Genetic Processes Commons^™

Nitric Oxide Is Required For Lung Alveolarization Revealed By Deficiency Of Argininosuccinate Lyase, Zixue Jin, Ming-Ming Jiang, Brendan Lee

Faculty, Staff and Students Publications

Inhaled nitric oxide (NO) therapy has been reported to improve lung growth in premature newborns. However, the underlying mechanisms by which NO regulates lung development remain largely unclear. NO is enzymatically produced by three isoforms of nitric oxide synthase (NOS) enzymes. NOS knockout mice are useful tools to investigate NO function in the lung. Each single NOS knockout mouse does not show obvious lung alveolar phenotype, likely due to compensatory mechanisms. While mice lacking all three NOS isoforms display impaired lung alveolarization, implicating NO plays a pivotal role in lung alveolarization. Argininosuccinate lyase (ASL) is the only mammalian enzyme capable …

Boosting Glycolysis To Combat Fragile Bone In Type 1 Diabetes, Zixue Jin, Brendan Lee

Faculty, Staff and Students Publications

Individuals with type 1 diabetes (T1D) have an increased risk of osteoporosis and fracture. In this issue of Cell Chemical Biology, Ji et al.1 show that impaired glucose metabolism in the bone-forming osteoblast drives diabetic osteoporosis in Akita mice, a mouse model of T1D.

A Single Dominant Locus Restricts Retrovirus Replication In Ybr/Ei Mice, Helen A Beilinson, Amanda Sevilleja, Jessica Spring, Fernando Benavides, Vera Beilinson, Nickolas Neokosmidis, Tatyana Golovkina

Faculty, Staff and Student Publications

Differential responses to viral infections are influenced by the genetic makeup of the host. Studies of resistance to retroviruses in human populations are complicated due to the inability to conduct proof-of-principle studies. Inbred mouse lines, which have a range of susceptible phenotypes to retroviruses, are an ideal tool to identify and characterize mechanisms of resistance and define their genetic underpinnings. YBR/Ei mice become infected with Mouse Mammary Tumor Virus, a mucosally transmitted murine retrovirus, but eliminate the virus from their pedigrees. Virus elimination correlates with a lack of virus-specific neonatal oral tolerance, which is a major mechanism for blocking the …

Spatial Organization Of The Mouse Retina At Single Cell Resolution By Merfish, Jongsu Choi, Jin Li, Salma Ferdous, Qingnan Liang, Jeffrey R Moffitt, Rui Chen

Faculty, Staff and Students Publications

The visual signal processing in the retina requires the precise organization of diverse neuronal types working in concert. While single-cell omics studies have identified more than 120 different neuronal subtypes in the mouse retina, little is known about their spatial organization. Here, we generated the single-cell spatial atlas of the mouse retina using multiplexed error-robust fluorescence in situ hybridization (MERFISH). We profiled over 390,000 cells and identified all major cell types and nearly all subtypes through the integration with reference single-cell RNA sequencing (scRNA-seq) data. Our spatial atlas allowed simultaneous examination of nearly all cell subtypes in the retina, revealing …

Effective Methods For Bulk Rna-Seq Deconvolution Using Scnrna-Seq Transcriptomes, Francisco Avila Cobos, Mohammad Javad Najaf Panah, Jessica Epps, Xiaochen Long, Tsz-Kwong Man, Hua-Sheng Chiu, Elad Chomsky, Evgeny Kiner, Michael J Krueger, Diego Di Bernardo, Luis Voloch, Jan Molenaar, Sander R Van Hooff, Frank Westermann, Selina Jansky, Michele L Redell, Pieter Mestdagh, Pavel Sumazin

Faculty, Staff and Students Publications

Background: RNA profiling technologies at single-cell resolutions, including single-cell and single-nuclei RNA sequencing (scRNA-seq and snRNA-seq, scnRNA-seq for short), can help characterize the composition of tissues and reveal cells that influence key functions in both healthy and disease tissues. However, the use of these technologies is operationally challenging because of high costs and stringent sample-collection requirements. Computational deconvolution methods that infer the composition of bulk-profiled samples using scnRNA-seq-characterized cell types can broaden scnRNA-seq applications, but their effectiveness remains controversial.

Results: We produced the first systematic evaluation of deconvolution methods on datasets with either known or scnRNA-seq-estimated compositions. Our analyses revealed …

Ywhae Loss Of Function Causes A Rare Neurodevelopmental Disease With Brain Abnormalities In Human And Mouse, Anne-Sophie Denommé-Pichon, Stephan C Collins, Ange-Line Bruel, Anna Mikhaleva, Christel Wagner, Valerie E Vancollie, Quentin Thomas, Martin Chevarin, Mathys Weber, Carlos E Prada, Alexis Overs, María Palomares-Bralo, Fernando Santos-Simarro, Marta Pacio-Míguez, Tiffany Busa, Eric Legius, Carlos A Bacino, Jill A Rosenfeld, Gwenaël Le Guyader, Matthieu Egloff, Xavier Le Guillou, Maria Antonietta Mencarelli, Alessandra Renieri, Salvatore Grosso, Jonathan Levy, Blandine Dozières, Isabelle Desguerre, Antonio Vitobello, Yannis Duffourd, Christopher J Lelliott, Christel Thauvin-Robinet, Christophe Philippe, Laurence Faivre, Binnaz Yalcin

Faculty, Staff and Students Publications

Purpose: Miller-Dieker syndrome is caused by a multiple gene deletion, including PAFAH1B1 and YWHAE. Although deletion of PAFAH1B1 causes lissencephaly unambiguously, deletion of YWHAE alone has not clearly been linked to a human disorder.

Methods: Cases with YWHAE variants were collected through international data sharing networks. To address the specific impact of YWHAE loss of function, we phenotyped a mouse knockout of Ywhae.

Results: We report a series of 10 individuals with heterozygous loss-of-function YWHAE variants (3 single-nucleotide variants and 7 deletions < 1 Mb encompassing YWHAE but not PAFAH1B1), including 8 new cases and 2 follow-ups, added with 5 cases (copy number variants) from literature review. Although, until now, only 1 intragenic deletion has been described in YWHAE, we report 4 new variants specifically in YWHAE (3 splice variants and 1 intragenic deletion). The most frequent manifestations are developmental delay, delayed speech, seizures, and brain malformations, including corpus callosum hypoplasia, delayed myelination, and ventricular dilatation. Individuals with variants affecting YWHAE alone have milder features than those with larger deletions. Neuroanatomical studies in Ywhae-/- mice revealed brain structural defects, including thin cerebral cortex, corpus callosum dysgenesis, and hydrocephalus paralleling those seen in humans.

Conclusion: This study further demonstrates that YWHAE loss-of-function variants cause a neurodevelopmental disease with brain abnormalities.

A Global Catalog Of Whole-Genome Diversity From 233 Primate Species\, Lukas F K Kuderna, Hong Gao, Mareike C Janiak, Martin Kuhlwilm, Joseph D Orkin, Thomas Bataillon, Shivakumara Manu, Alejandro Valenzuela, Juraj Bergman, Marjolaine Rousselle, Felipe Ennes Silva, Lidia Agueda, Julie Blanc, Marta Gut, Dorien De Vries, Ian Goodhead, R Alan Harris, Muthuswamy Raveendran, Axel Jensen, Idrissa S Chuma, Julie E Horvath, Christina Hvilsom, David Juan, Peter Frandsen, Joshua G Schraiber, Fabiano R De Melo, Fabrício Bertuol, Hazel Byrne, Iracilda Sampaio, Izeni Farias, João Valsecchi, Malu Messias, Maria N F Da Silva, Mihir Trivedi, Rogerio Rossi, Tomas Hrbek, Nicole Andriaholinirina, Clément J Rabarivola, Alphonse Zaramody, Clifford J Jolly, Jane Phillips-Conroy, Gregory Wilkerson, Christian Abee, Joe H Simmons, Eduardo Fernandez-Duque, Sree Kanthaswamy, Fekadu Shiferaw, Dongdong Wu, Long Zhou, Yong Shao, Guojie Zhang, Julius D Keyyu, Sascha Knauf, Minh D Le, Esther Lizano, Stefan Merker, Arcadi Navarro, Tilo Nadler, Chiea Chuen Khor, Jessica Lee, Patrick Tan, Weng Khong Lim, Andrew C Kitchener, Dietmar Zinner, Ivo Gut, Amanda D Melin, Katerina Guschanski, Mikkel Heide Schierup, Robin M D Beck, Govindhaswamy Umapathy, Christian Roos, Jean P Boubli, Jeffrey Rogers, Kyle Kai-How Farh, Tomas Marques Bonet

Faculty, Staff and Students Publications

The rich diversity of morphology and behavior displayed across primate species provides an informative context in which to study the impact of genomic diversity on fundamental biological processes. Analysis of that diversity provides insight into long-standing questions in evolutionary and conservation biology and is urgent given severe threats these species are facing. Here, we present high-coverage whole-genome data from 233 primate species representing 86% of genera and all 16 families. This dataset was used, together with fossil calibration, to create a nuclear DNA phylogeny and to reassess evolutionary divergence times among primate clades. We found within-species genetic diversity across families …

Genome-Wide Coancestry Reveals Details Of Ancient And Recent Male-Driven Reticulation In Baboons, Erik F Sørensen, R Alan Harris, Liye Zhang, Muthuswamy Raveendran, Lukas F K Kuderna, Jerilyn A Walker, Jessica M Storer, Martin Kuhlwilm, Claudia Fontsere, Lakshmi Seshadri, Christina M Bergey, Andrew S Burrell, Juraj Bergman, Jane E Phillips-Conroy, Fekadu Shiferaw, Kenneth L Chiou, Idrissa S Chuma, Julius D Keyyu, Julia Fischer, Marie-Claude Gingras, Sejal Salvi, Harshavardhan Doddapaneni, Mikkel H Schierup, Mark A Batzer, Clifford J Jolly, Sascha Knauf, Dietmar Zinner, Kyle K-H Farh, Tomas Marques-Bonet, Kasper Munch, Christian Roos, Jeffrey Rogers

Faculty, Staff and Students Publications

Baboons (genus Papio) are a morphologically and behaviorally diverse clade of catarrhine monkeys that have experienced hybridization between phenotypically and genetically distinct phylogenetic species. We used high-coverage whole-genome sequences from 225 wild baboons representing 19 geographic localities to investigate population genomics and interspecies gene flow. Our analyses provide an expanded picture of evolutionary reticulation among species and reveal patterns of population structure within and among species, including differential admixture among conspecific populations. We describe the first example of a baboon population with a genetic composition that is derived from three distinct lineages. The results reveal processes, both ancient and …

Phylogenomic Analyses Provide Insights Into Primate Evolution, Yong Shao, Long Zhou, Fang Li, Lan Zhao, Bao-Lin Zhang, Feng Shao, Jia-Wei Chen, Chun-Yan Chen, Xupeng Bi, Xiao-Lin Zhuang, Hong-Liang Zhu, Jiang Hu, Zongyi Sun, Xin Li, Depeng Wang, Iker Rivas-González, Sheng Wang, Yun-Mei Wang, Wu Chen, Gang Li, Hui-Meng Lu, Yang Liu, Lukas F K Kuderna, Kyle Kai-How Farh, Peng-Fei Fan, Li Yu, Ming Li, Zhi-Jin Liu, George P Tiley, Anne D Yoder, Christian Roos, Takashi Hayakawa, Tomas Marques-Bonet, Jeffrey Rogers, Peter D Stenson, David N Cooper, Mikkel Heide Schierup, Yong-Gang Yao, Ya-Ping Zhang, Wen Wang, Xiao-Guang Qi, Guojie Zhang, Dong-Dong Wu

Faculty, Staff and Students Publications

Comparative analysis of primate genomes within a phylogenetic context is essential for understanding the evolution of human genetic architecture and primate diversity. We present such a study of 50 primate species spanning 38 genera and 14 families, including 27 genomes first reported here, with many from previously less well represented groups, the New World monkeys and the Strepsirrhini. Our analyses reveal heterogeneous rates of genomic rearrangement and gene evolution across primate lineages. Thousands of genes under positive selection in different lineages play roles in the nervous, skeletal, and digestive systems and may have contributed to primate innovations and adaptations. Our …

The Clinical And Molecular Spectrum Of The Kdm6b-Related Neurodevelopmental Disorder, Dmitrijs Rots, Taryn E Jakub, Crystal Keung, Adam Jackson, Siddharth Banka, Rolph Pfundt, Bert B A De Vries, Richard H Van Jaarsveld, Saskia M J Hopman, Ellen Van Binsbergen, Irene Valenzuela, Maja Hempel, Tatjana Bierhals, Fanny Kortüm, Francois Lecoquierre, Alice Goldenberg, Jens Michael Hertz, Charlotte Brasch Andersen, Maria Kibæk, Eloise J Prijoles, Roger E Stevenson, David B Everman, Wesley G Patterson, Linyan Meng, Charul Gijavanekar, Karl De Dios, Shenela Lakhani, Tess Levy, Matias Wagner, Dagmar Wieczorek, Paul J Benke, María Soledad Lopez Garcia, Renee Perrier, Sergio B Sousa, Pedro M Almeida, Maria José Simões, Bertrand Isidor, Wallid Deb, Andrew A Schmanski, Omar Abdul-Rahman, Christophe Philippe, Ange-Line Bruel, Laurence Faivre, Antonio Vitobello, Christel Thauvin, Jeroen J Smits, Livia Garavelli, Stefano G Caraffi, Francesca Peluso, Laura Davis-Keppen, Dylan Platt, Erin Royer, Lisette Leeuwen, Margje Sinnema, Alexander P A Stegmann, Constance T R M Stumpel, George E Tiller, Daniëlle G M Bosch, Stephanus T Potgieter, Shelagh Joss, Miranda Splitt, Simon Holden, Matina Prapa, Nicola Foulds, Sofia Douzgou, Kaija Puura, Regina Waltes, Andreas G Chiocchetti, Christine M Freitag, F Kyle Satterstrom, Silvia De Rubeis, Joseph Buxbaum, Bruce D Gelb, Aleksic Branko, Itaru Kushima, Jennifer Howe, Stephen W Scherer, Alessia Arado, Chiara Baldo, Olivier Patat, Demeer Bénédicte, Diego Lopergolo, Filippo M Santorelli, Tobias B Haack, Andreas Dufke, Miriam Bertrand, Ruth J Falb, Angelika Rieß, Peter Krieg, Stephanie Spranger, Maria Francesca Bedeschi, Maria Iascone, Sarah Josephi-Taylor, Tony Roscioli, Michael F Buckley, Jan Liebelt, Aditi I Dagli, Emmelien Aten, Anna C E Hurst, Alesha Hicks, Mohnish Suri, Ermal Aliu, Sunil Naik, Richard Sidlow, Juliette Coursimault, Gaël Nicolas, Hanna Küpper, Florence Petit, Veyan Ibrahim, Deniz Top, Francesca Di Cara, Raymond J Louie, Elliot Stolerman, Han G Brunner, Lisenka E L M Vissers, Jamie M Kramer, Tjitske Kleefstra

Faculty, Staff and Students Publications

De novo variants are a leading cause of neurodevelopmental disorders (NDDs), but because every monogenic NDD is different and usually extremely rare, it remains a major challenge to understand the complete phenotype and genotype spectrum of any morbid gene. According to OMIM, heterozygous variants in KDM6B cause "neurodevelopmental disorder with coarse facies and mild distal skeletal abnormalities." Here, by examining the molecular and clinical spectrum of 85 reported individuals with mostly de novo (likely) pathogenic KDM6B variants, we demonstrate that this description is inaccurate and potentially misleading. Cognitive deficits are seen consistently in all individuals, but the overall phenotype is …

Using Cancer Proteomics Data To Identify Gene Candidates For Therapeutic Targeting, Diana Monsivais, Sydney E Parks, Darshan S Chandrashekar, Sooryanarayana Varambally, Chad J Creighton

Faculty, Staff and Students Publications

Gene-level associations obtained from mass-spectrometry-based cancer proteomics datasets represent a resource for identifying gene candidates for functional studies. When recently surveying proteomic correlates of tumor grade across multiple cancer types, we identified specific protein kinases having a functional impact on uterine endometrial cancer cells. This previously published study provides just one template for utilizing public molecular datasets to discover potential novel therapeutic targets and approaches for cancer patients. Proteomic profiling data combined with corresponding multi-omics data on human tumors and cell lines can be analyzed in various ways to prioritize genes of interest for interrogating biology. Across hundreds of cancer …

N-Terminal Egfp-Tagging Of Rabbit Apolipoprotein A-I Decreases Expression And Impairs Cholesterol-Efflux Activity, Bradley K Wacker, Lianxiang Bi, Li Liu, Mary G Sorci-Thomas, Philip Ng, Donna J Palmer, Chongren Tang, David A Dichek

Faculty, Staff and Students Publications

No abstract provided.

Defects In Lipid Homeostasis Reflect The Function Of Tango2 In Phospholipid And Neutral Lipid Metabolism, Agustin Leonardo Lujan, Ombretta Foresti, Conor Sugden, Nathalie Brouwers, Alex Mateo Farre, Alessio Vignoli, Mahshid Azamian, Alicia Turner, Jose Wojnacki, Vivek Malhotra

Faculty, Staff and Students Publications

We show that TANGO2 in mammalian cells localizes predominantly to mitochondria and partially at mitochondria sites juxtaposed to lipid droplets (LDs) and the endoplasmic reticulum. HepG2 cells and fibroblasts of patients lacking TANGO2 exhibit enlarged LDs. Quantitative lipidomics revealed a marked increase in lysophosphatidic acid (LPA) and a concomitant decrease in its biosynthetic precursor phosphatidic acid (PA). These changes were exacerbated in nutrient-starved cells. Based on our data, we suggest that TANGO2 function is linked to acyl-CoA metabolism, which is necessary for the acylation of LPA to generate PA. The defect in acyl-CoA availability impacts the metabolism of many other …

Biallelic Variants In Ogdh Encoding Oxoglutarate Dehydrogenase Lead To A Neurodevelopmental Disorder Characterized By Global Developmental Delay, Movement Disorder, And Metabolic Abnormalities, Ella F Whittle, Madison Chilian, Ehsan Ghayoor Karimiani, Helga Progri, Daniela Buhas, Melis Kose, Rebecca D Ganetzky, Mehran Beiraghi Toosi, Paria Najarzadeh Torbati, Reza Shervin Badv, Ivan Shelihan, Hui Yang, Houda Zghal Elloumi, Sukyeong Lee, Yalda Jamshidi, Alan M Pittman, Henry Houlden, Erika Ignatius, Shamima Rahman, Reza Maroofian, Wan Hee Yoon, Christopher J Carroll

Faculty, Staff and Students Publications

PURPOSE: This study aimed to establish the genetic cause of a novel autosomal recessive neurodevelopmental disorder characterized by global developmental delay, movement disorder, and metabolic abnormalities.

METHODS: We performed a detailed clinical characterization of 4 unrelated individuals from consanguineous families with a neurodevelopmental disorder. We used exome sequencing or targeted-exome sequencing, cosegregation, in silico protein modeling, and functional analyses of variants in HEK293 cells and Drosophila melanogaster, as well as in proband-derived fibroblast cells.

RESULTS: In the 4 individuals, we identified 3 novel homozygous variants in oxoglutarate dehydrogenase (OGDH) (NM_002541.3), which encodes a subunit of the tricarboxylic acid cycle enzyme …

Brain Monoamine Vesicular Transport Disease Caused By Homozygous Slc18a2 Variants: A Study In 42 Affected Individuals, Ken Saida, Reza Maroofian, Toru Sengoku, Tadahiro Mitani, Alistair T Pagnamenta, Dana Marafi, Maha S Zaki, Thomas J O'Brien, Ehsan Ghayoor Karimiani, Rauan Kaiyrzhanov, Marina Takizawa, Sachiko Ohori, Huey Yin Leong, Gulsen Akay, Hamid Galehdari, Mina Zamani, Ratna Romy, Christopher J Carroll, Mehran Beiraghi Toosi, Farah Ashrafzadeh, Shima Imannezhad, Hadis Malek, Najmeh Ahangari, Hoda Tomoum, Vykuntaraju K Gowda, Varunvenkat M Srinivasan, David Murphy, Natalia Dominik, Hasnaa M Elbendary, Karima Rafat, Sanem Yilmaz, Seda Kanmaz, Mine Serin, Deepa Krishnakumar, Alice Gardham, Anna Maw, Tekki Sreenivasa Rao, Sarah Alsubhi, Myriam Srour, Daniela Buhas, Tamison Jewett, Rachel E Goldberg, Hanan Shamseldin, Eirik Frengen, Doriana Misceo, Petter Strømme, José Ricardo Magliocco Ceroni, Chong Ae Kim, Gozde Yesil, Esma Sengenc, Serhat Guler, Mariam Hull, Mered Parnes, Dilek Aktas, Banu Anlar, Yavuz Bayram, Davut Pehlivan, Jennifer E Posey, Shahryar Alavi, Seyed Ali Madani Manshadi, Hamad Alzaidan, Mohammad Al-Owain, Lama Alabdi, Ferdous Abdulwahab, Futoshi Sekiguchi, Kohei Hamanaka, Atsushi Fujita, Yuri Uchiyama, Takeshi Mizuguchi, Satoko Miyatake, Noriko Miyake, Reem M Elshafie, Kamran Salayev, Ulviyya Guliyeva, Fowzan S Alkuraya, Joseph G Gleeson, Kristin G Monaghan, Katherine G Langley, Hui Yang, Mahsa Motavaf, Saeid Safari, Mozhgan Alipour, Kazuhiro Ogata, André E X Brown, James R Lupski, Henry Houlden, Naomichi Matsumoto

Faculty, Staff and Students Publications

Purpose: Brain monoamine vesicular transport disease is an infantile-onset movement disorder that mimics cerebral palsy. In 2013, the homozygous SLC18A2 variant, p.Pro387Leu, was first reported as a cause of this rare disorder, and dopamine agonists were efficient for treating affected individuals from a single large family. To date, only 6 variants have been reported. In this study, we evaluated genotype-phenotype correlations in individuals with biallelic SLC18A2 variants.

Methods: A total of 42 affected individuals with homozygous SLC18A2 variant alleles were identified. We evaluated genotype-phenotype correlations and the missense variants in the affected individuals based on the structural modeling of rat …

The Assembled Genome Of The Stroke-Prone Spontaneously Hypertensive Rat, Theodore S Kalbfleisch, Nahla A Hussien Abouel Ela, Kai Li, Wesley A Brashear, Kelli J Kochan, Andrew E Hillhouse, Yaming Zhu, Isha S Dhande, Eric J Kline, Elizabeth A Hudson, Terence D Murphy, Françoise Thibaud-Nissen, Melissa L Smith, Peter A Doris

Faculty, Staff and Student Publications

BACKGROUND: We report the creation and evaluation of a de novo assembly of the genome of the spontaneously hypertensive rat, the most widely used model of human cardiovascular disease.

METHODS: The genome is assembled from long read sequencing (PacBio HiFi and continuous long read data [CLR]) and scaffolded with long-range structural information obtained from Bionano optical maps and proximity ligation sequencing proximity analysis of the genome. The genome assembly was polished with Illumina short reads. Completeness of the assembly was investigated using Benchmarking Universal Single Copy Orthologs analysis. The genome assembly was also evaluated with the rat reference gene set, …

Treatment Of Epilepsy Using A Targeted P38Γ Kinase Gene Therapy, Nicolle Morey, Magdalena Przybyla, Julia Van Der Hoven, Yazi D Ke, Fabien Delerue, Janet Van Eersel, Lars M Ittner

Faculty, Staff and Student Publications

Hyperphosphorylated microtubule-associated protein tau has been implicated in dementia, epilepsy, and other neurological disorders. In contrast, site-specific phosphorylation of tau at threonine 205 (T205) by the kinase p38γ was shown to disengage tau from toxic pathways, serving a neuroprotective function in Alzheimer's disease. Using a viral-mediated gene delivery approach in different mouse models of epilepsy, we show that p38γ activity-enhancing treatment reduces seizure susceptibility, restores neuronal firing patterns, reduces behavioral deficits, and ameliorates epilepsy-induced deaths. Furthermore, we show that p38γ-mediated phosphorylation of tau at T205 is essential for this protection in epilepsy, as a lack of this critical interaction reinstates …

Crispr/Cas13 Effectors Have Differing Extents Of Off-Target Effects That Limit Their Utility In Eukaryotic Cells, Yuxi Ai, Dongming Liang, Jeremy E Wilusz

Faculty, Staff and Students Publications

CRISPR/Cas13 effectors have garnered increasing attention as easily customizable tools for detecting and depleting RNAs of interest. Near perfect complementarity between a target RNA and the Cas13-associated guide RNA is required for activation of Cas13 ribonuclease activity. Nonetheless, the specificity of Cas13 effectors in eukaryotic cells has been debated as the Cas13 nuclease domains can be exposed on the enzyme surface, providing the potential for promiscuous cleavage of nearby RNAs (so-called collateral damage). Here, using co-transfection assays in Drosophila and human cells, we found that the off-target effects of RxCas13d, a commonly used Cas13 effector, can be as strong as …

Systematic Expression Profiling Of Dpr And Dip Genes Reveals Cell Surface Codes In Drosophila Larval Motor And Sensory Neurons, Yupu Wang, Meike Lobb-Rabe, James Ashley, Purujit Chatterjee, Veera Anand, Hugo J Bellen, Oguz Kanca, Robert A Carrillo

Faculty, Staff and Students Publications

In complex nervous systems, neurons must identify their correct partners to form synaptic connections. The prevailing model to ensure correct recognition posits that cell-surface proteins (CSPs) in individual neurons act as identification tags. Thus, knowing what cells express which CSPs would provide insights into neural development, synaptic connectivity, and nervous system evolution. Here, we investigated expression of Dpr and DIP genes, two CSP subfamilies belonging to the immunoglobulin superfamily, in Drosophila larval motor neurons (MNs), muscles, glia and sensory neurons (SNs) using a collection of GAL4 driver lines. We found that Dpr genes are more broadly expressed than DIP genes …

Retrospective Genomics Highlights Changes In Genetic Composition Of Tiger Sharks (Galeocerdo Cuvier) And Potential Loss Of A South-Eastern Australia Population, Alice Manuzzi, Belen Jiménez-Mena, Romina Henriques, Bonnie J Holmes, Julian Pepperell, Janette Edson, Mike B Bennett, Charlie Huveneers, Jennifer R Ovenden, Einar E Nielsen

Staff and Researcher Publications

Over the last century, many shark populations have declined, primarily due to overexploitation in commercial, artisanal and recreational fisheries. In addition, in some locations the use of shark control programs also has had an impact on shark numbers. Still, there is a general perception that populations of large ocean predators cover wide areas and therefore their diversity is less susceptible to local anthropogenic disturbance. Here we report on temporal genomic analyses of tiger shark (Galeocerdo cuvier) DNA samples that were collected from eastern Australia over the past century. Using Single Nucleotide Polymorphism (SNP) loci, we documented a significant change in …

Mutations In Hcfc1 And Ronin Result In An Inborn Error Of Cobalamin Metabolism And Ribosomopathy, Tiffany Chern, Annita Achilleos, Xuefei Tong, Matthew C Hill, Alexander B Saltzman, Lucas C Reineke, Arindam Chaudhury, Swapan K Dasgupta, Yushi Redhead, David Watkins, Joel R Neilson, Perumal Thiagarajan, Jeremy B A Green, Anna Malovannaya, James F Martin, David S Rosenblatt, Ross A Poché

Faculty, Staff and Students Publications

Combined methylmalonic acidemia and homocystinuria (cblC) is the most common inborn error of intracellular cobalamin metabolism and due to mutations in Methylmalonic Aciduria type C and Homocystinuria (MMACHC). Recently, mutations in the transcriptional regulators HCFC1 and RONIN (THAP11) were shown to result in cellular phenocopies of cblC. Since HCFC1/RONIN jointly regulate MMACHC, patients with mutations in these factors suffer from reduced MMACHC expression and exhibit a cblC-like disease. However, additional de-regulated genes and the resulting pathophysiology is unknown. Therefore, we have generated mouse models of this disease. In addition to exhibiting loss of Mmachc, metabolic perturbations, and developmental defects previously …

Nrf1 Association With Auts2-Polycomb Mediates Specific Gene Activation In The Brain, Sanxiong Liu, Kimberly A Aldinger, Chi Vicky Cheng, Takae Kiyama, Mitali Dave, Hanna K Mcnamara, Wukui Zhao, James M Stafford, Nicolas Descostes, Pedro Lee, Stefano G Caraffi, Ivan Ivanovski, Edoardo Errichiello, Christiane Zweier, Orsetta Zuffardi, Michael Schneider, Antigone S Papavasiliou, M Scott Perry, Jennifer Humberson, Megan T Cho, Astrid Weber, Andrew Swale, Tudor C Badea, Chai-An Mao, Livia Garavelli, William B Dobyns, Danny Reinberg

Faculty, Staff and Student Publications

The heterogeneous family of complexes comprising Polycomb repressive complex 1 (PRC1) is instrumental for establishing facultative heterochromatin that is repressive to transcription. However, two PRC1 species, ncPRC1.3 and ncPRC1.5, are known to comprise novel components, AUTS2, P300, and CK2, that convert this repressive function to that of transcription activation. Here, we report that individuals harboring mutations in the HX repeat domain of AUTS2 exhibit defects in AUTS2 and P300 interaction as well as a developmental disorder reflective of Rubinstein-Taybi syndrome, which is mainly associated with a heterozygous pathogenic variant in CREBBP/EP300. Moreover, the absence of AUTS2 or mutation in its …

Multiplexed Drug-Based Selection And Counterselection Genetic Manipulations In Drosophila, Nick Matinyan, Mansi S Karkhanis, Yezabel Gonzalez, Antrix Jain, Alexander Saltzman, Anna Malovannaya, Alejandro Sarrion-Perdigones, Herman A Dierick, Koen J T Venken

Faculty, Staff and Students Publications

The power of Drosophila melanogaster as a model system relies on tractable germline genetic manipulations. Despite Drosophila's expansive genetics toolbox, such manipulations are still accomplished one change at a time and depend predominantly on phenotypic screening. We describe a drug-based genetic platform consisting of four selection and two counterselection markers, eliminating the need to screen for modified progeny. These markers work reliably individually or in combination to produce specific genetic outcomes. We demonstrate three example applications of multiplexed drug-based genetics by generating (1) transgenic animals, expressing both components of binary overexpression systems in a single transgenesis step; (2) dual selectable …

Discovery And Characterization Of Bromodomain 2-Specific Inhibitors Of Brdt, Zhifeng Yu, Angela F Ku, Justin L Anglin, Rajesh Sharma, Melek Nihan Ucisik, John C Faver, Feng Li, Pranavanand Nyshadham, Nicholas Simmons, Kiran L Sharma, Sureshbabu Nagarajan, Kevin Riehle, Gundeep Kaur, Banumathi Sankaran, Marta Storl-Desmond, Stephen S Palmer, Damian W Young, Choel Kim, Martin M Matzuk

Faculty, Staff and Students Publications

Bromodomain testis (BRDT), a member of the bromodomain and extraterminal (BET) subfamily that includes the cancer targets BRD2, BRD3, and BRD4, is a validated contraceptive target. All BET subfamily members have two tandem bromodomains (BD1 and BD2). Knockout mice lacking BRDT-BD1 or both bromodomains are infertile. Treatment of mice with JQ1, a BET BD1/BD2 nonselective inhibitor with the highest affinity for BRD4, disrupts spermatogenesis and reduces sperm number and motility. To assess the contribution of each BRDT bromodomain, we screened our collection of DNA-encoded chemical libraries for BRDT-BD1 and BRDT-BD2 binders. High-enrichment hits were identified and resynthesized off-DNA and examined …

Phenytoin Inhibits Cell Proliferation Through Microrna-196a-5p In Mouse Lip Mesenchymal Cells, Hiroki Yoshioka, Sai Shankar Ramakrishnan, Akiko Suzuki, Junichi Iwata

Faculty, Staff and Student Publications

Cleft lip (CL) is one of the most common birth defects. It is caused by either genetic mutations or environmental factors. Recent studies suggest that environmental factors influence the expression of noncoding RNAs [e.g., microRNA (miRNA)], which can regulate the expression of genes crucial for cellular functions. In this study, we examined which miRNAs are associated with CL. Among 10 candidate miRNAs (miR-98-3p, miR-101a-3p, miR-101b-3p, miR-141-3p, miR-144-3p, miR-181a-5p, miR-196a-5p, miR-196b-5p, miR-200a-3p, and miR-710) identified through our bioinformatic analysis of CL-associated genes, overexpression of miR-181a-5p, miR-196a-5p, miR-196b-5p, and miR-710 inhibited cell proliferation through suppression of genes associated with CL in cultured …

Evidence For Craniofacial Enhancer Variation Underlying Nonsyndromic Cleft Lip And Palate, Vershanna E Morris, S Shahrukh Hashmi, Lisha Zhu, Lorena Maili, Christian Urbina, Steven Blackwell, Matthew R Greives, Edward P Buchanan, John B Mulliken, Susan H Blanton, W Jim Zheng, Jacqueline T Hecht, Ariadne Letra

Faculty, Staff and Student Publications

Nonsyndromic cleft lip with or without cleft palate (NSCLP) is a common birth defect for which only ~ 20% of the underlying genetic variation has been identified. Variants in noncoding regions have been increasingly suggested to contribute to the missing heritability. In this study, we investigated whether variation in craniofacial enhancers contributes to NSCLP. Candidate enhancers were identified using VISTA Enhancer Browser and previous publications. Prioritization was based on patterning defects in knockout mice, deletion/duplication of craniofacial genes in animal models and results of whole exome/whole genome sequencing studies. This resulted in 20 craniofacial enhancers to be investigated. Custom amplicon-based …

Aav-Crispr Gene Editing Is Negated By Pre-Existing Immunity To Cas9, Ang Li, Mark R Tanner, Ciaran M Lee, Ayrea E Hurley, Marco De Giorgi, Kelsey E Jarrett, Timothy H Davis, Alexandria M Doerfler, Gang Bao, Christine Beeton, William R Lagor

Faculty, Staff and Students Publications

Adeno-associated viral (AAV) vectors are a leading candidate for the delivery of CRISPR-Cas9 for therapeutic genome editing in vivo. However, AAV-based delivery involves persistent expression of the Cas9 nuclease, a bacterial protein. Recent studies indicate a high prevalence of neutralizing antibodies and T cells specific to the commonly used Cas9 orthologs from Streptococcus pyogenes (SpCas9) and Staphylococcus aureus (SaCas9) in humans. We tested in a mouse model whether pre-existing immunity to SaCas9 would pose a barrier to liver genome editing with AAV packaging CRISPR-Cas9. Although efficient genome editing occurred in mouse liver with pre-existing SaCas9 immunity, this was accompanied by …

Integrating Mouse And Human Genetic Data To Move Beyond Gwas And Identify Causal Genes In Cholesterol Metabolism, Zhonggang Li, James A Votava, Gregory J M Zajac, Jenny N Nguyen, Fernanda B Leyva Jaimes, Sophia M Ly, Jacqueline A Brinkman, Marco De Giorgi, Sushma Kaul, Cara L Green, Samantha L St Clair, Sabrina L Belisle, Julia M Rios, David W Nelson, Mary G Sorci-Thomas, William R Lagor, Dudley W Lamming, Chi-Liang Eric Yen, Brian W Parks

Faculty, Staff and Students Publications

Identifying the causal gene(s) that connects genetic variation to a phenotype is a challenging problem in genome-wide association studies (GWASs). Here, we develop a systematic approach that integrates mouse liver co-expression networks with human lipid GWAS data to identify regulators of cholesterol and lipid metabolism. Through our approach, we identified 48 genes showing replication in mice and associated with plasma lipid traits in humans and six genes on the X chromosome. Among these 54 genes, 25 have no previously identified role in lipid metabolism. Based on functional studies and integration with additional human lipid GWAS datasets, we pinpoint Sestrin1 as …

The Zinc Transporter Zipt-7.1 Regulates Sperm Activation In Nematodes, Yanmei Zhao, Chieh-Hsiang Tan, Amber Krauchunas, Andrea Scharf, Nicholas Dietrich, Kurt Warnhoff, Zhiheng Yuan, Marina Druzhinina, Sam Guoping Gu, Long Miao, Andrew Singson, Ronald E Ellis, Kerry Kornfeld

Rowan-Virtua School of Osteopathic Medicine Departmental Research

Sperm activation is a fascinating example of cell differentiation, in which immotile spermatids undergo a rapid and dramatic transition to become mature, motile sperm. Because the sperm nucleus is transcriptionally silent, this transition does not involve transcriptional changes. Although Caenorhabditis elegans is a leading model for studies of sperm activation, the mechanisms by which signaling pathways induce this transformation remain poorly characterized. Here we show that a conserved transmembrane zinc transporter, ZIPT-7.1, regulates the induction of sperm activation in Caenorhabditis nematodes. The zipt-7.1 mutant hermaphrodites cannot self-fertilize, and males reproduce poorly, because mutant spermatids are defective in responding to activating …

Persistent Stress-Induced Neuroplastic Changes In The Locus Coeruleus/Norepinephrine System, Olga Borodovitsyna, Neal Joshi, Daniel Chandler

Rowan-Virtua School of Osteopathic Medicine Departmental Research

Neural plasticity plays a critical role in mediating short- and long-term brain responses to environmental stimuli. A major effector of plasticity throughout many regions of the brain is stress. Activation of the locus coeruleus (LC) is a critical step in mediating the neuroendocrine and behavioral limbs of the stress response. During stressor exposure, activation of the hypothalamic-pituitary-adrenal axis promotes release of corticotropin-releasing factor in LC, where its signaling promotes a number of physiological and cellular changes. While the acute effects of stress on LC physiology have been described, its long-term effects are less clear. This review will describe how stress …