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Superresolution Imaging Identifies That Conventional Trafficking Pathways Are Not Essential For Endoplasmic Reticulum To Outer Mitochondrial Membrane Protein Transport., Kyle Salka, Shivaprasad Bhuvanendran, Kassandra Wilson, Petros Bozidis, Mansi Mehta, Kristin Rainey, Hiromi Sesaki, George H Patterson, Jyoti K. Jaiswal, Anamaris M. Colberg-Poley 2017 George Washington University

Superresolution Imaging Identifies That Conventional Trafficking Pathways Are Not Essential For Endoplasmic Reticulum To Outer Mitochondrial Membrane Protein Transport., Kyle Salka, Shivaprasad Bhuvanendran, Kassandra Wilson, Petros Bozidis, Mansi Mehta, Kristin Rainey, Hiromi Sesaki, George H Patterson, Jyoti K. Jaiswal, Anamaris M. Colberg-Poley

Integrative Systems Biology Faculty Publications

Most nuclear-encoded mitochondrial proteins traffic from the cytosol to mitochondria. Some of these proteins localize at mitochondria-associated membranes (MAM), where mitochondria are closely apposed with the endoplasmic reticulum (ER). We have previously shown that the human cytomegalovirus signal-anchored protein known as viral mitochondria-localized inhibitor of apoptosis (vMIA) traffics from the ER to mitochondria and clusters at the outer mitochondrial membrane (OMM). Here, we have examined the host pathways by which vMIA traffics from the ER to mitochondria and clusters at the OMM. By disruption of phosphofurin acidic cluster sorting protein 2 (PACS-2), mitofusins (Mfn1/2), and dynamin related protein 1 ...


Stem Cell Derived Osteoprogenitors And Their Role In Bone Repair Using Morphogenic Activators, Angelina Miller, Sylvanna Islas, George Washington Brownridge III, Warren Hoeffler 2017 Dominican University of California

Stem Cell Derived Osteoprogenitors And Their Role In Bone Repair Using Morphogenic Activators, Angelina Miller, Sylvanna Islas, George Washington Brownridge Iii, Warren Hoeffler

Scholarly & Creative Works Conference 2017

Bone constantly cycles through a dynamic process of breakdown and remodeling. Osteoblasts are the specialized mesenchymal stem cells that have a major role in bone formation and the remodeling process whereas their counterpart osteoclasts, handle bone resorption. Embryonic stem cells can be partially differentiated into Progenitor cells, and we worked with #18, a candidate for being an osteoprogenitor that has the potential to respond to morphogenic activators. In the case of bone remodeling, TGF-β 2, BMP-2 and an abundance of CA++ have been shown to be potential activators of differentiation into osteoblasts. Eight different trials were conducted with the cells ...


Identification Of Novel Interactors Of Sec6 Via Genetic Suppressor Screen Using A Saccharomyces Cerevisiae Genomic Dna Library, Y. Ellen France PhD, Emily Parrish, Raira Ank, Tristan Peterman, Sarah Mork, Billie Mills, Lindsey Duke, Jyoti Lama 2017 Georgia college and state university

Identification Of Novel Interactors Of Sec6 Via Genetic Suppressor Screen Using A Saccharomyces Cerevisiae Genomic Dna Library, Y. Ellen France Phd, Emily Parrish, Raira Ank, Tristan Peterman, Sarah Mork, Billie Mills, Lindsey Duke, Jyoti Lama

Georgia College Student Research Events

Polarized protein secretion is a fundamental process for all organisms, from yeast to higher eukaryotes. The secretory pathway in eukaryotes includes many steps mediated by hundreds of essential proteins. Our interest lies on Sec6, which is an 88kDa protein subunit of the tethering complex named exocyst, which is known to play diverse roles in recognition, tethering, and SNARE-mediated fusion of secretory vesicles. Previous work on Sec6 done by Munson and Songer (2009) explored phenotypes of two novel SEC6 mutant proteins, whose conserved surface amino acids had been altered. Both sec6-49 and sec6-54 mutant alleles displayed severe temperature sensitive growth and ...


Investigation Of Chitosan-Based Hydrogels As A Cell Delivery Platform For Adipose-Derived Stem/Stromal Cell Transplantation To Promote Angiogenesis In Ischemic Tissues, Jobanpreet Singh Dhillon 2017 The University of Western Ontario

Investigation Of Chitosan-Based Hydrogels As A Cell Delivery Platform For Adipose-Derived Stem/Stromal Cell Transplantation To Promote Angiogenesis In Ischemic Tissues, Jobanpreet Singh Dhillon

Electronic Thesis and Dissertation Repository

Stem cell transplantation is under investigation to stimulate angiogenesis in patients with peripheral artery disease. To develop a cell-delivery platform that enhances cell retention and function post-transplantation, the response of human adipose-derived stem/stromal cells (ASCs) encapsulated within N-methacrylate glycol chitosan (MGC) hydrogels with or without integrin-binding RGD or IKVAV motifs was explored. ASC viability was enhanced in the MGC and MGC-RGD hydrogels relative to the MGC-IKVAV group under hypoxic (2% O­2) culture conditions, with cell spreading and higher metabolic activity noted in MGC-RGD at 14 days. Analysis of angiogenic gene expression revealed similar patterns between all hydrogel ...


Alcohol-Related Changes In The Intestinal Microbiome Influence Neutrophil Infiltration, Inflammation And Steatosis In Early Alcoholic Hepatitis In Mice, Patrick Lowe, Benedek Gyongyosi, Abhishek Satishchandran, Arvin Iracheta-Vellve, Aditya Ambade, Karen Kodys, Donna Catalano, Doyle V. Ward, Gyongyi Szabo 2017 University of Massachusetts Medical School

Alcohol-Related Changes In The Intestinal Microbiome Influence Neutrophil Infiltration, Inflammation And Steatosis In Early Alcoholic Hepatitis In Mice, Patrick Lowe, Benedek Gyongyosi, Abhishek Satishchandran, Arvin Iracheta-Vellve, Aditya Ambade, Karen Kodys, Donna Catalano, Doyle V. Ward, Gyongyi Szabo

University of Massachusetts Medical School Faculty Publications

BACKGROUND: Alcohol-induced intestinal dysbiosis disrupts homeostatic gut-liver axis function and is essential in the development of alcoholic liver disease. Here, we investigate changes in enteric microbiome composition in a model of early alcoholic steatohepatitis and dissect the pathogenic role of intestinal microbes in alcohol-induced liver pathology.

MATERIALS AND METHODS: Wild type mice received a 10-day diet that was either 5% alcohol-containing or an isocaloric control diet plus a single binge. 16S rDNA sequencing defined the bacterial communities in the cecum of alcohol- and pair-fed animals. Some mice were treated with an antibiotic cocktail prior to and throughout alcohol feeding. Liver ...


Sumo-Targeted Ubiquitin Ligase (Stubl) Slx5 Regulates Proteolysis Of Centromeric Histone H3 Variant Cse4 And Prevents Its Mislocalization To Euchromatin, Kentaro Ohkuni, Yoshimitsu Takahashi, Alyona Fulp, Josh Lawrimore, Wei-Chun Au, Nagesh Pasupala, Reuben Levy-Myers, Jack Warren, Alexander Strunnikov, Richard E. Baker, Oliver Kerscher, Kerry Bloom, Munira A. Basrai 2017 National Cancer Institute

Sumo-Targeted Ubiquitin Ligase (Stubl) Slx5 Regulates Proteolysis Of Centromeric Histone H3 Variant Cse4 And Prevents Its Mislocalization To Euchromatin, Kentaro Ohkuni, Yoshimitsu Takahashi, Alyona Fulp, Josh Lawrimore, Wei-Chun Au, Nagesh Pasupala, Reuben Levy-Myers, Jack Warren, Alexander Strunnikov, Richard E. Baker, Oliver Kerscher, Kerry Bloom, Munira A. Basrai

Oliver Kerscher

Centromeric histone H3, CENP-ACse4, is essential for faithful chromosome segregation. Stringent regulation of cellular levels of CENP-ACse4 restricts its localization to centromeres. Mislocalization of CENP-ACse4 is associated with aneuploidy in yeast, flies and tumorigenesis in human cells; thus, defining pathways that regulate CENP-A levels is critical for understanding how mislocalization of CENP-A contributes to aneuploidy in human cancers. Previous work in budding yeast has shown that ubiquitination of overexpressed Cse4 by Psh1, an E3 ligase, partially contributes to proteolysis of Cse4. Here, we provide the first evidence that Cse4 is sumoylated by E3 ligases Siz1 and Siz2 in vivo and ...


Tim18p Is A New Component Of The Tim54p-Tim22p Translocon In The Mitochondrial Inner Membrane, Oliver Kerscher, Naresh B. Sepuri, Robert E. Jensen 2017 College of William and Mary

Tim18p Is A New Component Of The Tim54p-Tim22p Translocon In The Mitochondrial Inner Membrane, Oliver Kerscher, Naresh B. Sepuri, Robert E. Jensen

Oliver Kerscher

The mitochondrial inner membrane contains two separate translocons: one required for the translocation of matrix-targeted proteins (the Tim23p-Tim17p complex) and one for the insertion of polytopic proteins into the mitochondrial inner membrane (the Tim54p-Tim22p complex). To identify new members of the Tim54p-Tim22p complex, we screened for high-copy suppressors of the temperature-sensitivetim54-1 mutant. We identified a new gene,TIM18, that encodes an integral protein of the inner membrane. The following genetic and biochemical observations suggest that the Tim18 protein is part of the Tim54p-Tim22p complex in the inner membrane: multiple copies of TIM18 suppress thetim54-1 growth defect; thetim18::HIS3 disruption is ...


Recognizing Chromosomes In Trouble: Association Of The Spindle Checkpoint Protein Bub3p With Altered Kinetochores And A Unique Defective Centromere, Oliver Kerscher, Luciana B. Crotti, Munira A. Basrai 2017 College of William and Mary

Recognizing Chromosomes In Trouble: Association Of The Spindle Checkpoint Protein Bub3p With Altered Kinetochores And A Unique Defective Centromere, Oliver Kerscher, Luciana B. Crotti, Munira A. Basrai

Oliver Kerscher

Spindle checkpoint proteins monitor the interaction of the spindle apparatus with the kinetochores, halting anaphase even if the microtubule attachment of only a single chromosome is altered. In this study, we show that Bub3p of Saccharomyces cerevisiae, an evolutionarily conserved spindle checkpoint protein, exhibits distinct interactions with an altered or defective kinetochore(s). We show for the first time that green fluorescent protein-tagged S. cerevisiae Bub3p (Bub3-GFP) exhibits not only a diffuse nuclear localization pattern but also forms distinct nuclear foci in unperturbed growing and G2/M-arrested cells. As Bub3-GFP foci overlap only a subset of kinetochores, we tested ...


Sumo-Targeted Ubiquitin Ligase (Stubl) Slx5 Regulates Proteolysis Of Centromeric Histone H3 Variant Cse4 And Prevents Its Mislocalization To Euchromatin, Kentaro Ohkuni, Yoshimitsu Takahashi, Alyona Flup, Josh La, Wei-Chun Au, Nagesh Pasupala, Ruben Levy-Meyers, Jack Warren, Alexander Strunnikov, Richard E. Baker, Oliver Kerscher, Kerry Bloom, Munira A. Basrai 2017 College of William and Mary

Sumo-Targeted Ubiquitin Ligase (Stubl) Slx5 Regulates Proteolysis Of Centromeric Histone H3 Variant Cse4 And Prevents Its Mislocalization To Euchromatin, Kentaro Ohkuni, Yoshimitsu Takahashi, Alyona Flup, Josh La, Wei-Chun Au, Nagesh Pasupala, Ruben Levy-Meyers, Jack Warren, Alexander Strunnikov, Richard E. Baker, Oliver Kerscher, Kerry Bloom, Munira A. Basrai

Oliver Kerscher

Centromeric histone H3, CENP-ACse4, is essential for faithful chromosome segregation. Stringent regulation of cellular levels of CENP-ACse4 restricts its localization to centromeres. Mislocalization of CENP-ACse4 is associated with aneuploidy in yeast and flies and tumorigenesis in human cells; thus defining pathways that regulate CENP-A levels is critical for understanding how mislocalization of CENP-A contributes to aneuploidy in human cancers. Previous work in budding yeast shows that ubiquitination of overexpressed Cse4 by Psh1, an E3 ligase, partially contributes to proteolysis of Cse4. Here we provide the first evidence that Cse4 is sumoylated by E3 ligases Siz1 and Siz2 in vivo and ...


The Yeast Nuclear Pore Complex Functionally Interacts With Components Of The Spindle Assembly Checkpoint, Tatiana Louk, Oliver Kerscher, Robert J. Scott, Munira A. Basrai, Richard W. Wozniak 2017 College of William and Mary

The Yeast Nuclear Pore Complex Functionally Interacts With Components Of The Spindle Assembly Checkpoint, Tatiana Louk, Oliver Kerscher, Robert J. Scott, Munira A. Basrai, Richard W. Wozniak

Oliver Kerscher

Aphysical and functional link between the nuclear pore complex (NPC) and the spindle checkpoint machinery has been established in the yeast Saccharomyces cerevisiae. We show that two proteins required for the execution of the spindle checkpoint, Mad1p and Mad2p, reside predominantly at the NPC throughout the cell cycle. There they are associated with a subcomplex of nucleoporins containing Nup53p, Nup170p, and Nup157p. The association of the Mad1p–Mad2p complex with the NPC requires Mad1p and is mediated in part by Nup53p. On activation of the spindle checkpoint, we detect changes in the interactions between these proteins, including the release of ...


Functional Redundancy Between Trans-Golgi Network Snare Family Members In Arabidopsis Thaliana, Sang-Jin Kim, Diane C. Bassham 2017 Iowa State University

Functional Redundancy Between Trans-Golgi Network Snare Family Members In Arabidopsis Thaliana, Sang-Jin Kim, Diane C. Bassham

Diane Bassham

Background

Vesicle fusion is an essential process for maintaining the structure and function of the endomembrane system. Fusion is mediated by t-SNARE (soluble N-ethylmaleimide-sensitive factor attachment protein receptor) fusion proteins on the target membrane and v-SNAREs on the vesicle membrane; v-and t-SNAREs interact with each other, driving vesicle fusion with the target membrane. The Arabidopsis thaliana trans-Golgi network resident SNAREs SYP41 and VTI12, along with YKT61/62, have been shown to function in vesicle fusion in vitro, consistent with immunoprecipitation results showing their interaction in Arabidopsis cell extracts. Conflicting published results have indicated that SYP4 family members are ...


Guidelines For The Use And Interpretation Of Assays For Monitoring Autophagy In Higher Eukaryotes, Daniel J. Klionsky, Michael J. Lenardo, Diane C. Bassham 2017 University of Michigan-Ann Arbor

Guidelines For The Use And Interpretation Of Assays For Monitoring Autophagy In Higher Eukaryotes, Daniel J. Klionsky, Michael J. Lenardo, Diane C. Bassham

Diane Bassham

Research in autophagy continues to accelerate,1 and as a result many new scientists are entering the field. Accordingly, it is important to establish a standard set of criteria for monitoring macroautophagy in different organisms. Recent reviews have described the range of assays that have been used for this purpose.2,3 There are many useful and convenient methods that can be used to monitor macroautophagy in yeast, but relatively few in other model systems, and there is much confusion regarding acceptable methods to measure macroautophagy in higher eukaryotes. A key point that needs to be emphasized is that there ...


Autophagy In Plants And Algae, Diane C. Bassham, Jose L. Crespo 2017 Iowa State University

Autophagy In Plants And Algae, Diane C. Bassham, Jose L. Crespo

Diane Bassham

Autophagy is a major cellular degradation pathway in which materials are delivered to the vacuole in double-membrane vesicles known as autophagosomes, broken down, and recycled (Li and Vierstra, 2012; Liu and Bassham, 2012). In photosynthetic organisms, the pathway is strongly activated by biotic and abiotic stresses, including nutrient limitation, oxidative, salt and drought stress and pathogen infection, and during senescence (Perez-Perez et al., 2012; Lv et al., 2014). Mutation of genes required for autophagy causes hypersensitivity to stress, indicating that autophagy is important for tolerance of multiple stresses. While autophagy is often non-selective, a growing number of examples of selectivity ...


Guidelines For The Use And Interpretation Of Assays For Monitoring Autophagy, Daniel J. Klionsky, Shoba Amarnath, Vellareddy Anantharam, Diane C. Bassham, Bryan Bellaire 2017 University of Michigan-Ann Arbor

Guidelines For The Use And Interpretation Of Assays For Monitoring Autophagy, Daniel J. Klionsky, Shoba Amarnath, Vellareddy Anantharam, Diane C. Bassham, Bryan Bellaire

Diane Bassham

In 2008 we published the first set of guidelines for standardizing research in autophagy. Since then, research on this topic has continued to accelerate, and many new scientists have entered the field. Our knowledge base and relevant new technologies have also been expanding. Accordingly, it is important to update these guidelines for monitoring autophagy in different organisms. Various reviews have described the range of assays that have been used for this purpose. Nevertheless, there continues to be confusion regarding acceptable methods to measure autophagy, especially in multicellular eukaryotes. A key point that needs to be emphasized is that there is ...


Characterization Of A New Oda3 Allele, Oda3-6, Defective In Assembly Of The Outer Dynein Arm-Docking Complex In Chlamydomonas Reinhardtii, Jason M. Brown, Matthew Mosley, Daniela Montes-Berrueta, Yuqing Hou, Fan Yang, Chasity Scarbrough, George B. Witman, Maureen Wirschell 2017 Salem State University

Characterization Of A New Oda3 Allele, Oda3-6, Defective In Assembly Of The Outer Dynein Arm-Docking Complex In Chlamydomonas Reinhardtii, Jason M. Brown, Matthew Mosley, Daniela Montes-Berrueta, Yuqing Hou, Fan Yang, Chasity Scarbrough, George B. Witman, Maureen Wirschell

Biology Faculty Publications

We have used an insertional mutagenesis approach to generate new C. reinhardtii motility mutants. Of 56 mutants isolated, one is a new allele at the ODA3 locus, called oda3-6. Similar to the previously characterized oda3 alleles, oda3-6 has a slow-jerky swimming phenotype and reduced swimming speed. The oda3-6 mutant fails to assemble the outer dynein arm motor and outer dynein arm—docking complex (ODA-DC) in the ciliary axoneme due to an insertion in the 5’ end of the DCC1 gene, which encodes the DC1 subunit of the ODA-DC. Transformation of oda3-6 with the wild-type DCC1 gene rescues the mutant swimming ...


P32. Altered Sensory Processing In Response To Novel Dreadd-Induced Inactivation Of Gaba In Pedunculopontine Tegmental Nucleus, Niveen Fulcher 2017 Western University

P32. Altered Sensory Processing In Response To Novel Dreadd-Induced Inactivation Of Gaba In Pedunculopontine Tegmental Nucleus, Niveen Fulcher

Western Research Forum

Niveen Fulcher1, Cleusa De Oliveira2, & Susanne Schmid1,2

1Schulich School of Medicine & Dentistry, University of Western Ontario

2Anatomy and Cell Biology, University of Western Ontario

Altered sensory processing in response to novel DREADD-induced inactivation of GABA in pedunculopontine tegmental nucleus

Background: Sensory processing deficits are associated with certain psychiatric illnesses, such as schizophrenia and autism spectrum disorder (ASD). Sensory filtering and sensorimotor gating are evolutionarily conserved preattentive responses that filter and block redundant sensory stimuli that would otherwise overwhelm our brains. To date, underlying mechanisms of these deficits are undefined. Prepulse inhibition (PPI) of the ...


P23. Elucidation Of The Signaling Pathway Of Mertk, Kyle Taruc 2017 Western University

P23. Elucidation Of The Signaling Pathway Of Mertk, Kyle Taruc

Western Research Forum

Background

Mer Tyrosine Kinase (MERTK) is a receptor which mediates efferocytosis – removal of apoptotic cells by phagocytes such as macrophages. MERTK plays a critical role in homeostasis, with mutations in MERTK associated with the development and progression of atherosclerosis, the buildup of cholesterol-laden plaque in the sub-arterial space. Indeed, one MERTK allele is associated with 66% (heterozygous) to 75% (homozygous) protection from atherosclerosis, while other alleles and SNP’s pre-dispose to atherosclerosis and autoimmunity. Complications resulting from atherosclerosis, including heart attack and stroke, are currently the second leading cause of mortality in Canada. Despite the importance of MERTK in atherosclerosis ...


Pulmonary Surfactant Fortified With Cath-2 As A Novel Therapy For Bacterial Pneumonia, Brandon J. Baer 2017 University of Western Ontario

Pulmonary Surfactant Fortified With Cath-2 As A Novel Therapy For Bacterial Pneumonia, Brandon J. Baer

Western Research Forum

Background: Bacterial pneumonia is a leading cause of death worldwide, with high mortality rates persisting even after antibiotic treatment. Current treatments for pneumonia involve administration of antibiotics, however after the bacteria are killed they release toxic substances that induce inflammation and lung dysfunction. Host defense peptides represent a potential solution to this problem through their ability to down regulate inflammation. However, effective delivery to the lung is difficult because of the complex branching structure of the airways. My study addresses this delivery problem by using exogenous surfactant, a pulmonary delivery vehicle capable of improving spreading of these peptides throughout the ...


Antisense Oligonucleotides Used To Target The Dux4 Mrna As Therapeutic Approaches In Faciosscapulohumeral Muscular Dystrophy (Fshd), Eugenie Ansseau, Celine Vanderplanck, Armelle Wauters, Scott Q. Harper, Frederique Coppee, Alexandra Belayew 2017 University of Mons

Antisense Oligonucleotides Used To Target The Dux4 Mrna As Therapeutic Approaches In Faciosscapulohumeral Muscular Dystrophy (Fshd), Eugenie Ansseau, Celine Vanderplanck, Armelle Wauters, Scott Q. Harper, Frederique Coppee, Alexandra Belayew

Wellstone Center for FSHD Publications and Presentations

FacioScapuloHumeral muscular Dystrophy (FSHD) is one of the most prevalent hereditary myopathies and is generally characterized by progressive muscle atrophy affecting the face, scapular fixators; upper arms and distal lower legs. The FSHD locus maps to a macrosatellite D4Z4 repeat array on chromosome 4q35. Each D4Z4 unit contains a DUX4 gene; the most distal of which is flanked by a polyadenylation site on FSHD-permissive alleles, which allows for production of stable DUX4 mRNAs. In addition, an open chromatin structure is required for DUX4 gene transcription. FSHD thus results from a gain of function of the toxic DUX4 protein that normally ...


Evolutionary Adaptations Of Biofilms Infecting Cystic Fibrosis Lungs Promote Mechanical Toughness By Adjusting Polysaccharide Production, Kristin Kovach, Megan Davis-Fields, Yasuhiko Irie, Kanishk Jain, Shashvat Doorwar, Katherine Vuong, Numa Dhamani, Kishore Mohanty, Ahmed Touhami, Vernita D. Gordon 2017 University of Texas at Austin

Evolutionary Adaptations Of Biofilms Infecting Cystic Fibrosis Lungs Promote Mechanical Toughness By Adjusting Polysaccharide Production, Kristin Kovach, Megan Davis-Fields, Yasuhiko Irie, Kanishk Jain, Shashvat Doorwar, Katherine Vuong, Numa Dhamani, Kishore Mohanty, Ahmed Touhami, Vernita D. Gordon

Yasuhiko Irie

Biofilms are communities of microbes embedded in a matrix of extracellular polymeric substances, largely polysaccharides. Multiple types of extracellular polymeric substances can be produced by a single bacterial strain. The distinct polymer components of biofilms are known to provide chemical protection, but little is known about how distinct extracellular polysaccharides may also protect biofilms against mechanical stresses such as shear or phagocytic engulfment. Decades-long infections of Pseudomonas. aeruginosa biofilms in the lungs of cystic fibrosis patients are natural models for studies of biofilm fitness under pressure from antibiotics and the immune system. In cystic fibrosis infections, production of the extracellular ...


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