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Articles 1 - 30 of 68

Full-Text Articles in Biochemistry, Biophysics, and Structural Biology

Yeast Sirtuin Family Members Maintain Transcription Homeostasis To Ensure Genome Stability, Jessica L. Feldman, Craig L. Peterson Jun 2019

Yeast Sirtuin Family Members Maintain Transcription Homeostasis To Ensure Genome Stability, Jessica L. Feldman, Craig L. Peterson

Program in Molecular Medicine Publications and Presentations

The mammalian sirtuin, SIRT6, is a key tumor suppressor that maintains genome stability and regulates transcription, though how SIRT6 family members control genome stability is unclear. Here, we use multiple genome-wide approaches to demonstrate that the yeast SIRT6 homologs, Hst3 and Hst4, prevent genome instability by tuning levels of both coding and noncoding transcription. While nascent RNAs are elevated in the absence of Hst3 and Hst4, a global impact on steady-state mRNAs is masked by the nuclear exosome, indicating that sirtuins and the exosome provide two levels of regulation to maintain transcription homeostasis. We find that, in the absence of ...


Circular Dichroism And Molecular Modeling Yield A Structure For The Complex Of Human Immunodeficiency Virus Type 1 Trans-Activation Response Rna And The Binding Region Of Tat, The Trans-Acting Transcriptional Activator, Erwann P. Loret, Philippe T. Georgel, W. Curtis Johnson Jr., Pui Shing Ho May 2019

Circular Dichroism And Molecular Modeling Yield A Structure For The Complex Of Human Immunodeficiency Virus Type 1 Trans-Activation Response Rna And The Binding Region Of Tat, The Trans-Acting Transcriptional Activator, Erwann P. Loret, Philippe T. Georgel, W. Curtis Johnson Jr., Pui Shing Ho

Philippe T. Georgel

Transcription in the human immunodeficiency virus type 1 (HIV-1) retrovirus is regulated by binding the viral Tat protein (trans-acting transcriptional activator) to the trans-activation response (TAR) RNA sequence. Here, vacuum UV circular dichroism (VUV-CD) is used to study the structure of TAR and its complex with two peptide fragments that are important for Tat binding to TAR. The VUV-CD spectrum of TAR is typical of A-form RNA and is minimally perturbed when bound to either the short or the long Tat peptide. The CD spectra ofthe complexes indicate an extended structure in the argnine-rich region of Tat from amino acid ...


The Worlds Of Splicing And Chromatin Collide, J. Adam Hall, Philippe T. Georgel May 2019

The Worlds Of Splicing And Chromatin Collide, J. Adam Hall, Philippe T. Georgel

Philippe T. Georgel

Both transcription and splicing take place in a nuclear environment which, at face value, may seem refractory to the efficiency afforded by the coupling of both processes. This environment, chromatin, was once viewed as only a passive packaging system for genetic material, with very little contribution to the variety of nuclear activities occurring within and around it. However, overwhelming evidence now points to the chromatin environment as being highly dynamic, and an active player in nuclear activities.


Transient Kinetic Analysis Of Swr1c-Catalyzed H2a.Z Deposition Unravels The Impact Of Nucleosome Dynamics And The Asymmetry Of Histone Exchange, Raushan K. Singh, Jiayi Fan, Nathan Gioacchini, Shinya Watanabe, Osman Bilsel, Craig L. Peterson Apr 2019

Transient Kinetic Analysis Of Swr1c-Catalyzed H2a.Z Deposition Unravels The Impact Of Nucleosome Dynamics And The Asymmetry Of Histone Exchange, Raushan K. Singh, Jiayi Fan, Nathan Gioacchini, Shinya Watanabe, Osman Bilsel, Craig L. Peterson

Open Access Articles

The SWR1C chromatin remodeling enzyme catalyzes ATP-dependent replacement of nucleosomal H2A with the H2A.Z variant, regulating key DNA-mediated processes such as transcription and DNA repair. Here, we investigate the transient kinetic mechanism of the histone exchange reaction, employing ensemble FRET, fluorescence correlation spectroscopy (FCS), and the steady-state kinetics of ATP hydrolysis. Our studies indicate that SWR1C modulates nucleosome dynamics on both the millisecond and microsecond timescales, poising the nucleosome for the dimer exchange reaction. The transient kinetic analysis of the remodeling reaction performed under single turnover conditions unraveled a striking asymmetry in the ATP-dependent replacement of nucleosomal dimers, promoted ...


Protein Degradation Regulates Phospholipid Biosynthetic Gene Expression In Saccharomyces Cerevisiae, Bryan Salas-Santiago Jan 2019

Protein Degradation Regulates Phospholipid Biosynthetic Gene Expression In Saccharomyces Cerevisiae, Bryan Salas-Santiago

Doctoral Dissertations

Transcriptional regulation of most phospholipid biosynthetic genes in Saccharomyces cerevisiae is coordinated by inositol and choline. Inositol affects phosphatidic acid (PA) intracellular levels. Opi1p interacts physically with PA and is the main repressor of the phospholipid biosynthetic genes. It is localized in the endoplasmic reticulum (ER) bound to the ER membrane protein Scs2p. When PA levels drop, Opi1p is translocated into the nucleus repressing most phospholipid biosynthetic genes. The OPI1 locus was identified in a screen looking for overproduction and excretion of inositol (Opi-). Opi- mutants are generally associated with a defect in repression of the ...


Transcriptional Correlates Of Proximal-Distal Identify And Regeneration Timing In Axolotl Limbs, S. Randal Voss, David Murrugarra, Tyler B. Jensen, James R Monaghan Jun 2018

Transcriptional Correlates Of Proximal-Distal Identify And Regeneration Timing In Axolotl Limbs, S. Randal Voss, David Murrugarra, Tyler B. Jensen, James R Monaghan

Neuroscience Faculty Publications

Cells within salamander limbs retain memories that inform the correct replacement of amputated tissues at different positions along the length of the arm, with proximal and distal amputations completing regeneration at similar times. We investigated the possibility that positional memory is associated with variation in transcript abundances along the proximal-distal limb axis. Transcripts were deeply sampled from Ambystoma mexicanum limbs at the time they were administered fore arm vs upper arm amputations, and at 19 post-amputation time points. After amputation and prior to regenerative outgrowth, genes typically expressed by differentiated muscle cells declined more rapidly in upper arms while cell ...


Transcriptome-Wide Identification Of The Rna-Binding Landscape Of The Chromatin-Associated Protein Parp1 Reveals Functions In Rna Biogenesis, Manana Melikishvili, Julia H. Chariker, Eric C. Rouchka, Yvonne N. Fondufe-Mittendorf Nov 2017

Transcriptome-Wide Identification Of The Rna-Binding Landscape Of The Chromatin-Associated Protein Parp1 Reveals Functions In Rna Biogenesis, Manana Melikishvili, Julia H. Chariker, Eric C. Rouchka, Yvonne N. Fondufe-Mittendorf

Molecular and Cellular Biochemistry Faculty Publications

Recent studies implicate Poly (ADP-ribose) polymerase 1 (PARP1) in alternative splicing regulation, and PARP1 may be an RNA-binding protein. However, detailed knowledge of RNA targets and the RNA-binding region for PARP1 are unknown. Here we report the first global study of PARP1–RNA interactions using PAR–CLIP in HeLa cells. We identified a largely overlapping set of 22 142 PARP1–RNA-binding peaks mapping to mRNAs, with 20 484 sites located in intronic regions. PARP1 preferentially bound RNA containing GC-rich sequences. Using a Bayesian model, we determined positional effects of PARP1 on regulated exon-skipping events: PARP1 binding upstream and downstream of ...


Casein Kinase 2-Mediated Phosphorylation Of Brahma-Related Gene 1 Controls Myoblast Proliferation And Contributes To Swi/Snf Complex Composition, Teresita Padilla-Benavides, Brian T. Nasipak, Amanda L. Paskavitz, Dominic T. Haokip, Jake M. Schnabl, Jeffrey A. Nickerson, Anthony N. Imbalzano Nov 2017

Casein Kinase 2-Mediated Phosphorylation Of Brahma-Related Gene 1 Controls Myoblast Proliferation And Contributes To Swi/Snf Complex Composition, Teresita Padilla-Benavides, Brian T. Nasipak, Amanda L. Paskavitz, Dominic T. Haokip, Jake M. Schnabl, Jeffrey A. Nickerson, Anthony N. Imbalzano

UMass Metabolic Network Publications

Transcriptional regulation is modulated in part by chromatin-remodeling enzymes that control gene accessibility by altering chromatin compaction or nucleosome positioning. Brahma-related gene 1 (Brg1), a catalytic subunit of the mammalian SWI/SNF chromatin-remodeling enzymes, is required for both myoblast proliferation and differentiation, and the control of Brg1 phosphorylation by calcineurin, PKCbeta1, and p38 regulates the transition to differentiation. However, we hypothesized that Brg1 activity might be regulated by additional kinases. Here, we report that Brg1 is also a target of casein kinase 2 (CK2), a serine/threonine kinase, in proliferating myoblasts. We found that CK2 interacts with Brg1, and mutation ...


Structure Of Rna Polymerase Bound To Ribosomal 30s Subunit, Gabriel Demo, Aviram Rasouly, Nikita Vasilyev, Vladimir Svetlov, Anna B. Loveland, Ruben Diaz-Avalos, Nikolaus Grigorieff, Evgeny Nudler, Andrei A. Korostelev Oct 2017

Structure Of Rna Polymerase Bound To Ribosomal 30s Subunit, Gabriel Demo, Aviram Rasouly, Nikita Vasilyev, Vladimir Svetlov, Anna B. Loveland, Ruben Diaz-Avalos, Nikolaus Grigorieff, Evgeny Nudler, Andrei A. Korostelev

Open Access Articles

In bacteria, mRNA transcription and translation are coupled to coordinate optimal gene expression and maintain genome stability. Coupling is thought to involve direct interactions between RNA polymerase (RNAP) and the translational machinery. We present cryo-EM structures of E. coli RNAP core bound to the small ribosomal 30S subunit. The complex is stable under cell-like ionic conditions, consistent with functional interaction between RNAP and the 30S subunit. The RNA exit tunnel of RNAP aligns with the Shine-Dalgarno-binding site of the 30S subunit. Ribosomal protein S1 forms a wall of the tunnel between RNAP and the 30S subunit, consistent with its role ...


A Synthetic Biology Approach To Probing Nucleosome Symmetry, Yuichi Ichikawa, Caitlin M. Connolly, Hsin-Jung Chou, Yuanyuan Chen, Upasna Sharma, Hsuiyi V. Chen, Vineeta Bajaj, Daniel Na. Bolon, Oliver J. Rando, Paul D. Kaufman Sep 2017

A Synthetic Biology Approach To Probing Nucleosome Symmetry, Yuichi Ichikawa, Caitlin M. Connolly, Hsin-Jung Chou, Yuanyuan Chen, Upasna Sharma, Hsuiyi V. Chen, Vineeta Bajaj, Daniel Na. Bolon, Oliver J. Rando, Paul D. Kaufman

UMass Metabolic Network Publications

The repeating subunit of chromatin, the nucleosome, includes two copies of each of the four core histones, and several recent studies have reported that asymmetrically-modified nucleosomes occur at regulatory elements in vivo. To probe the mechanisms by which histone modifications are read out, we designed an obligate pair of H3 heterodimers, termed H3X and H3Y, which we extensively validated genetically and biochemically. Comparing the effects of asymmetric histone tail point mutants with those of symmetric double mutants revealed that a single methylated H3K36 per nucleosome was sufficient to silence cryptic transcription in vivo. We also demonstrate the utility of this ...


Transcription Factor Hlx Controls A Systematic Switch From White To Brown Fat Through Prdm16-Mediated Co-Activation, Lei Huang, Dongning Pan, Qingbo Chen, Lihua (Julie) Zhu, Jianhong Ou, Martin Wabitsch, Yong-Xu Wang Jul 2017

Transcription Factor Hlx Controls A Systematic Switch From White To Brown Fat Through Prdm16-Mediated Co-Activation, Lei Huang, Dongning Pan, Qingbo Chen, Lihua (Julie) Zhu, Jianhong Ou, Martin Wabitsch, Yong-Xu Wang

Open Access Articles

Browning of subcutaneous white fat (iWAT) involves several reprograming events, but the underlying mechanisms are incompletely understood. Here we show that the transcription factor Hlx is selectively expressed in brown adipose tissue (BAT) and iWAT, and is translationally upregulated by beta3-adrenergic signaling-mediated suppression of the translational inhibitor 4E-BP1. Hlx interacts with and is co-activated by Prdm16 to control BAT-selective gene expression and mitochondrial biogenesis. Hlx heterozygous knockout mice have defects in brown-like adipocyte formation in iWAT, and develop glucose intolerance and high fat-induced hepatic steatosis. Conversely, transgenic expression of Hlx at a physiological level drives a full program of thermogenesis ...


Effects Of Nicotine On The Cyp6a8 Gene Promoter Of Drosophila Melanogaster, Leslie M. Stroud May 2017

Effects Of Nicotine On The Cyp6a8 Gene Promoter Of Drosophila Melanogaster, Leslie M. Stroud

Chancellor’s Honors Program Projects

No abstract provided.


Zhx2 (Zinc Fingers And Homeoboxes 2) Regulates Major Urinary Protein Gene Expression In The Mouse Liver, Jieyun Jiang, Kate Townsend Creasy, Justin Purnell, Martha L. Peterson, Brett T. Spear Mar 2017

Zhx2 (Zinc Fingers And Homeoboxes 2) Regulates Major Urinary Protein Gene Expression In The Mouse Liver, Jieyun Jiang, Kate Townsend Creasy, Justin Purnell, Martha L. Peterson, Brett T. Spear

Microbiology, Immunology, and Molecular Genetics Faculty Publications

The mouse major urinary proteins (Mups) are encoded by a large family of highly related genes clustered on chromosome 4. Mups, synthesized primarily and abundantly in the liver and secreted through the kidneys, exhibit male-biased expression. Mups bind a variety of volatile ligands; these ligands, and Mup proteins themselves, influence numerous behavioral traits. Although urinary Mup protein levels vary between inbred mouse strains, this difference is most pronounced in BALB/cJ mice, which have dramatically low urinary Mup levels; this BALB/cJ trait had been mapped to a locus on chromosome 15. We previously identified Zhx2 (zinc fingers and homeoboxes ...


Development Of A Novel Method To Express And Purify Vaccinia Virus Early Transcription Factors A7 And D6 Using Bacteria, Omkar M. Gandbhir Jan 2017

Development Of A Novel Method To Express And Purify Vaccinia Virus Early Transcription Factors A7 And D6 Using Bacteria, Omkar M. Gandbhir

Seton Hall University Dissertations and Theses (ETDs)

The eradication of Smallpox is one of the greatest human achievements. However other poxviruses still exist infecting humans (Molluscum Contagiosum, monkeypox, and Vaccinia) or livestock (cowpox, sheeppox). Though vaccines and anti-virals exist, the side effects are serious, and viral resistance has been detected. The genome amongst poxviruses is highly conserved, early gene promoters maintain a specific critical consensus region that can be targeted for development of novel broad spectrum therapeutics. Using Vaccinia early transcription factors (VETFs) a novel method of expressing and purifying VETF A7 and D6 was developed. A7 and D6 are required for transcription of early genes recruiting ...


The Regenerating Fin As A Model To Examine The Skeletal Defects Of Roberts Syndrome, Rajeswari Banerji Jan 2017

The Regenerating Fin As A Model To Examine The Skeletal Defects Of Roberts Syndrome, Rajeswari Banerji

Theses and Dissertations

Roberts Syndrome (RBS) is a human developmental disorder characterized by craniofacial abnormalities, limb malformation and often severe mental retardation. RBS arises from mutations in the cohesin auxiliary factor ESCO2 that targets the SMC3 subunit of the cohesin complex. Mutations in cohesin subunits and a subset of cohesin auxiliary factors gives rise to a related developmental malady termed Cornelia de Lange Syndrome (CdLS). They are collectively termed cohesinopathies since both disorders comprise overlapping phenotypes and the causative genes for both syndromes perform common activities. The underlying cause of CdLS is largely modeled as occurring through transcriptional deregulation. Whereas, the mechanism that ...


Significance Of Pten Phosphorylation And Its Nuclear Function In Lung Cancer, Prerna Malaney Nov 2016

Significance Of Pten Phosphorylation And Its Nuclear Function In Lung Cancer, Prerna Malaney

Graduate Theses and Dissertations

Phosphorylation mediated inactivation of PTEN leads to multiple malignancies with increased severity. However, the consequence of such inactivation on downstream functions of PTEN are poorly understood. Therefore, the objective of my thesis is to ascertain the molecular mechanisms by which PTEN phosphorylation drives lung cancer. PTEN phosphorylation at the C-terminal serine/threonine cluster abrogates its tumor suppressor function. Despite the critical role of the PTEN C-tail in regulating its function, the crystal structure of the C-tail remains unknown. Using bioinformatics and structural analysis, I determined that the PTEN C-tail is an intrinsically disordered region and is a hot spot for ...


Rna Polymerase Ii Depletion Promotes Transcription Of Alternative Mrna Species, Lijian Yu, Mayuri Rege, Craig L. Peterson, Michael R. Volkert Aug 2016

Rna Polymerase Ii Depletion Promotes Transcription Of Alternative Mrna Species, Lijian Yu, Mayuri Rege, Craig L. Peterson, Michael R. Volkert

Open Access Articles

BACKGROUND: Cells respond to numerous internal and external stresses, such as heat, cold, oxidative stress, DNA damage, and osmotic pressure changes. In most cases, the primary response to stress is transcriptional induction of genes that assist the cells in tolerating the stress and facilitate the repair of the cellular damage. However, when the transcription machinery itself is stressed, responding by such standard mechanisms may not be possible.

RESULTS: In this study, we demonstrate that depletion or inactivation of RNA polymerase II (RNAPII) changes the preferred polyadenylation site usage for several transcripts, and leads to increased transcription of a specific subset ...


Regulation Of Chaperone Binding And Nucleosome Dynamics By Key Residues Within The Globular Domain Of Histone H3, Sarah J. Hainer, Joseph A. Martens Apr 2016

Regulation Of Chaperone Binding And Nucleosome Dynamics By Key Residues Within The Globular Domain Of Histone H3, Sarah J. Hainer, Joseph A. Martens

Open Access Articles

BACKGROUND: Nucleosomes have an important role in modulating access of DNA by regulatory factors. The role specific histone residues have in this process has been shown to be an important mechanism of transcription regulation. Previously, we identified eight amino acids in histones H3 and H4 that are required for nucleosome occupancy over highly transcribed regions of the genome.

RESULTS: We investigate the mechanism through which three of these previously identified histone H3 amino acids regulate nucleosome architecture. We find that histone H3 K122, Q120, and R49 are required for Spt2, Spt6, and Spt16 occupancies at genomic locations where transcription rates ...


The Mitotic Genome: Accessibility And Transcriptional Control, Chris Hsiung Jan 2016

The Mitotic Genome: Accessibility And Transcriptional Control, Chris Hsiung

Publicly Accessible Penn Dissertations

Mitosis entails dramatic global alterations to genome structure and regulation, including

chromosome condensation, dissociation of the transcriptional machinery from chromosomes, and transcriptional silencing. Here I report studies that address the macromolecular accessibility of the mitotic genome and the control of transcriptional reactivation upon mitotic exit in a mammalian cell line. The results obtained from measuring the sensitivity of chromatin to DNase I cleavage by sequencing (DNase-seq) in pure mitotic cell populations demonstrate that macromolecular accessibility of the mitotic genome is widely preserved. Thus, steric hindrance from chromatin condensation is insufficient for explaining the eviction of transcription factors from mitotic chromatin ...


Examining The Role Of Rna Secondary Structure On Human Rna Polymerase Ii Pausing, Meagan N. Paul Jan 2016

Examining The Role Of Rna Secondary Structure On Human Rna Polymerase Ii Pausing, Meagan N. Paul

Undergraduate Honors Theses

The process of transcription, in which DNA is converted to RNA, is an integral process in the expression of our genetic information. Regulation of transcription is critical for healthy, functioning cells and defects in transcriptional regulation can cause a plethora of ill health effects including cancer and improper cardiac or neuronal development. Human RNA Polymerase II (Pol II) is the enzyme that transcribes all protein-coding and most non-coding RNA genes in the human genome. Within the past 10 years, it has become evident that Pol II function is regulated after it initiates transcription; specifically, the Pol II enzyme pauses after ...


Cadmium-Induced Transcription By Nfatc2 In Breast Cancer Cells, Natalie Miller Jan 2016

Cadmium-Induced Transcription By Nfatc2 In Breast Cancer Cells, Natalie Miller

Undergraduate Honors Theses

Although breast cancer is the second leading cause of cancer death among women in the United States, many of the mechanisms behind breast cancer development are unknown. It is known, however, that mis-regulated gene expression contributes to cancer development. Gene expression is tightly regulated at transcriptional initiation both by the formation of the pre-initiation complex and assembly of a host of other transcription factors, which act to promote expression of specific genes. NFATc2 is one such transcription factor implicated in breast cancer progression. This protein is regulated by intracellular calcium levels that may be induced by exposure to the environmental ...


Regulation Of Gene Expression By The Mediator Kinase Cdk19, Katherine Audrey Audetat Jan 2016

Regulation Of Gene Expression By The Mediator Kinase Cdk19, Katherine Audrey Audetat

Molecular, Cellular, and Developmental Biology Graduate Theses & Dissertations

The Mediator complex is a required co-activator of RNA Polymerase II (Pol II), the enzyme responsible for the transcription of all protein coding genes in eukaryotes. Mediator facilitates transcription factor-dependent gene expression by directly interacting with gene specific transcription factors and Pol II and the general transcription machinery. Mediator consists of up to 29 subunits; a four-subunit “CDK module” reversibly associates with Core-Mediator and alters its structure and activity. Most of the work on the CDK module has been done in yeast, and has focused on the CDK module subunits CDK8, MED12, and MED13. These three subunits have paralog proteins ...


Hepatic Nutrient And Hormonal Regulation Of The Pancreatic-Derived Factor (Pander) Promoter, Whitney Ratliff Nov 2015

Hepatic Nutrient And Hormonal Regulation Of The Pancreatic-Derived Factor (Pander) Promoter, Whitney Ratliff

Graduate Theses and Dissertations

PANcreatic-DERived factor (PANDER, FAM3B) has been shown to regulate glycemic levels via interactions with both pancreatic islets and the liver. Although PANDER is predominantly expressed from the endocrine pancreas, recent work has provided sufficient evidence that the liver may also be an additional tissue source of PANDER production. At physiological levels, PANDER is capable of disrupting insulin signaling and promoting increased hepatic glucose production. As shown in some animal models, strong expression of PANDER, induced by viral delivery within the liver, induces hepatic steatosis. However, no studies to date have explicitly characterized the transcriptional regulation of PANDER from the liver ...


Genome-Wide Profiling Of Parp1 Reveals An Interplay With Gene Regulatory Regions And Dna Methylation, Narasimharao Nalabothula, Taha Al-Jumaily, Abdallah M. Eteleeb, Robert M. Flight, Shao Xiaorong, Hunter Moseley, Eric C. Rouchka, Yvonne N. Fondufe-Mittendorf Aug 2015

Genome-Wide Profiling Of Parp1 Reveals An Interplay With Gene Regulatory Regions And Dna Methylation, Narasimharao Nalabothula, Taha Al-Jumaily, Abdallah M. Eteleeb, Robert M. Flight, Shao Xiaorong, Hunter Moseley, Eric C. Rouchka, Yvonne N. Fondufe-Mittendorf

Molecular and Cellular Biochemistry Faculty Publications

Poly (ADP-ribose) polymerase-1 (PARP1) is a nuclear enzyme involved in DNA repair, chromatin remodeling and gene expression. PARP1 interactions with chromatin architectural multi-protein complexes (i.e. nucleosomes) alter chromatin structure resulting in changes in gene expression. Chromatin structure impacts gene regulatory processes including transcription, splicing, DNA repair, replication and recombination. It is important to delineate whether PARP1 randomly associates with nucleosomes or is present at specific nucleosome regions throughout the cell genome. We performed genome-wide association studies in breast cancer cell lines to address these questions. Our studies show that PARP1 associates with epigenetic regulatory elements genome-wide, such as active ...


Transcriptional Activity Of The Islet Β Cell Factor Pdx1 Is Augmented By Lysine Methylation Catalyzed By The Methyltransferase Set7/9, Aarthi V. Maganti, Bernhard Maier, Sarah A. Tersey, Megan L. Sampley, Amber L. Mosley, Sabire Özcan, Boobalan Pachaiyappan, Patrick M. Woster, Chad S. Hunter, Roland Stein, Raghavendra G. Mirmira Apr 2015

Transcriptional Activity Of The Islet Β Cell Factor Pdx1 Is Augmented By Lysine Methylation Catalyzed By The Methyltransferase Set7/9, Aarthi V. Maganti, Bernhard Maier, Sarah A. Tersey, Megan L. Sampley, Amber L. Mosley, Sabire Özcan, Boobalan Pachaiyappan, Patrick M. Woster, Chad S. Hunter, Roland Stein, Raghavendra G. Mirmira

Molecular and Cellular Biochemistry Faculty Publications

The transcription factor Pdx1 is crucial to islet β cell function and regulates target genes in part through interaction with coregulatory factors. Set7/9 is a Lys methyltransferase that interacts with Pdx1. Here we tested the hypothesis that Lys methylation of Pdx1 by Set7/9 augments Pdx1 transcriptional activity. Using mass spectrometry and mutational analysis of purified proteins, we found that Set7/9 methylates the N-terminal residues Lys-123 and Lys-131 of Pdx1. Methylation of these residues occurred only in the context of intact, full-length Pdx1, suggesting a specific requirement of secondary and/or tertiary structural elements for catalysis by Set7 ...


Suppression Of Pervasive Noncoding Transcription In Embryonic Stem Cells By Esbaf, Sarah J. Hainer, Weifeng Gu, Benjamin R. Carone, Benjamin D. Landry, Oliver J. Rando, Craig C. Mello, Thomas G. Fazzio Feb 2015

Suppression Of Pervasive Noncoding Transcription In Embryonic Stem Cells By Esbaf, Sarah J. Hainer, Weifeng Gu, Benjamin R. Carone, Benjamin D. Landry, Oliver J. Rando, Craig C. Mello, Thomas G. Fazzio

Molecular, Cell and Cancer Biology Publications

Approximately 75% of the human genome is transcribed, the majority of which does not encode protein. However, many noncoding RNAs (ncRNAs) are rapidly degraded after transcription, and relatively few have established functions, questioning the significance of this observation. Here we show that esBAF, a SWI/SNF family nucleosome remodeling factor, suppresses transcription of ncRNAs from approximately 57,000 nucleosome-depleted regions (NDRs) throughout the genome of mouse embryonic stem cells (ESCs). We show that esBAF functions to both keep NDRs nucleosome-free and promote elevated nucleosome occupancy adjacent to NDRs. Reduction of adjacent nucleosome occupancy upon esBAF depletion is strongly correlated with ...


Functional Analysis Of The Ovarian Cancer Susceptibility Locus At 9p22.2 Reveals A Transcription Regulatory Network Mediated By Bnc2 In Ovarian Cells, Melissa Buckley Jan 2015

Functional Analysis Of The Ovarian Cancer Susceptibility Locus At 9p22.2 Reveals A Transcription Regulatory Network Mediated By Bnc2 In Ovarian Cells, Melissa Buckley

Graduate Theses and Dissertations

GWAS have identified several chromosomal loci associated with ovarian cancer risk. However, the mechanism underlying these associations remains elusive. We identify candidate functional Single Nucleotide Polymorphisms (SNPs) at the 9p22.2 ovarian cancer susceptibility locus, several of which map to transcriptional regulatory elements active in ovarian cells identified by FAIRE-seq (Formaldehyde assisted isolation of regulatory elements followed by sequencing) and ChIP-seq (Chromatin Immunoprecipitation followed by sequencing) in relevant cell types. Reporter and electrophoretic mobility shift assays (EMSA) determined the extent to which candidate SNPs had allele specific effects. Chromosome conformation capture (3C) reveals a physical association between Basonuclin 2 (BNC2 ...


Single Human Cells Use Transcriptional Mechanisms To Compensate For Differences In Cell Size And Dna Content, Olivia Padovan-Merhar Jan 2015

Single Human Cells Use Transcriptional Mechanisms To Compensate For Differences In Cell Size And Dna Content, Olivia Padovan-Merhar

Publicly Accessible Penn Dissertations

Human cells are dynamic: they grow, replicate their genetic information (DNA), and divide. Clonal populations of cells can display marked heterogeneity in size, leading to significant variability in the ratio of DNA to cellular volume. Despite this variability, cells must maintain a constant concentration of RNA and protein, produced from DNA, to ensure proper functionality. How do larger cells produce more output from the same amount of DNA? How do cells that have replicated their DNA prior to cellular division produce the same output as before? Using RNA fluorescence in situ hybridization (RNA FISH), we visualize and count individual RNA ...


A Distinct Subunit Composition Of Chromatin-Bound Mediator, Eliza Foster Jan 2015

A Distinct Subunit Composition Of Chromatin-Bound Mediator, Eliza Foster

Undergraduate Honors Theses

Mediator is a multi-subunit protein complex that plays an essential role in transcription by integrating DNA-binding transcription factors (TFs) bound to the enhancer and general transcription factors (GTFs) bound to the promoter of a gene. Mediator is similar to other general transcription factors in that it associates with the promoter region of DNA and interacts with RNA polymerase II (RNAPII), but its large size and many subunits allows it to interact with both GTFs at the promoter and other TFs bound to enhancer DNA. Mediator may regulate transcription by interacting with chromatin. Mediator can exist in different structural states and ...


Transcription Of The Streptococcus Pyogenes Hyaluronic Acid Capsule Biosynthesis Operon Is Regulated By Previously Unknown Upstream Elements, Marina Falaleeva, Oliwia W. Zurek, Robert L. Watkins, Robert W. Reed, Hadeel Ali, Paul Sumby, Jovanka M. Voyich, Natalia Korotkova Dec 2014

Transcription Of The Streptococcus Pyogenes Hyaluronic Acid Capsule Biosynthesis Operon Is Regulated By Previously Unknown Upstream Elements, Marina Falaleeva, Oliwia W. Zurek, Robert L. Watkins, Robert W. Reed, Hadeel Ali, Paul Sumby, Jovanka M. Voyich, Natalia Korotkova

Molecular and Cellular Biochemistry Faculty Publications

The important human pathogen Streptococcus pyogenes (group A Streptococcus [GAS]) produces a hyaluronic acid (HA) capsule that plays critical roles in immune evasion. Previous studies showed that the hasABC operon encoding the capsule biosynthesis enzymes is under the control of a single promoter, P1, which is negatively regulated by the two-component regulatory system CovR/S. In this work, we characterize the sequence upstream of P1 and identify a novel regulatory region controlling transcription of the capsule biosynthesis operon in the M1 serotype strain MGAS2221. This region consists of a promoter, P2, which initiates transcription of a novel small RNA, HasS ...