Digital Commons Network^™

Monospecific And Bispecific Monoclonal Sars-Cov-2 Neutralizing Antibodies That Maintain Potency Against B.1.617, Lei Peng, Yingxia Hu, Madeleine C Mankowski, Ping Ren, Rita E Chen, Michael S Diamond, Et Al

Open Access Publications

COVID-19 pathogen SARS-CoV-2 has infected hundreds of millions and caused over 5 million deaths to date. Although multiple vaccines are available, breakthrough infections occur especially by emerging variants. Effective therapeutic options such as monoclonal antibodies (mAbs) are still critical. Here, we report the development, cryo-EM structures, and functional analyses of mAbs that potently neutralize SARS-CoV-2 variants of concern. By high-throughput single cell sequencing of B cells from spike receptor binding domain (RBD) immunized animals, we identify two highly potent SARS-CoV-2 neutralizing mAb clones that have single-digit nanomolar affinity and low-picomolar avidity, and generate a bispecific antibody. Lead antibodies show strong …

The Effects Of The Combination Of Mesenchymal Stromal Cells And Nanofiber-Hydrogel Composite On Repair Of The Contused Spinal Cord, Agnes E Haggerty, Ines Maldonado-Lasunción, Yohshiro Nitobe, Kentaro Yamane, Megan M Marlow, Hua You, Chi Zhang, Brian Cho, Xiaowei Li, Sashank Reddy, Hai-Quan Mao, Martin Oudega

Open Access Publications

A bone marrow-derived mesenchymal stromal cell (MSC) transplant and a bioengineered nanofiber-hydrogel composite (NHC) have been shown to stimulate nervous tissue repair in the contused spinal cord in rodent models. Here, these two modalities were combined to assess their repair effects in the contused spinal cord in adult rats. Cohorts of contused rats were treated with MSC in NHC (MSC-NHC), MSC in phosphate-buffered saline (MSC-PBS), NHC, or PBS injected into the contusion site at 3 days post-injury. One week after injury, there were significantly fewer CD68+ cells in the contusion with MSC-NHC and NHC, but not MSC-PBS. The reduction in …

Wwox Binding To The Murine Brca1-Brct Domain Regulates Timing Of Brip1 And Ctip Phospho-Protein Interactions With This Domain At Dna Double-Strand Breaks, And Repair Pathway Choice, Dongju Park, Mehdi Gharghabi, Colleen R Reczek, Rebecca Plow, Charles Yungvirt, C Marcelo Aldaz, Kay Huebner

Faculty, Staff and Student Publications

Wwox-deficient human cells show elevated homologous recombination, leading to resistance to killing by double-strand break-inducing agents. Human Wwox binds to the Brca1 981-PPLF-984 Wwox-binding motif, likely blocking the pChk2 phosphorylation site at Brca1-S988. This phosphorylation site is conserved across mammalian species; the PPLF motif is conserved in primates but not in rodents. We now show that murine Wwox does not bind Brca1 near the conserved mouse Brca1 phospho-S971 site, leaving it open for Chk2 phosphorylation and Brca1 activation. Instead, murine Wwox binds to Brca1 through its BRCT domain, where pAbraxas, pBrip1, and pCtIP, of the A, B, and C binding …

Cross-Omics Analysis Of Fenugreek Supplementation Reveals Beneficial Effects Are Caused By Gut Microbiome Changes Not Mammalian Host Physiology, Katarina A. Jones, Allison J. Richard, J Michael Salbaum, Susan Newman Phd, Rn, Crrn, Faan, Richard Carmouche, Sara Webb, Annadora J. Bruce-Keller, Jacqueline M. Stephens, Shawn R. Campagna

Publications

Herbal remedies are increasing in popularity as treatments for metabolic conditions such as obesity and Type 2 Diabetes. One potential therapeutic option is fenugreek seeds (Trigonella foenum-graecum), which have been used for treating high cholesterol and Type 2 diabetes. A proposed mechanism for these benefits is through alterations in the microbiome, which impact mammalian host metabolic function. This study used untargeted metabolomics to investigate the fenugreek-induced alterations in the intestinal, liver, and serum profiles of mice fed either a 60% high-fat or low-fat control diet each with or without fenugreek supplementation (2% w/w) for 14 weeks. Metagenomic analyses of intestinal …

Cross-Omics Analysis Of Fenugreek Supplementation Reveals Beneficial Effects Are Caused By Gut Microbiome Changes Not Mammalian Host Physiology, Katarina A. Jones, Allison J. Richard, J Michael Salbaum, Susan Newman Phd, Rn, Crrn, Faan, Richard Carmouche, Sara Webb, Annadora J. Bruce-Keller, Jacqueline M. Stephens, Shawn R. Campagna

Publications

Herbal remedies are increasing in popularity as treatments for metabolic conditions such as obesity and Type 2 Diabetes. One potential therapeutic option is fenugreek seeds (Trigonella foenum-graecum), which have been used for treating high cholesterol and Type 2 diabetes. A proposed mechanism for these benefits is through alterations in the microbiome, which impact mammalian host metabolic function. This study used untargeted metabolomics to investigate the fenugreek-induced alterations in the intestinal, liver, and serum profiles of mice fed either a 60% high-fat or low-fat control diet each with or without fenugreek supplementation (2% w/w) for 14 weeks. Metagenomic analyses of intestinal …

The Bacterial Microbiota Regulates Normal Hematopoiesis Via Metabolite-Induced Type 1 Interferon Signaling, Hannah Yan, Forrest C Walker, Arushana Ali, Hyojeong Han, Lin Tan, Lucas Veillon, Philip L Lorenzi, Megan T Baldridge, Katherine Y King

Open Access Publications

Antibiotic therapy, especially when administered long term, is associated with adverse hematologic effects such as cytopenia. Signals from the intestinal microbiota are critical to maintain normal hematopoiesis, and antibiotics can cause bone marrow suppression through depletion of the microbiota. We reported previously that STAT1 signaling is necessary for microbiota-dependent hematopoiesis, but the precise mechanisms by which the gut microbiota signals to the host bone marrow to regulate hematopoiesis remain undefined. We sought to identify the cell type(s) through which STAT1 promotes microbiota-mediated hematopoiesis and to elucidate which upstream signaling pathways trigger STAT1 signaling. Using conditional knockout and chimeric mice, we …

Mechanisms Of Mitochondrial Promoter Recognition In Humans And Other Mammalian Species, Angelica Zamudio-Ochoa, Yaroslav I Morozov, Azadeh Sarfallah, Michael Anikin, Dmitry Temiakov

Department of Biochemistry and Molecular Biology Faculty Papers

Recognition of mammalian mitochondrial promoters requires the concerted action of mitochondrial RNA polymerase (mtRNAP) and transcription initiation factors TFAM and TFB2M. In this work, we found that transcript slippage results in heterogeneity of the human mitochondrial transcripts in vivo and in vitro. This allowed us to correctly interpret the RNAseq data, identify the bona fide transcription start sites (TSS), and assign mitochondrial promoters for > 50% of mammalian species and some other vertebrates. The divergent structure of the mammalian promoters reveals previously unappreciated aspects of mtDNA evolution. The correct assignment of TSS also enabled us to establish the precise register of …

Adult Mouse Fibroblasts Retain Organ-Specific Transcriptomic Identity., Elvira Forte, Mirana Ramialison, Hieu T Nim, Madison Mara, Jacky Y Li, Rachel Cohn, Sandra Daigle, Sarah Boyd, Edouard G Stanley, Andrew G Elefanty, J Travis Hinson, Mauro W Costa, Nadia Rosenthal, Milena B Furtado

Faculty Research 2022

Organ fibroblasts are essential components of homeostatic and diseased tissues. They participate in sculpting the extracellular matrix, sensing the microenvironment, and communicating with other resident cells. Recent studies have revealed transcriptomic heterogeneity among fibroblasts within and between organs. To dissect the basis of interorgan heterogeneity, we compare the gene expression of murine fibroblasts from different tissues (tail, skin, lung, liver, heart, kidney, and gonads) and show that they display distinct positional and organ-specific transcriptome signatures that reflect their embryonic origins. We demonstrate that expression of genes typically attributed to the surrounding parenchyma by fibroblasts is established in embryonic development and …

Drosophila Functional Screening Of De Novo Variants In Autism Uncovers Damaging Variants And Facilitates Discovery Of Rare Neurodevelopmental Diseases, Paul C Marcogliese, Samantha L Deal, Jonathan Andrews, J Michael Harnish, V Hemanjani Bhavana, Hillary K Graves, Sharayu Jangam, Xi Luo, Ning Liu, Danqing Bei, Yu-Hsin Chao, Brooke Hull, Pei-Tseng Lee, Hongling Pan, Pradnya Bhadane, Mei-Chu Huang, Colleen M Longley, Hsiao-Tuan Chao, Hyung-Lok Chung, Nele A Haelterman, Oguz Kanca, Sathiya N Manivannan, Linda Z Rossetti, Ryan J German, Amanda Gerard, Eva Maria Christina Schwaibold, Sarah Fehr, Renzo Guerrini, Annalisa Vetro, Eleina England, Chaya N Murali, Tahsin Stefan Barakat, Marieke F Van Dooren, Martina Wilke, Marjon Van Slegtenhorst, Gaetan Lesca, Isabelle Sabatier, Nicolas Chatron, Catherine A Brownstein, Jill A Madden, Pankaj B Agrawal, Boris Keren, Thomas Courtin, Laurence Perrin, Melanie Brugger, Timo Roser, Steffen Leiz, Frederic Tran Mau-Them, Julian Delanne, Elena Sukarova-Angelovska, Slavica Trajkova, Erik Rosenhahn, Vincent Strehlow, Konrad Platzer, Roberto Keller, Lisa Pavinato, Alfredo Brusco, Jill A Rosenfeld, Ronit Marom, Michael F Wangler, Shinya Yamamoto

Duncan NRI Faculty and Staff Publications

Individuals with autism spectrum disorder (ASD) exhibit an increased burden of de novo mutations (DNMs) in a broadening range of genes. While these studies have implicated hundreds of genes in ASD pathogenesis, which DNMs cause functional consequences in vivo remains unclear. We functionally test the effects of ASD missense DNMs using Drosophila through "humanization" rescue and overexpression-based strategies. We examine 79 ASD variants in 74 genes identified in the Simons Simplex Collection and find 38% of them to cause functional alterations. Moreover, we identify GLRA2 as the cause of a spectrum of neurodevelopmental phenotypes beyond ASD in 13 previously undiagnosed …

Discovery And Preclinical Pharmacology Of Ine963, A Potent And Fast-Acting Blood-Stage Antimalarial With A High Barrier To Resistance And Potential For Single-Dose Cures In Uncomplicated Malaria., Benjamin R. Taft, Fumiaki Yokokawa, Tom Kirrane, Anne-Catherine Mata, Richard Huang, Nicole Blaquiere, Grace Waldron, Bin Zou, Oliver Simon, Subramanyam Vankadara, Wai Ling Chan, Mei Ding, Sandra Sim, Judith Straimer, Armand Guiguemde, Suresh B Lakshminarayana, Jay Prakash Jain, Christopher Bodenreider, Christopher Thompson, Christian Lanshoeft, Wei Shu, Eric Fang, Jafri Qumber, Katherine Chan, Luying Pei, Yen-Liang Chen, Hanna Schulz, Jessie Lim, Siti Nurdiana Abas, Xiaoman Ang, Yugang Liu, Iñigo Angulo-Barturen, María Belén Jiménez-Díaz, Glaxosmithkline Gamo, Benigno Crespo-Fernandez, Philip J. Rosenthal, Roland A. Cooper, Patrick Tumwebaze, Anna Caroline Campos Aguiar, Brice Campo, Simon Campbell, Jürgen Wagner, Thierry T Diagana, Christopher Sarko

Natural Sciences and Mathematics | Faculty Scholarship

A series of 5-aryl-2-amino-imidazothiadiazole (ITD) derivatives were identified by a phenotype-based high-throughput screening using a blood stage Plasmodium falciparum (Pf) growth inhibition assay. A lead optimization program focused on improving antiplasmodium potency, selectivity against human kinases, and absorption, distribution, metabolism, excretion, and toxicity properties and extended pharmacological profiles culminated in the identification of INE963 (1), which demonstrates potent cellular activity against Pf 3D7 (EC50 = 0.006 μM) and achieves "artemisinin-like" kill kinetics in vitro with a parasite clearance time of/kg is fully curative in the Pf-humanized severe combined immunodeficient mouse model. INE963 (1) also exhibits a high barrier to resistance …

Pftaire Kinase L63 Interactor 1a (Pif1a Protein) Is Required For Actin Cone Movement During Spermatid Individualization In Drosophila Melanogaster, Harrison D Pravder, Dorota Grabowska, Kaushik Roychoudhury, Betty Zhang, Deborah Frank, Przemysław Zakrzewski, Marta Lenartowska, Kathryn G Miller

Open Access Publications

A useful model for determining the mechanisms by which actin and actin binding proteins control cellular architecture is the

Key Factors For Improving Rigor And Reproducibility: Guidelines, Peer Reviews, And Journal Technical Reviews, Hong S. Lu, Alan Daugherty

Saha Cardiovascular Research Center Faculty Publications

To respond to the NIH's policy for rigor and reproducibility in preclinical research, many journals have implemented guidelines and checklists to guide authors in improving the rigor and reproducibility of their research. Transparency in developing detailed prospective experimental designs and providing raw data are essential premises of rigor and reproducibility. Standard peer reviews and journal-specific technical and statistical reviews are critical factors for enhancing rigor and reproducibility. This brief review also shares some experience from Arteriosclerosis, Thrombosis, and Vascular Biology, an American Heart Association journal, that has implemented several mechanisms to enhance rigor and reproducibility for preclinical research.

Defining The Mammalian Coactivation Of Hepatic 12-H Clock And Lipid Metabolism, Huan Meng, Naomi M Gonzales, Sung Yun Jung, Yue Lu, Nagireddy Putluri, Bokai Zhu, Clifford C Dacso, David M Lonard, Bert W O'Malley

Faculty, Staff and Students Publications

The 12-h clock coordinates lipid homeostasis, energy metabolism, and stress rhythms via the transcriptional regulator XBP1. However, the biochemical and physiological bases for integrated control of the 12-h clock and diverse metabolic pathways remain unclear. Here, we show that steroid receptor coactivator SRC-3 coactivates XBP1 transcription and regulates hepatic 12-h cistrome and gene rhythmicity. Mice lacking SRC-3 show abnormal 12-h rhythms in hepatic transcription, metabolic functions, systemic energetics, and rate-limiting lipid metabolic processes, including triglyceride, phospholipid, and cardiolipin pathways. Notably, 12-h clock coactivation is not only preserved, with its cistromic activation priming ahead of the zeitgeber cue of light, but …

Ezh2 And Endometrial Cancer Development: Insights From A Mouse Model, Xin Fang, Nan Ni, Xiaofang Wang, Yanan Tian, Ivan Ivanov, Monique Rijnkels, Kayla J Bayless, John P Lydon, Qinglei Li

Faculty, Staff and Students Publications

Enhancer of zeste homolog 2 (EZH2), a core component of polycomb repressive complex 2, plays an important role in cancer development. As both oncogenic and tumor suppressive functions of EZH2 have been documented in the literature, the objective of this study is to determine the impact of Ezh2 deletion on the development and progression of endometrial cancer induced by inactivation of phosphatase and tensin homolog (PTEN), a tumor suppressor gene frequently dysregulated in endometrial cancer patients. To this end, we created mice harboring uterine deletion of both Ezh2 and Pten using Cre recombinase driven by the progesterone receptor …

Glynac (Glycine And N-Acetylcysteine) Supplementation In Mice Increases Length Of Life By Correcting Glutathione Deficiency, Oxidative Stress, Mitochondrial Dysfunction, Abnormalities In Mitophagy And Nutrient Sensing, And Genomic Damage, Premranjan Kumar, Ob W Osahon, Rajagopal V Sekhar

Faculty, Staff and Students Publications

Determinants of length of life are not well understood, and therefore increasing lifespan is a challenge. Cardinal theories of aging suggest that oxidative stress (OxS) and mitochondrial dysfunction contribute to the aging process, but it is unclear if they could also impact lifespan. Glutathione (GSH), the most abundant intracellular antioxidant, protects cells from OxS and is necessary for maintaining mitochondrial health, but GSH levels decline with aging. Based on published human studies where we found that supplementing glycine and N-acetylcysteine (GlyNAC) improved/corrected GSH deficiency, OxS and mitochondrial dysfunction, we hypothesized that GlyNAC supplementation could increase longevity. We tested our hypothesis …

Fly Cell Atlas: A Single-Nucleus Transcriptomic Atlas Of The Adult Fruit Fly, Hongjie Li, Jasper Janssens, Maxime De Waegeneer, Sai Saroja Kolluru, Kristofer Davie, Vincent Gardeux, Wouter Saelens, Fabrice P A David, Maria Brbić, Katina Spanier, Jure Leskovec, Colleen N Mclaughlin, Qijing Xie, Robert C Jones, Katja Brueckner, Jiwon Shim, Sudhir Gopal Tattikota, Frank Schnorrer, Katja Rust, Todd G Nystul, Zita Carvalho-Santos, Carlos Ribeiro, Soumitra Pal, Sharvani Mahadevaraju, Teresa M Przytycka, Aaron M Allen, Stephen F Goodwin, Cameron W Berry, Margaret T Fuller, Helen White-Cooper, Erika L Matunis, Stephen Dinardo, Anthony Galenza, Lucy Erin O'Brien, Julian A T Dow, Fca Consortium, Heinrich Jasper, Brian Oliver, Norbert Perrimon, Bart Deplancke, Stephen R Quake, Liqun Luo, Stein Aerts, Devika Agarwal, Yasir Ahmed-Braimah, Michelle Arbeitman, Majd M Ariss, Jordan Augsburger, Kumar Ayush, Catherine C Baker, Torsten Banisch, Katja Birker, Rolf Bodmer, Benjamin Bolival, Susanna E Brantley, Julie A Brill, Nora C Brown, Norene A Buehner, Xiaoyu Tracy Cai, Rita Cardoso-Figueiredo, Fernando Casares, Amy Chang, Thomas R Clandinin, Sheela Crasta, Claude Desplan, Angela M Detweiler, Darshan B Dhakan, Erika Donà, Stefanie Engert, Swann Floc'hlay, Nancy George, Amanda J González-Segarra, Andrew K Groves, Samantha Gumbin, Yanmeng Guo, Devon E Harris, Yael Heifetz, Stephen L Holtz, Felix Horns, Bruno Hudry, Ruei-Jiun Hung, Yuh Nung Jan, Jacob S Jaszczak, Gregory S X E Jefferis, Jim Karkanias, Timothy L Karr, Nadja Sandra Katheder, James Kezos, Anna A Kim, Seung K Kim, Lutz Kockel, Nikolaos Konstantinides, Thomas B Kornberg, Henry M Krause, Andrew Thomas Labott, Meghan Laturney, Ruth Lehmann, Sarah Leinwand, Jiefu Li, Joshua Shing Shun Li, Kai Li, Ke Li, Liying Li, Tun Li, Maria Litovchenko, Han-Hsuan Liu, Yifang Liu, Tzu-Chiao Lu, Jonathan Manning, Anjeli Mase, Mikaela Matera-Vatnick, Neuza Reis Matias, Caitlin E Mcdonough-Goldstein, Aaron Mcgeever, Alex D Mclachlan, Paola Moreno-Roman, Norma Neff, Megan Neville, Sang Ngo, Tanja Nielsen, Caitlin E O'Brien, David Osumi-Sutherland, Mehmet Neset Özel, Irene Papatheodorou, Maja Petkovic, Clare Pilgrim, Angela Oliveira Pisco, Carolina Reisenman, Erin Nicole Sanders, Gilberto Dos Santos, Kristin Scott, Aparna Sherlekar, Philip Shiu, David Sims, Rene V Sit, Maija Slaidina, Harold E Smith, Gabriella Sterne, Yu-Han Su, Daniel Sutton, Marco Tamayo, Michelle Tan, Ibrahim Tastekin, Christoph Treiber, David Vacek, Georg Vogler, Scott Waddell, Wanpeng Wang, Rachel I Wilson, Mariana F Wolfner, Yiu-Cheung E Wong, Anthony Xie, Jun Xu, Shinya Yamamoto, Jia Yan, Zepeng Yao, Kazuki Yoda, Ruijun Zhu, Robert P Zinzen

Duncan NRI Faculty and Staff Publications

For more than 100 years, the fruit fly Drosophila melanogaster has been one of the most studied model organisms. Here, we present a single-cell atlas of the adult fly, Tabula Drosophilae, that includes 580,000 nuclei from 15 individually dissected sexed tissues as well as the entire head and body, annotated to >250 distinct cell types. We provide an in-depth analysis of cell type-related gene signatures and transcription factor markers, as well as sexual dimorphism, across the whole animal. Analysis of common cell types between tissues, such as blood and muscle cells, reveals rare cell types and tissue-specific subtypes. This …

Elucidating The Clinical Spectrum And Molecular Basis Of Hyal2 Deficiency, James Fasham, Siying Lin, Promita Ghosh, Francesca Clementina Radio, Emily G Farrow, Isabelle Thiffault, Jennifer Kussman, Dihong Zhou, Rick Hemming, Kenneth Zahka, Barry A Chioza, Lettie E Rawlins, Olivia K Wenger, Adam C Gunning, Simone Pizzi, Roberta Onesimo, Giuseppe Zampino, Emily Barker, Natasha Osawa, Megan Christine Rodriguez, Teresa M Neuhann, Elaine H Zackai, Beth Keena, Jenina Capasso, Alex V Levin, Elizabeth Bhoj, Dong Li, Hakon Hakonarson, Ingrid M Wentzensen, Adam Jackson, Kate E Chandler, Zeynep H Coban-Akdemir, Jennifer E Posey, Siddharth Banka, James R Lupski, Sarah E Sheppard, Marco Tartaglia, Barbara Triggs-Raine, Andrew H Crosby, Emma L Baple

Faculty, Staff and Student Publications

PURPOSE: We previously defined biallelic HYAL2 variants causing a novel disorder in 2 families, involving orofacial clefting, facial dysmorphism, congenital heart disease, and ocular abnormalities, with Hyal2 knockout mice displaying similar phenotypes. In this study, we better define the phenotype and pathologic disease mechanism.

METHODS: Clinical and genomic investigations were undertaken alongside molecular studies, including immunoblotting and immunofluorescence analyses of variant/wild-type human HYAL2 expressed in mouse fibroblasts, and in silico modeling of putative pathogenic variants.

RESULTS: Ten newly identified individuals with this condition were investigated, and they were associated with 9 novel pathogenic variants. Clinical studies defined genotype-phenotype correlations and …

Tissue-Targeted Crispr-Cas9-Mediated Genome Editing Of Multiple Homeologs In F0 Generation Xenopus Laevis Embryos, Mark E Corkins, Bridget D Delay, Rachel K Miller

Faculty, Staff and Student Publications

Xenopus laevis frogs are a powerful developmental model that enables studies combining classical embryology and molecular manipulation. Because of the large embryo size, ease of microinjection, and ability to target tissues through established fate maps, X. laevis have become the predominant amphibian research model. Given that their allotetraploid genome has complicated the generation of gene knockouts, strategies need to be established for efficient mutagenesis of multiple homeologs to evaluate gene function. Here we describe a protocol to utilize CRISPR-Cas9-mediated genome editing to target either single alleles or multiple alloalleles in F0 X. laevis embryos. A single guide (sg) RNA is …

Isoform-Specific Functions Of Pparγ In Gene Regulation And Metabolism, Wenxiang Hu, Chunjie Jiang, Mindy Kim, Yang Xiao, Hannah J Richter, Dongyin Guan, Kun Zhu, Brianna M Krusen, Arielle N Roberts, Jessica Miller, David J Steger, Mitchell A Lazar

Faculty, Staff and Students Publications

Peroxisome proliferator-activated receptor γ (PPARγ) is a nuclear receptor that is a vital regulator of adipogenesis, insulin sensitivity, and lipid metabolism. Activation of PPARγ by antidiabetic thiazolidinediones (TZD) reverses insulin resistance but also leads to weight gain that limits the use of these drugs. There are two main PPARγ isoforms, but the specific functions of each are not established. Here we generated mouse lines in which endogenous PPARγ1 and PPARγ2 were epitope-tagged to interrogate isoform-specific genomic binding, and mice deficient in either PPARγ1 or PPARγ2 to assess isoform-specific gene regulation. Strikingly, although PPARγ1 and PPARγ2 contain identical DNA binding domains, …

Elucidating The Clinical Spectrum And Molecular Basis Of Hyal2 Deficiency., James Fasham, Siying Lin, Promita Ghosh, Francesca Clementina Radio, Emily G. Farrow, Isabelle Thiffault, Jennifer Kussman, Dihong Zhou, Rick Hemming, Kenneth Zahka, Barry A. Chioza, Lettie E. Rawlins, Olivia K. Wenger, Adam C. Gunning, Simone Pizzi, Roberta Onesimo, Giuseppe Zampino, Emily Barker, Natasha Osawa, Megan Christine Rodriguez, Teresa M. Neuhann, Elaine H. Zackai, Beth Keena, Jenina Capasso, Alex V. Levin, Elizabeth Bhoj, Dong Li, Hakon Hakonarson, Ingrid M. Wentzensen, Adam Jackson, Kate E. Chandler, Zeynep H. Coban-Akdemir, Jennifer E. Posey, Siddharth Banka, James R. Lupski, Sarah E. Sheppard, Marco Tartaglia, Barbara Triggs-Raine, Andrew H. Crosby, Emma L. Baple

Manuscripts, Articles, Book Chapters and Other Papers

PURPOSE: We previously defined biallelic HYAL2 variants causing a novel disorder in 2 families, involving orofacial clefting, facial dysmorphism, congenital heart disease, and ocular abnormalities, with Hyal2 knockout mice displaying similar phenotypes. In this study, we better define the phenotype and pathologic disease mechanism.

METHODS: Clinical and genomic investigations were undertaken alongside molecular studies, including immunoblotting and immunofluorescence analyses of variant/wild-type human HYAL2 expressed in mouse fibroblasts, and in silico modeling of putative pathogenic variants.

RESULTS: Ten newly identified individuals with this condition were investigated, and they were associated with 9 novel pathogenic variants. Clinical studies defined genotype-phenotype correlations and …

Metabolic Stress Induces Gd2+ Cancer Stem Cell-Like Phenotype In Triple-Negative Breast Cancer, Appalaraju Jaggupilli, Stanley Ly, Khoa Nguyen, Vivek Anand, Bin Yuan, Fouad El-Dana, Yuanqing Yan, Zoe Arvanitis, Danthasinghe Waduge Badrajee Piyarathna, Nagireddy Putluri, Helen Piwnica-Worms, Henry Charles Manning, Michael Andreeff, V Lokesh Battula

Faculty, Staff and Student Publications

Background: Metabolic stress resulting from nutrient deficiency is one of the hallmarks of a growing tumour. Here, we tested the hypothesis that metabolic stress induces breast cancer stem-like cell (BCSC) phenotype in triple-negative breast cancer (TNBC).

Methods: Flow cytometry for GD2 expression, mass spectrometry and Ingenuity Pathway Analysis for metabolomics, bioinformatics, in vitro tumorigenesis and in vivo models were used.

Results: Serum/glucose deprivation not only increased stress markers but also enhanced GD2+ BCSC phenotype and function in TNBC cells. Global metabolomics profiling identified upregulation of glutathione biosynthesis in GD2high cells, suggesting a role of glutamine in the BCSC phenotype. Cueing …

Chronic Estrus Disrupts Uterine Gland Development And Homeostasis, C Allison Stewart, M David Stewart, Ying Wang, Rachel D Mullen, Bonnie K Kircher, Rui Liang, Yu Liu, Richard R Behringer

Faculty, Staff and Student Publications

Female mice homozygous for an engineered Gnrhr E90K mutation have reduced gonadotropin-releasing hormone signaling, leading to infertility. Their ovaries have numerous antral follicles but no corpora lutea, indicating a block to ovulation. These mutants have high levels of circulating estradiol and low progesterone, indicating a state of persistent estrus. This mouse model provided a unique opportunity to examine the lack of cyclic levels of ovarian hormones on uterine gland biology. Although uterine gland development appeared similar to controls during prepubertal development, it was compromised during adolescence in the mutants. By age 20 weeks, uterine gland development was comparable to controls, …

Treatment Preferences For Pharmacological Versus Psychological Interventions Among Primary Care Providers In Nepal: Mixed Methods Analysis Of A Pilot Cluster Randomized Controlled Trial., Anvita Bhardwaj, Dristy Gurung, Sauharda Rai, Bonnie N Kaiser, Cori L Cafaro, Kathleen J Sikkema, Crick Lund, Nagendra P Luitel, Brandon A. Kohrt

Psychiatry and Behavioral Sciences Faculty Publications

There is increasing evidence supporting the effectiveness of psychological interventions in low- and middle-income countries. However, primary care providers (PCPs) may prefer treating patients with medication. A secondary exploratory analysis of a pilot cluster randomized controlled trial was conducted to evaluate psychological vs. pharmacological treatment preferences among PCPs. Thirty-four health facilities, including 205 PCPs, participated in the study, with PCPs in 17 facilities assigned to a standard version of the mental health Gap Action Programme (mhGAP) training delivered by mental health specialists. PCPs in the other 17 facilities received mhGAP instruction delivered by specialists and people with lived experience of …

Aloin Isoforms (A And B) Selectively Inhibits Proteolytic And Deubiquitinating Activity Of Papain Like Protease (Plpro) Of Sars-Cov-2 In Vitro., Devin S M Lewis, Joanna Ho, Savannah Wills, Anasha Kawall, Avini Sharma, Krishna Chavada, Maximilian C C J C Ebert, Stefania Evoli, Ajay Singh, Srujana Rayalam, Vicky Mody, Shashidharamurthy Taval

PCOM Scholarly Works

The most common host entry point of human adapted coronaviruses (CoV) including SARS-CoV-2 is through the initial colonization in the nostril and mouth region which is responsible for spread of the infection. Most recent studies suggest that the commercially available oral and nasal rinse products are effective in inhibiting the viral replication. However, the anti-viral mechanism of the active ingredients present in the oral rinses have not been studied. In the present study, we have assessed in vitro enzymatic inhibitory activity of active ingredients in the oral mouth rinse products: aloin A and B, chlorhexidine, eucalyptol, hexetidine, menthol, triclosan, methyl …

Ddr1 Contributes To Kidney Inflammation And Fibrosis By Promoting The Phosphorylation Of Bcr And Stat3, Corina M. Borza, Gema Bolas, Fabian Bock, Xiuqi Zhang, Favour C Akabogu, Ming-Zhi Zhang, Mark De Caestecker, Min Yang, Haichun Yang, Ethan Lee, Leslie Gewin, Agnes B Fogo, W Hayes Mcdonald, Roy Zent, Ambra Pozzi

Open Access Publications

Discoidin domain receptor 1 (DDR1), a receptor tyrosine kinase activated by collagen, contributes to chronic kidney disease. However, its role in acute kidney injury and subsequent development of kidney fibrosis is not clear. Thus, we performed a model of severe ischemia/reperfusion-induced acute kidney injury that progressed to kidney fibrosis in WT and Ddr1-null mice. We showed that Ddr1-null mice had reduced acute tubular injury, inflammation, and tubulointerstitial fibrosis with overall decreased renal monocyte chemoattractant protein (MCP-1) levels and STAT3 activation. We identified breakpoint cluster region (BCR) protein as a phosphorylated target of DDR1 that controls MCP-1 production in renal proximal …

Cryo-Et Of Toxoplasma Parasites Gives Subnanometer Insight Into Tubulin-Based Structures, Stella Y Sun, Li-Av Segev-Zarko, Muyuan Chen, Grigore D Pintilie, Michael F Schmid, Steven J Ludtke, John C Boothroyd, Wah Chiu

Faculty, Staff and Students Publications

Tubulin is a conserved protein that polymerizes into different forms of filamentous structures in Toxoplasma gondii, an obligate intracellular parasite in the phylum Apicomplexa. Two key tubulin-containing cytoskeletal components are subpellicular microtubules (SPMTs) and conoid fibrils (CFs). The SPMTs help maintain shape and gliding motility, while the CFs are implicated in invasion. Here, we use cryogenic electron tomography to determine the molecular structures of the SPMTs and CFs in vitrified intact and detergent-extracted parasites. Subvolume densities from detergent-extracted parasites yielded averaged density maps at subnanometer resolutions, and these were related back to their architecture in situ. An intralumenal spiral …

Myocardial Rev-Erb-Mediated Diurnal Metabolic Rhythm And Obesity Paradox, Shiyang Song, Chih-Liang Tien, Hao Cui, Paul Basil, Ningxia Zhu, Yingyun Gong, Wenbo Li, Hui Li, Qiying Fan, Jong Min Choi, Weijia Luo, Yanfeng Xue, Rui Cao, Wenjun Zhou, Andrea R Ortiz, Brittany Stork, Vatsala Mundra, Nagireddy Putluri, Brian York, Maoping Chu, Jiang Chang, Sung Yun Jung, Liang Xie, Jiangping Song, Lilei Zhang, Zheng Sun

Faculty, Staff and Students Publications

BACKGROUND: The nuclear receptor Rev-erbα/β, a key component of the circadian clock, emerges as a drug target for heart diseases, but the function of cardiac Rev-erb has not been studied in vivo. Circadian disruption is implicated in heart diseases, but it is unknown whether cardiac molecular clock dysfunction is associated with the progression of any naturally occurring human heart diseases. Obesity paradox refers to the seemingly protective role of obesity for heart failure, but the mechanism is unclear.

METHODS: We generated mouse lines with cardiac-specific Rev-erbα/β knockout (KO), characterized cardiac phenotype, conducted multi-omics (RNA-sequencing, chromatin immunoprecipitation sequencing, proteomics, and metabolomics) …

Live-Dead Assay On Unlabeled Cells Using Phase Imaging With Computational Specificity, Chenfei Hu, Shenghua He, Young Jae Lee, Yuchen He, Edward M Kong, Hua Li, Mark A Anastasio, Gabriel Popescu

Open Access Publications

Existing approaches to evaluate cell viability involve cell staining with chemical reagents. However, the step of exogenous staining makes these methods undesirable for rapid, nondestructive, and long-term investigation. Here, we present an instantaneous viability assessment of unlabeled cells using phase imaging with computation specificity. This concept utilizes deep learning techniques to compute viability markers associated with the specimen measured by label-free quantitative phase imaging. Demonstrated on different live cell cultures, the proposed method reports approximately 95% accuracy in identifying live and dead cells. The evolution of the cell dry mass and nucleus area for the labeled and unlabeled populations reveal …

Srgn-Triggered Aggressive And Immunosuppressive Phenotype In A Subset Of Ttf-1-Negative Lung Adenocarcinomas, Ichidai Tanaka, Delphine Dayde, Mei Chee Tai, Haruki Mori, Luisa M Solis, Satyendra C Tripathi, Johannes F Fahrmann, Nese Unver, Gargy Parhy, Rekha Jain, Edwin R Parra, Yoshiko Murakami, Clemente Aguilar-Bonavides, Barbara Mino, Muge Celiktas, Dilsher Dhillon, Julian Phillip Casabar, Masahiro Nakatochi, Francesco Stingo, Veera Baladandayuthapani, Hong Wang, Hiroyuki Katayama, Jennifer B Dennison, Philip L Lorenzi, Kim-Anh Do, Junya Fujimoto, Carmen Behrens, Edwin J Ostrin, Jaime Rodriguez-Canales, Tetsunari Hase, Takayuki Fukui, Taisuke Kajino, Seiichi Kato, Yasushi Yatabe, Waki Hosoda, Koji Kawaguchi, Kohei Yokoi, Toyofumi F Chen-Yoshikawa, Yoshinori Hasegawa, Adi F Gazdar, Ignacio I Wistuba, Samir Hanash, Ayumu Taguchi

Faculty, Staff and Student Publications

BACKGROUND: Approximately 20% of lung adenocarcinoma (LUAD) is negative for the lineage-specific oncogene Thyroid transcription factor 1 (TTF-1) and exhibits worse clinical outcome with a low frequency of actionable genomic alterations. To identify molecular features associated with TTF-1-negative LUAD, we compared the transcriptomic and proteomic profiles of LUAD cell lines. SRGN , a chondroitin sulfate proteoglycan Serglycin, was identified as a markedly overexpressed gene in TTF-1-negative LUAD. We therefore investigated the roles and regulation of SRGN in TTF-1-negative LUAD.

METHODS: Proteomic and metabolomic analyses of 41 LUAD cell lines were done using mass spectrometry. The function of SRGN was investigated …

Highly Reliable Creation Of Floxed Alleles By Electroporating Single-Cell Embryos, Monica F Sentmanat, J Michael White, Evguenia Kouranova, Xiaoxia Cui

2020-Current year OA Pubs

BACKGROUND: Floxed (flanked by loxP) alleles are a crucial portion of conditional knockout mouse models. However, an efficient and reliable strategy to flox genomic regions of any desired size is still lacking.

RESULTS: Here, we demonstrate that the method combining electroporation of fertilized eggs with gRNA/Cas9 complexes and single-stranded oligodeoxynucleotides (ssODNs), assessing phasing of loxP insertions in founders using an in vitro Cre assay and an optional, highly specific and efficient second-round targeting ensures the generation of floxed F1 animals in roughly five months for a wide range of sequence lengths (448 bp to 160 kb reported here).

CONCLUSIONS: Floxed …