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

In Situ Structure Of Rotavirus Vp1 Rna-Dependent Rna Polymerase, Simon Jenni, Eric N. Salgado, Tobias Herrmann, Zongli Li, Timothy Grant, Nikolaus Grigorieff, Stefano Trapani, Leandro F. Estrozi, Stephen C. Harrison Jun 2019

In Situ Structure Of Rotavirus Vp1 Rna-Dependent Rna Polymerase, Simon Jenni, Eric N. Salgado, Tobias Herrmann, Zongli Li, Timothy Grant, Nikolaus Grigorieff, Stefano Trapani, Leandro F. Estrozi, Stephen C. Harrison

Open Access Articles

Rotaviruses, like other non-enveloped, double-strand RNA viruses, package an RNA-dependent RNA polymerase (RdRp) with each duplex of their segmented genomes. Rotavirus cell entry results in loss of an outer protein layer and delivery into the cytosol of an intact, inner capsid particle (the "double-layer particle," or DLP). The RdRp, designated VP1, is active inside the DLP; each VP1 achieves many rounds of mRNA transcription from its associated genome segment. Previous work has shown that one VP1 molecule lies close to each 5-fold axis of the icosahedrally symmetric DLP, just beneath the inner surface of its protein shell, embedded in tightly ...


Extracellular-Signal Regulated Kinase: A Central Molecule Driving Epithelial-Mesenchymal Transition In Cancer, Monserrat Olea-Flores, Miriam Daniela Zuniga-Eulogio, Miguel Angel Mendoza-Catalan, Hugo Alberto Rodriguez-Ruiz, Eduardo Castaneda-Saucedo, Carlos Ortuno-Pineda, Teresita Padilla-Benavides, Napoleon Navarro-Tito Jun 2019

Extracellular-Signal Regulated Kinase: A Central Molecule Driving Epithelial-Mesenchymal Transition In Cancer, Monserrat Olea-Flores, Miriam Daniela Zuniga-Eulogio, Miguel Angel Mendoza-Catalan, Hugo Alberto Rodriguez-Ruiz, Eduardo Castaneda-Saucedo, Carlos Ortuno-Pineda, Teresita Padilla-Benavides, Napoleon Navarro-Tito

Open Access Articles

Epithelial-mesenchymal transition (EMT) is a reversible cellular process, characterized by changes in gene expression and activation of proteins, favoring the trans-differentiation of the epithelial phenotype to a mesenchymal phenotype. This process increases cell migration and invasion of tumor cells, progression of the cell cycle, and resistance to apoptosis and chemotherapy, all of which support tumor progression. One of the signaling pathways involved in tumor progression is the MAPK pathway. Within this family, the ERK subfamily of proteins is known for its contributions to EMT. The ERK subfamily is divided into typical (ERK 1/2/5), and atypical (ERK 3/4 ...


An Order-To-Disorder Structural Switch Activates The Foxm1 Transcription Factor, Aimee H. Marceau, Caileen M. Brison, Santrupti Nerli, Heather E. Arsenault, Andrew C. Mcshan, Eefei Chen, Hsiau-Wei Lee, Jennifer A. Benanti, Nikolaos G. Sgourakis, Seth M. Rubin May 2019

An Order-To-Disorder Structural Switch Activates The Foxm1 Transcription Factor, Aimee H. Marceau, Caileen M. Brison, Santrupti Nerli, Heather E. Arsenault, Andrew C. Mcshan, Eefei Chen, Hsiau-Wei Lee, Jennifer A. Benanti, Nikolaos G. Sgourakis, Seth M. Rubin

Open Access Articles

Intrinsically disordered transcription factor transactivation domains (TADs) function through structural plasticity, adopting ordered conformations when bound to transcriptional co-regulators. Many transcription factors contain a negative regulatory domain (NRD) that suppresses recruitment of transcriptional machinery through autoregulation of the TAD. We report the solution structure of an autoinhibited NRD-TAD complex within FoxM1, a critical activator of mitotic gene expression. We observe that while both the FoxM1 NRD and TAD are primarily intrinsically disordered domains, they associate and adopt a structured conformation. We identify how Plk1 and Cdk kinases cooperate to phosphorylate FoxM1, which releases the TAD into a disordered conformation that ...


Serum Deprivation Of Mesenchymal Stem Cells Improves Exosome Activity And Alters Lipid And Protein Composition, Reka A. Haraszti, Rachael Miller, Michelle L. Dubuke, Andrew H. Coles, Marie C. Didiot, Dimas Echeverria, Matteo Stoppato, Yves Y. Sere, John D. Leszyk, Julia F. Alterman, Bruno M. D. C. Godinho, Matthew R. Hassler, Rachel Wollacott, Yan Wang, Scott A. Shaffer, Neil Aronin, Anastasia Khvorova May 2019

Serum Deprivation Of Mesenchymal Stem Cells Improves Exosome Activity And Alters Lipid And Protein Composition, Reka A. Haraszti, Rachael Miller, Michelle L. Dubuke, Andrew H. Coles, Marie C. Didiot, Dimas Echeverria, Matteo Stoppato, Yves Y. Sere, John D. Leszyk, Julia F. Alterman, Bruno M. D. C. Godinho, Matthew R. Hassler, Rachel Wollacott, Yan Wang, Scott A. Shaffer, Neil Aronin, Anastasia Khvorova

Open Access Articles

Exosomes can serve as delivery vehicles for advanced therapeutics. The components necessary and sufficient to support exosomal delivery have not been established. Here we connect biochemical composition and activity of exosomes to optimize exosome-mediated delivery of small interfering RNAs (siRNAs). This information is used to create effective artificial exosomes. We show that serum-deprived mesenchymal stem cells produce exosomes up to 22-fold more effective at delivering siRNAs to neurons than exosomes derived from control cells. Proteinase treatment of exosomes stops siRNA transfer, indicating that surface proteins on exosomes are involved in trafficking. Proteomic and lipidomic analyses show that exosomes derived in ...


Brown Fat Organogenesis And Maintenance Requires Akt1 And Akt2, Joan Sanchez-Gurmaches, Camila Martinez Calejman, Su Myung Jung, Huawei Li, David A. Guertin May 2019

Brown Fat Organogenesis And Maintenance Requires Akt1 And Akt2, Joan Sanchez-Gurmaches, Camila Martinez Calejman, Su Myung Jung, Huawei Li, David A. Guertin

Open Access Articles

OBJECTIVE: Understanding the signaling mechanisms that control brown adipose tissue (BAT) development is relevant to understanding energy homeostasis and obesity. The AKT kinases are insulin effectors with critical in vivo functions in adipocytes; however, their role in adipocyte development remains poorly understood. The goal of this study was to investigate AKT function in BAT development.

METHODS: We conditionally deleted Akt1 and Akt2 either individually or together with Myf5-Cre, which targets early mesenchymal precursors that give rise to brown adipocytes. Because Myf5-Cre also targets skeletal muscle and some white adipocyte lineages, comparisons were made between AKT function in BAT versus white ...


Regulation Of The Drosophila Imd Pathway By Signaling Amyloids, Anni Kleino, Neal S. Silverman May 2019

Regulation Of The Drosophila Imd Pathway By Signaling Amyloids, Anni Kleino, Neal S. Silverman

Open Access Articles

Fruit flies elicit effective defense responses against numerous microbes. The responses against Gram-negative bacteria are mediated by the Imd pathway, an evolutionarily conserved NF-kappaB pathway recognizing meso-diaminopimelic acid (DAP)-type peptidoglycan from bacterial cell walls. Several reviews already provide a detailed view of ligand recognition and signal transduction during Imd signaling, but the formation and regulation of the signaling complex immediately downstream of the peptidoglycan-sensing receptors is still elusive. In this review, we focus on the formation of the Imd amyloidal signaling center and post-translational modifications in the assembly and disassembly of the Imd signaling complex.


Ouabain Enhances Cell-Cell Adhesion Mediated By Beta1 Subunits Of The Na(+),K(+)-Atpase In Cho Fibroblasts, Claudia Andrea Vilchis-Nestor, Maria Luisa Roldan, Angelina Leonardi, Juan G. Navea, Teresita Padilla-Benavides, Liora Shoshani Apr 2019

Ouabain Enhances Cell-Cell Adhesion Mediated By Beta1 Subunits Of The Na(+),K(+)-Atpase In Cho Fibroblasts, Claudia Andrea Vilchis-Nestor, Maria Luisa Roldan, Angelina Leonardi, Juan G. Navea, Teresita Padilla-Benavides, Liora Shoshani

Open Access Articles

Adhesion is a crucial characteristic of epithelial cells to form barriers to pathogens and toxic substances from the environment. Epithelial cells attach to each other using intercellular junctions on the lateral membrane, including tight and adherent junctions, as well as the Na(+),K(+)-ATPase. Our group has shown that non-adherent chinese hamster ovary (CHO) cells transfected with the canine beta1 subunit become adhesive, and those homotypic interactions amongst beta1 subunits of the Na(+),K(+)-ATPase occur between neighboring epithelial cells. Ouabain, a cardiotonic steroid, binds to the alpha subunit of the Na(+),K(+)-ATPase, inhibits the pump activity and induces ...


Exercise Rescues Gene Pathways Involved In Vascular Expansion And Promotes Functional Angiogenesis In Subcutaneous White Adipose Tissue, So Yun Min, Heather Learnard, Shashi Kant, Olga Gaelikman, Raziel Rojas-Rodriguez, Tiffany Desouza, Anand Desai, John F. Keaney Jr., Silvia Corvera, Siobhan M. Craige Apr 2019

Exercise Rescues Gene Pathways Involved In Vascular Expansion And Promotes Functional Angiogenesis In Subcutaneous White Adipose Tissue, So Yun Min, Heather Learnard, Shashi Kant, Olga Gaelikman, Raziel Rojas-Rodriguez, Tiffany Desouza, Anand Desai, John F. Keaney Jr., Silvia Corvera, Siobhan M. Craige

Open Access Articles

Exercise mitigates chronic diseases such as diabetes, cardiovascular diseases, and obesity; however, the molecular mechanisms governing protection from these diseases are not completely understood. Here we demonstrate that exercise rescues metabolically compromised high fat diet (HFD) fed mice, and reprograms subcutaneous white adipose tissue (scWAT). Using transcriptomic profiling, scWAT was analyzed for HFD gene expression changes that were rescued by exercise. Gene networks involved in vascularization were identified as prominent targets of exercise, which led us to investigate the vasculature architecture and endothelial phenotype. Vascular density in scWAT was found to be compromised in HFD, and exercise rescued this defect ...


The Nua4 Acetyltransferase And Histone H4 Acetylation Promote Replication Recovery After Topoisomerase I-Poisoning, Chiaki Noguchi, Tanu Singh, Melissa A. Ziegler, Jasmine D. Peake, Lyne Khair, Ana Aza, Toru M. Nakamura, Eishi Noguchi Apr 2019

The Nua4 Acetyltransferase And Histone H4 Acetylation Promote Replication Recovery After Topoisomerase I-Poisoning, Chiaki Noguchi, Tanu Singh, Melissa A. Ziegler, Jasmine D. Peake, Lyne Khair, Ana Aza, Toru M. Nakamura, Eishi Noguchi

Open Access Articles

BACKGROUND: Histone acetylation plays an important role in DNA replication and repair because replicating chromatin is subject to dynamic changes in its structures. However, its precise mechanism remains elusive. In this report, we describe roles of the NuA4 acetyltransferase and histone H4 acetylation in replication fork protection in the fission yeast Schizosaccharomyces pombe.

RESULTS: Downregulation of NuA4 subunits renders cells highly sensitive to camptothecin, a compound that induces replication fork breakage. Defects in NuA4 function or mutations in histone H4 acetylation sites lead to impaired recovery of collapsed replication forks and elevated levels of Rad52 DNA repair foci, indicating the ...


The Erk Mapk Pathway Is Essential For Skeletal Development And Homeostasis, Jung-Min Kim, Yeon-Suk Yang, Kwang Hwan Park, Hwanhee Oh, Matthew B. Greenblatt, Jae-Hyuck Shim Apr 2019

The Erk Mapk Pathway Is Essential For Skeletal Development And Homeostasis, Jung-Min Kim, Yeon-Suk Yang, Kwang Hwan Park, Hwanhee Oh, Matthew B. Greenblatt, Jae-Hyuck Shim

Open Access Articles

Mitogen-activated protein kinases (MAPKs) are a family of protein kinases that function as key signal transducers of a wide spectrum of extracellular stimuli, including growth factors and pro-inflammatory cytokines. Dysregulation of the extracellular signal-regulated kinase (ERK) MAPK pathway is associated with human skeletal abnormalities including Noonan syndrome, neurofibromatosis type 1, and cardiofaciocutaneous syndrome. Here, we demonstrate that ERK activation in osteoprogenitors is required for bone formation during skeletal development and homeostasis. Deletion of Mek1 and Mek2, kinases upstream of ERK MAPK, in osteoprogenitors (Mek1(Osx)Mek2(-/-)), resulted in severe osteopenia and cleidocranial dysplasia (CCD), similar to that seen in humans ...


The Er-Localized Protein Dfcp1 Modulates Er-Lipid Droplet Contact Formation, Dongfang Li, Yan G. Zhao, Di Li, Hongyu Zhao, Jie Huang, Guangyan Miao, Du Feng, Pingsheng Liu, Dong Li, Hong Zhang Apr 2019

The Er-Localized Protein Dfcp1 Modulates Er-Lipid Droplet Contact Formation, Dongfang Li, Yan G. Zhao, Di Li, Hongyu Zhao, Jie Huang, Guangyan Miao, Du Feng, Pingsheng Liu, Dong Li, Hong Zhang

Open Access Articles

Very little is known about the spatiotemporal generation of lipid droplets (LDs) from the endoplasmic reticulum (ER) and the factors that mediate ER-LD contacts for LD growth. Using super-resolution grazing incidence structured illumination microscopy (GI-SIM) live-cell imaging, we reveal that upon LD induction, the ER-localized protein DFCP1 redistributes to nascent puncta on the ER, whose formation depends on triglyceride synthesis. These structures move along the ER and fuse to form expanding LDs. Fusion and expansion of DFCP1-labeled nascent structures is controlled by BSCL2. BSCL2 depletion causes accumulation of nascent DFCP1 structures. DFCP1 overexpression increases LD size and enhances ER-LD contacts ...


Transfer Rna Genes Affect Chromosome Structure And Function Via Local Effects, Omar Hamdani, Tsung-Han S. Hsieh, Oliver J. Rando, Rohinton T. Kamakaka Apr 2019

Transfer Rna Genes Affect Chromosome Structure And Function Via Local Effects, Omar Hamdani, Tsung-Han S. Hsieh, Oliver J. Rando, Rohinton T. Kamakaka

Open Access Articles

The genome is packaged and organized in an ordered, non-random manner and specific chromatin segments contact nuclear substructures to mediate this organization. Transfer RNA genes (tDNAs) are binding sites for transcription factors and architectural proteins and are thought to play an important role in the organization of the genome. In this study, we investigate the role of tDNAs in genomic organization and chromosome function by editing a chromosome so that it lacks any tDNAs. Surprisingly our analyses of this tDNA-less chromosome show that loss of tDNAs does not grossly affect chromatin architecture or chromosome tethering and mobility. However, loss of ...


Genome-Wide Prediction Of Synthetic Rescue Mediators Of Resistance To Targeted And Immunotherapy, Avinash Das Sahu, Olga Ponomarova, Eytan Ruppin Mar 2019

Genome-Wide Prediction Of Synthetic Rescue Mediators Of Resistance To Targeted And Immunotherapy, Avinash Das Sahu, Olga Ponomarova, Eytan Ruppin

Open Access Articles

Most patients with advanced cancer eventually acquire resistance to targeted therapies, spurring extensive efforts to identify molecular events mediating therapy resistance. Many of these events involve synthetic rescue (SR) interactions, where the reduction in cancer cell viability caused by targeted gene inactivation is rescued by an adaptive alteration of another gene (the rescuer). Here, we perform a genome-wide in silico prediction of SR rescuer genes by analyzing tumor transcriptomics and survival data of 10,000 TCGA cancer patients. Predicted SR interactions are validated in new experimental screens. We show that SR interactions can successfully predict cancer patients' response and emerging ...


Anti-Drug Antibody Responses Impair Prophylaxis Mediated By Aav-Delivered Hiv-1 Broadly Neutralizing Antibodies, Matthew R. Gardner, Ina Fetzer, Lisa M. Kattenhorn, Meredith E. Davis-Gardner, Amber S. Zhou, Barnett Alfant, Jesse A. Weber, Hema R. Kondur, Jose M. Martinez-Navio, Sebastian P. Fuchs, Ronald C. Desrosiers, Guangping Gao, Jeffrey D. Lifson, Michael Farzan Mar 2019

Anti-Drug Antibody Responses Impair Prophylaxis Mediated By Aav-Delivered Hiv-1 Broadly Neutralizing Antibodies, Matthew R. Gardner, Ina Fetzer, Lisa M. Kattenhorn, Meredith E. Davis-Gardner, Amber S. Zhou, Barnett Alfant, Jesse A. Weber, Hema R. Kondur, Jose M. Martinez-Navio, Sebastian P. Fuchs, Ronald C. Desrosiers, Guangping Gao, Jeffrey D. Lifson, Michael Farzan

Open Access Articles

Adeno-associated virus (AAV) delivery of potent and broadly neutralizing antibodies (bNAbs is a promising approach for the prevention of HIV-1 infection. The immunoglobulin G (IgG)1 subtype is usually selected for this application, because it efficiently mediates antibody effector functions and has a somewhat longer half-life. However, the use of IgG1-Fc has been associated with the generation of anti-drug antibodies (ADAs) that correlate with loss of antibody expression. In contrast, we have shown that expression of the antibody-like molecule eCD4-Ig bearing a rhesus IgG2-Fc domain showed reduced immunogenicity and completely protected rhesus macaques from simian-HIV (SHIV)-AD8 challenges. To directly ...


Hypomorphic Mutations Of Trip11 Cause Odontochondrodysplasia, Anika Wehrle, John A. Follit, Gregory J. Pazour, Andrea Superti-Furga, Martin Lowe, Ekkehart Lausch Feb 2019

Hypomorphic Mutations Of Trip11 Cause Odontochondrodysplasia, Anika Wehrle, John A. Follit, Gregory J. Pazour, Andrea Superti-Furga, Martin Lowe, Ekkehart Lausch

Open Access Articles

Odontochondrodysplasia (ODCD) is an unresolved genetic disorder of skeletal and dental development. Here, we show that ODCD is caused by hypomorphic TRIP11 mutations, and we identify ODCD as the nonlethal counterpart to achondrogenesis 1A (ACG1A), the known null phenotype in humans. TRIP11 encodes Golgi-associated microtubule-binding protein 210 (GMAP-210), an essential tether protein of the Golgi apparatus that physically interacts with intraflagellar transport 20 (IFT20), a component of the ciliary intraflagellar transport complex B. This association and extraskeletal disease manifestations in ODCD point to a cilium-dependent pathogenesis. However, our functional studies in patient-derived primary cells clearly support a Golgi-based disease mechanism ...


Bridging From Intramuscular To Limb Perfusion Delivery Of Raav: Optimization In A Non-Human Primate Study, Alisha Gruntman, Gwladys Gernoux, Qiushi Tang, Guo-Jie Ye, Dave R. Knop, Gensheng Wang, Janet Benson, Kristen E. Coleman, Allison M. Keeler, Christian Mueller, Louis G. Chicoine, Jeffrey D. Chulay, Terence R. Flotte Feb 2019

Bridging From Intramuscular To Limb Perfusion Delivery Of Raav: Optimization In A Non-Human Primate Study, Alisha Gruntman, Gwladys Gernoux, Qiushi Tang, Guo-Jie Ye, Dave R. Knop, Gensheng Wang, Janet Benson, Kristen E. Coleman, Allison M. Keeler, Christian Mueller, Louis G. Chicoine, Jeffrey D. Chulay, Terence R. Flotte

Open Access Articles

Phase 1 and phase 2 gene therapy trials using intramuscular (IM) administration of a recombinant adeno-associated virus serotype 1 (rAAV1) for replacement of serum alpha-1 antitrypsin (AAT) deficiency have shown long-term (5-year) stable transgene expression at approximately 2% to 3% of therapeutic levels, arguing for the long-term viability of this approach to gene replacement of secreted serum protein deficiencies. However, achieving these levels required 100 IM injections to deliver 135 mL of vector, and further dose escalation is limited by the scalability of direct IM injection. To further advance the dose escalation, we sought to bridge the rAAV-AAT clinical development ...


An Automated Bayesian Pipeline For Rapid Analysis Of Single-Molecule Binding Data, Carlas Smith, Karina Jouravleva, Maximiliaan Huisman, Samson M. Jolly, Phillip D. Zamore, David Grünwald Jan 2019

An Automated Bayesian Pipeline For Rapid Analysis Of Single-Molecule Binding Data, Carlas Smith, Karina Jouravleva, Maximiliaan Huisman, Samson M. Jolly, Phillip D. Zamore, David Grünwald

Open Access Articles

Single-molecule binding assays enable the study of how molecular machines assemble and function. Current algorithms can identify and locate individual molecules, but require tedious manual validation of each spot. Moreover, no solution for high-throughput analysis of single-molecule binding data exists. Here, we describe an automated pipeline to analyze single-molecule data over a wide range of experimental conditions. In addition, our method enables state estimation on multivariate Gaussian signals. We validate our approach using simulated data, and benchmark the pipeline by measuring the binding properties of the well-studied, DNA-guided DNA endonuclease, TtAgo, an Argonaute protein from the Eubacterium Thermus thermophilus. We ...


Ninjurin1 Positively Regulates Osteoclast Development By Enhancing The Survival Of Prefusion Osteoclasts, Sung-Jin Bae, Min Wook Shin, Taekwon Son, Hye Shin. Lee, Ji Soo Chae, Sejin Jeon, Goo Taeg Oh, Kyu-Won Kim Jan 2019

Ninjurin1 Positively Regulates Osteoclast Development By Enhancing The Survival Of Prefusion Osteoclasts, Sung-Jin Bae, Min Wook Shin, Taekwon Son, Hye Shin. Lee, Ji Soo Chae, Sejin Jeon, Goo Taeg Oh, Kyu-Won Kim

Open Access Articles

Osteoclasts (OCs) are bone-resorbing cells that originate from hematopoietic stem cells and develop through the fusion of mononuclear myeloid precursors. Dysregulation of OC development causes bone disorders such as osteopetrosis, osteoporosis, and rheumatoid arthritis. Although the molecular mechanisms underlying osteoclastogenesis have been well established, the means by which OCs maintain their survival during OC development remain unknown. We found that Ninjurin1 (Ninj1) expression is dynamically regulated during osteoclastogenesis and that Ninj1(-/-) mice exhibit increased trabecular bone volume owing to impaired OC development. Ninj1 deficiency did not alter OC differentiation, transmigration, fusion, or actin ring formation but increased Caspase-9-dependent intrinsic apoptosis ...


A Persistence Detector For Metabolic Network Rewiring In An Animal, Jote T. Bulcha, Gabrielle E. Giese, Zulfikar Ali, Yong-Uk Lee, Melissa D. Walker, Amy D. Holdorf, L. Safak Yilmaz, Robert C. Brewster, Albertha J. M. Walhout Jan 2019

A Persistence Detector For Metabolic Network Rewiring In An Animal, Jote T. Bulcha, Gabrielle E. Giese, Zulfikar Ali, Yong-Uk Lee, Melissa D. Walker, Amy D. Holdorf, L. Safak Yilmaz, Robert C. Brewster, Albertha J. M. Walhout

Open Access Articles

Biological systems must possess mechanisms that prevent inappropriate responses to spurious environmental inputs. Caenorhabditis elegans has two breakdown pathways for the short-chain fatty acid propionate: a canonical, vitamin B12-dependent pathway and a propionate shunt that is used when vitamin B12 levels are low. The shunt pathway is kept off when there is sufficient flux through the canonical pathway, likely to avoid generating shunt-specific toxic intermediates. Here, we discovered a transcriptional regulatory circuit that activates shunt gene expression upon propionate buildup. Nuclear hormone receptor 10 (NHR-10) and NHR-68 function together as a "persistence detector" in a type 1, coherent feed-forward loop ...


The Non-Canonical Smc Protein Smchd1 Antagonises Tad Formation And Compartmentalisation On The Inactive X Chromosome, Michal R. Gdula, Tatyana B. Nesterova, Greta Pintacuda, Jonathan Godwin, Ye Zhan, Hakan Ozadam, Michael Mcclellan, Daniella Moralli, Felix Krueger, Catherine M. Green, Wolf Reik, Skirmantas Kriaucionis, Edith Heard, Job Dekker, Neil Brockdorff Jan 2019

The Non-Canonical Smc Protein Smchd1 Antagonises Tad Formation And Compartmentalisation On The Inactive X Chromosome, Michal R. Gdula, Tatyana B. Nesterova, Greta Pintacuda, Jonathan Godwin, Ye Zhan, Hakan Ozadam, Michael Mcclellan, Daniella Moralli, Felix Krueger, Catherine M. Green, Wolf Reik, Skirmantas Kriaucionis, Edith Heard, Job Dekker, Neil Brockdorff

Open Access Articles

The inactive X chromosome (Xi) in female mammals adopts an atypical higher-order chromatin structure, manifested as a global loss of local topologically associated domains (TADs), A/B compartments and formation of two mega-domains. Here we demonstrate that the non-canonical SMC family protein, SmcHD1, which is important for gene silencing on Xi, contributes to this unique chromosome architecture. Specifically, allelic mapping of the transcriptome and epigenome in SmcHD1 mutant cells reveals the appearance of sub-megabase domains defined by gene activation, CpG hypermethylation and depletion of Polycomb-mediated H3K27me3. These domains, which correlate with sites of SmcHD1 enrichment on Xi in wild-type cells ...


Hla-Do Modulates The Diversity Of The Mhc-Ii Self-Peptidome, Padma P. Nanaware, Mollie M. Jurewicz, John D. Leszyk, Scott A. Shaffer, Lawrence J. Stern Dec 2018

Hla-Do Modulates The Diversity Of The Mhc-Ii Self-Peptidome, Padma P. Nanaware, Mollie M. Jurewicz, John D. Leszyk, Scott A. Shaffer, Lawrence J. Stern

Open Access Articles

Presentation of antigenic peptides on MHC-II molecules is essential for tolerance to self and for initiation of immune responses against foreign antigens. DO (HLA-DO in humans, H2-O in mice) is a non-classical MHC-II protein that has been implicated in control of autoimmunity and regulation of neutralizing antibody responses to viruses. These effects likely are related to a role of DO in selecting MHC-II epitopes, but previous studies examining the effect of DO on presentation of selected CD4 T cell epitopes have been contradictory. To understand how DO modulates MHC-II antigen presentation, we characterized the full spectrum of peptides presented by ...


Membrane Protein Nanoparticles: The Shape Of Things To Come, Kailene S. Simon, Naomi L. Pollock, Sarah C. Lee Dec 2018

Membrane Protein Nanoparticles: The Shape Of Things To Come, Kailene S. Simon, Naomi L. Pollock, Sarah C. Lee

Open Access Articles

The use of styrene-maleic acid (SMA) for the purification of a wide range of membrane proteins (MPs) from both prokaryotic and eukaryotic sources has begun to make an impact in the field of MP biology. This method is growing in popularity as a means to purify and thoroughly investigate the structure and function of MPs and biological membranes. The amphiphilic SMA copolymer can effectively extract MPs directly from a native lipid bilayer to form discs approximately 10 nm in diameter. The resulting lipid particles, or styrene-maleic acid lipid particles (SMALPs), contain SMA, protein and membrane lipid. MPs purified in SMALPs ...


Orbit: A New Paradigm For Genetic Engineering Of Mycobacterial Chromosomes, Kenan C. Murphy, Samantha J. Nelson, Subhalaxmi Nambi, Kadamba Papavinasasundaram, Christina E. Baer, Christopher M. Sassetti Dec 2018

Orbit: A New Paradigm For Genetic Engineering Of Mycobacterial Chromosomes, Kenan C. Murphy, Samantha J. Nelson, Subhalaxmi Nambi, Kadamba Papavinasasundaram, Christina E. Baer, Christopher M. Sassetti

Open Access Articles

Two efficient recombination systems were combined to produce a versatile method for chromosomal engineering that obviates the need to prepare double-stranded DNA (dsDNA) recombination substrates. A synthetic "targeting oligonucleotide" is incorporated into the chromosome via homologous recombination mediated by the phage Che9c RecT annealase. This oligonucleotide contains a site-specific recombination site for the directional Bxb1 integrase (Int), which allows the simultaneous integration of a "payload plasmid" that contains a cognate recombination site and a selectable marker. The targeting oligonucleotide and payload plasmid are cotransformed into a RecT- and Int-expressing strain, and drug-resistant homologous recombinants are selected in a single step ...


General Decapping Activators Target Different Subsets Of Inefficiently Translated Mrnas, Feng He, Alper Celik, Chan Wu, Allan Jacobson Dec 2018

General Decapping Activators Target Different Subsets Of Inefficiently Translated Mrnas, Feng He, Alper Celik, Chan Wu, Allan Jacobson

Open Access Articles

The Dcp1-Dcp2 decapping enzyme and the decapping activators Pat1, Dhh1, and Lsm1 regulate mRNA decapping, but their mechanistic integration is unknown. We analyzed the gene expression consequences of deleting PAT1, LSM1, or DHH1, or the DCP2 C-terminal domain, and found that: i) the Dcp2 C-terminal domain is an effector of both negative and positive regulation; ii) rather than being global activators of decapping, Pat1, Lsm1, and Dhh1 directly target specific subsets of yeast mRNAs and loss of the functions of each of these factors has substantial indirect consequences for genome-wide mRNA expression; and iii) transcripts targeted by Pat1, Lsm1, and ...


Potent Cas9 Inhibition In Bacterial And Human Cells By Acriic4 And Acriic5 Anti-Crispr Proteins, Jooyoung Lee, Aamir Mir, Alireza Edraki, Bianca Garcia, Nadia Amrani, Hannah E. Lou, Ildar Gainetdinov, April Pawluk, Raed Ibraheim, Xin D. Gao, Pengpeng Liu, Alan R. Davidson, Karen L. Maxwell, Erik J. Sontheimer Dec 2018

Potent Cas9 Inhibition In Bacterial And Human Cells By Acriic4 And Acriic5 Anti-Crispr Proteins, Jooyoung Lee, Aamir Mir, Alireza Edraki, Bianca Garcia, Nadia Amrani, Hannah E. Lou, Ildar Gainetdinov, April Pawluk, Raed Ibraheim, Xin D. Gao, Pengpeng Liu, Alan R. Davidson, Karen L. Maxwell, Erik J. Sontheimer

Open Access Articles

In their natural settings, CRISPR-Cas systems play crucial roles in bacterial and archaeal adaptive immunity to protect against phages and other mobile genetic elements, and they are also widely used as genome engineering technologies. Previously we discovered bacteriophage-encoded Cas9-specific anti-CRISPR (Acr) proteins that serve as countermeasures against host bacterial immunity by inactivating their CRISPR-Cas systems (A. Pawluk, N. Amrani, Y. Zhang, B. Garcia, et al., Cell 167:1829-1838.e9, 2016, https://doi.org/10.1016/j.cell.2016.11.017). We hypothesized that the evolutionary advantages conferred by anti-CRISPRs would drive the widespread occurrence of these proteins in nature (K ...


Deciphering The Molecular Basis Of Mycobacteria And Lipoglycan Recognition By The C-Type Lectin Dectin-2, Alexiane Decout, Devinder Kaur, Jerome Nigou Nov 2018

Deciphering The Molecular Basis Of Mycobacteria And Lipoglycan Recognition By The C-Type Lectin Dectin-2, Alexiane Decout, Devinder Kaur, Jerome Nigou

Open Access Articles

Dectin-2 is a C-type lectin involved in the recognition of several pathogens such as Aspergillus fumigatus, Candida albicans, Schistosoma mansonii, and Mycobacterium tuberculosis that triggers Th17 immune responses. Identifying pathogen ligands and understanding the molecular basis of their recognition is one of the current challenges. Purified M. tuberculosis mannose-capped lipoarabinomannan (ManLAM) was shown to induce signaling via Dectin-2, an activity that requires the (alpha1 --> 2)-linked mannosides forming the caps. Here, using isogenic M. tuberculosis mutant strains, we demonstrate that ManLAM is a bona fide and actually the sole ligand mediating bacilli recognition by Dectin-2, although M. tuberculosis produces a ...


Signaling Pathways Induced By Leptin During Epithelial(-)Mesenchymal Transition In Breast Cancer, Monserrat Olea-Flores, Juan Carlos Juarez-Cruz, Miguel A. Mendoza-Catalan, Teresita Padilla-Benavides, Napoleon Navarro-Tito Nov 2018

Signaling Pathways Induced By Leptin During Epithelial(-)Mesenchymal Transition In Breast Cancer, Monserrat Olea-Flores, Juan Carlos Juarez-Cruz, Miguel A. Mendoza-Catalan, Teresita Padilla-Benavides, Napoleon Navarro-Tito

Open Access Articles

Leptin is an adipokine that is overexpressed in obese and overweight people. Interestingly, women with breast cancer present high levels of leptin and of its receptor ObR. Leptin plays an important role in breast cancer progression due to the biological processes it participates in, such as epithelial(-)mesenchymal transition (EMT). EMT consists of a series of orchestrated events in which cell(-)cell and cell(-)extracellular matrix interactions are altered and lead to the release of epithelial cells from the surrounding tissue. The cytoskeleton is also re-arranged, allowing the three-dimensional movement of epithelial cells into the extracellular matrix. This transition provides ...


Rare Gene Fusion Rearrangement Sptnb1-Pdgfrb In An Atypical Myeloproliferative Neoplasm, Vanessa Fiorini Furtado, Neeraj Y. Saini, William V. Walsh, Venu G. Bathini, Patricia M. Miron Oct 2018

Rare Gene Fusion Rearrangement Sptnb1-Pdgfrb In An Atypical Myeloproliferative Neoplasm, Vanessa Fiorini Furtado, Neeraj Y. Saini, William V. Walsh, Venu G. Bathini, Patricia M. Miron

Open Access Articles

The 2016 revision to the World Health Organization classification of myeloid neoplasms and acute leukemia recognizes a distinct class of myeloid and lymphoid tumors with eosinophilia-related proliferations associated with specific gene rearrangements, one of which involves rearrangements of platelet-derived growth factor receptor B (PDGFRB) gene. We report a case of a rare PDGFRB rearrangement with SPTNB1 (spectrin beta, nonerythrocytic 1) that presented as atypical myeloproliferative neoplasm.


Adeno-Associated Virus Neutralizing Antibodies In Large Animals And Their Impact On Brain Intraparenchymal Gene Transfer, Dan Wang, Li Zhong, Mengxin Li, Jia Li, Karen Tran, Lingzhi Ren, Ran He, Jun Xie, Richard P. Moser, Cara Fraser, Tim Kuchel, Miguel Sena-Esteves, Terence R. Flotte, Neil Aronin, Guangping Gao Oct 2018

Adeno-Associated Virus Neutralizing Antibodies In Large Animals And Their Impact On Brain Intraparenchymal Gene Transfer, Dan Wang, Li Zhong, Mengxin Li, Jia Li, Karen Tran, Lingzhi Ren, Ran He, Jun Xie, Richard P. Moser, Cara Fraser, Tim Kuchel, Miguel Sena-Esteves, Terence R. Flotte, Neil Aronin, Guangping Gao

Open Access Articles

Pre-existing neutralizing antibody (NAb) against adeno-associated virus (AAV) commonly found in primates is a major host barrier that can severely compromise in vivo gene transfer by AAV vectors. To achieve proof-of-concept success in clinical development of recombinant AAV (rAAV)-based in vivo gene therapy, it is crucial to consider the potential interference of NAb and to enroll serologically compatible study subjects. In this study, we report a large AAV NAb dataset comprising multiple large animal species and AAV serotypes and compare two NAb assays based on in vitro or in vivo transduction inhibition, respectively. Together with previously published AAV seroepidemiology ...


Differential Expression Of Zinc Transporters Accompanies The Differentiation Of C2c12 Myoblasts, Amanda L. Paskavitz, Julia Quintana, Daniella Cangussu, Cristina Tavera-Montanez, Yao Xiao, Sonia Ortiz-Mirnada, Juan G. Navea, Teresita Padilla-Benavides Sep 2018

Differential Expression Of Zinc Transporters Accompanies The Differentiation Of C2c12 Myoblasts, Amanda L. Paskavitz, Julia Quintana, Daniella Cangussu, Cristina Tavera-Montanez, Yao Xiao, Sonia Ortiz-Mirnada, Juan G. Navea, Teresita Padilla-Benavides

Open Access Articles

Zinc transporters facilitate metal mobilization and compartmentalization, playing a key role in cellular development. Little is known about the mechanisms and pathways of Zn movement between Zn transporters and metalloproteins during myoblast differentiation. We analyzed the differential expression of ZIP and ZnT transporters during C2C12 myoblast differentiation. Zn transporters account for a transient decrease of intracellular Zn upon myogenesis induction followed by a gradual increase of Zn in myotubes. Considering the subcellular localization and function of each of the Zn transporters, our findings indicate that a fine regulation is necessary to maintain correct metal concentrations in the cytosol and subcellular ...