimino, Laura Mediani, Jonathan Vinet, Enza M. Verde, Valentina Secco, Alfred Yamoah, Priyanka Tripathi, Eleonora Aronica, Maria Elena Cicardi, Davide Trotti, Jared Sterneckert, Anand Goswami, Serena Carra
Farber Institute for Neuroscience Faculty Papers
Amyotrophic Lateral Sclerosis (ALS) and Frontotemporal Dementia (FTD) are two neurodegenerative disorders that share genetic causes and pathogenic mechanisms. The critical genetic players of ALS and FTD are the TARDBP, FUS and C9orf72 genes, whose protein products, TDP-43, FUS and the C9orf72-dipeptide repeat proteins, accumulate in form of cytoplasmic inclusions. The majority of the studies focus on the understanding of how cells control TDP-43 and FUS aggregation in the cytoplasm, overlooking how dysfunctions occurring at the nuclear level may influence the maintenance of protein solubility outside of the nucleus. However, protein quality control (PQC) systems that maintain protein homeostasis comprise …
High-Resolution Cryo-Em Structure Of The Pseudomonas Bacteriophage E217, Fenglin Li, Chun-Feng Hou, Ravi K. Lokareddy, Ruoyu Yang, Francesca Forti, Federica Briani, Gino Cingolani
High-Resolution Cryo-Em Structure Of The Pseudomonas Bacteriophage E217, Fenglin Li, Chun-Feng Hou, Ravi K. Lokareddy, Ruoyu Yang, Francesca Forti, Federica Briani, Gino Cingolani
Department of Biochemistry and Molecular Biology Faculty Papers
E217 is a Pseudomonas phage used in an experimental cocktail to eradicate cystic fibrosis-associated Pseudomonas aeruginosa. Here, we describe the structure of the whole E217 virion before and after DNA ejection at 3.1 Å and 4.5 Å resolution, respectively, determined using cryogenic electron microscopy (cryo-EM). We identify and build de novo structures for 19 unique E217 gene products, resolve the tail genome-ejection machine in both extended and contracted states, and decipher the complete architecture of the baseplate formed by 66 polypeptide chains. We also determine that E217 recognizes the host O-antigen as a receptor, and we resolve the N-terminal portion …
Fibrosis-The Tale Of H3k27 Histone Methyltransferases And Demethylases, Morgan D. Basta, Svetlana Petruk, Alexander Mazo, Janice L. Walker
Fibrosis-The Tale Of H3k27 Histone Methyltransferases And Demethylases, Morgan D. Basta, Svetlana Petruk, Alexander Mazo, Janice L. Walker
Department of Biochemistry and Molecular Biology Faculty Papers
Fibrosis, or excessive scarring, is characterized by the emergence of alpha-smooth muscle actin (αSMA)-expressing myofibroblasts and the excessive accumulation of fibrotic extracellular matrix (ECM). Currently, there is a lack of effective treatment options for fibrosis, highlighting an unmet need to identify new therapeutic targets. The acquisition of a fibrotic phenotype is associated with changes in chromatin structure, a key determinant of gene transcription activation and repression. The major repressive histone mark, H3K27me3, has been linked to dynamic changes in gene expression in fibrosis through alterations in chromatin structure. H3K27-specific homologous histone methylase (HMT) enzymes, Enhancer of zeste 1 and 2 …
Molecular Mechanisms Protecting Centromeres From Self-Sabotage And Implications For Cancer Therapy, Rim Nassar, Lily Thompson, Elise Fouquerel
Molecular Mechanisms Protecting Centromeres From Self-Sabotage And Implications For Cancer Therapy, Rim Nassar, Lily Thompson, Elise Fouquerel
Student Papers, Posters & Projects
Centromeres play a crucial role in DNA segregation by mediating the cohesion and separation of sister chromatids during cell division. Centromere dysfunction, breakage or compromised centromeric integrity can generate aneuploidies and chromosomal instability, which are cellular features associated with cancer initiation and progression. Maintaining centromere integrity is thus essential for genome stability. However, the centromere itself is prone to DNA breaks, likely due to its intrinsically fragile nature. Centromeres are complex genomic loci that are composed of highly repetitive DNA sequences and secondary structures and require the recruitment and homeostasis of a centromere-associated protein network. The molecular mechanisms engaged to …
A Label-Free Assay For Aminoacylation Of Trna, Howard Gamper, Ya-Ming Hou
A Label-Free Assay For Aminoacylation Of Trna, Howard Gamper, Ya-Ming Hou
Department of Biochemistry and Molecular Biology Faculty Papers
Aminoacylation of tRNA generates an aminoacyl-tRNA (aa-tRNA) that is active for protein synthesis on the ribosome. Quantification of aminoacylation of tRNA is critical to understand the mechanism of specificity and the flux of the aa-tRNA into the protein synthesis machinery, which determines the rate of cell growth. Traditional assays for the quantification of tRNA aminoacylation involve radioactivity, either with a radioactive amino acid or with a [3'-32P]-labeled tRNA. We describe here a label-free assay that monitors aminoacylation by biotinylation-streptavidin (SA) conjugation to the α-amine or the α-imine of the aminoacyl group on the aa-tRNA. The conjugated aa-tRNA product is readily …