Medical Specialties Commons^™

Multiple Quality Control Checkpoints Safeguard Small Nuclear Rna Biogenesis And Prevent Assembly Of Aberrant Spliceosomes, Tiantai Ma, Claire Huntington, Zhuoyi Song, Rea M Lardelli, Eric L Van Nostrand, Jens Lykke-Andersen

Faculty, Staff and Students Publications

Defective small nuclear (sn)RNAs are produced from hundreds of human snRNA pseudogenes and mutant snRNA genes associated with human developmental disorders. Machineries that prevent defective snRNAs from disrupting pre-mRNA splicing remain poorly defined. Here, we identify multiple checkpoints in snRNA biogenesis monitored by quality control machineries that subject defective snRNAs to degradation and prevent their assembly into spliceosomes. We show that variant U1 snRNAs produced from human pseudogenes, some at rates approaching canonical snRNAs, are impaired in 3' cleavage and targeted for degradation by the NEXT-exosome while failures in subsequent protein assembly steps promote NEXT-exosome- or terminal uridylyl transferase 4/7-mediated …

Notch2 Vus Interpretation In Pediatric Cholestasis: A 365-Variant Landscape And Genotype-Phenotype Analysis Of 7 Cases, Brett J. Hoskins, Tiziano Pramparo, Shannon M. Vandriel, Chaowapong Jarasvaraparn, Michael J. Wilsey, Voytek Slowik, Lakshmi Kunam, Janis M. Stoll, Ruben E. Quiros-Tejeira, Simon Lam, Wikrom Karnsakul

Manuscripts, Articles, Book Chapters and Other Papers

Background/Objectives: Interpretation of NOTCH2 variants of uncertain significance (VUSs) remains challenging in pediatric cholestasis and suspected Alagille syndrome (ALGS) due to incomplete penetrance, phenotypic variability, and extensive genetic variation. Although disease-specific variant classification criteria have identified pathogenic hotspots within epidermal growth factor (EGF)-like repeats 8-12 and ankyrin repeat domains 3-6, characterizing these VUSs requires a robust framework. Methods: We reviewed aggregate deidentified data from 519 multigene cholestasis panel entries and evaluated genotype-phenotype data from a consortium cohort of 7 pediatric patients with heterozygous NOTCH2 VUSs using a clinical-genomic prioritization framework informed by disease-specific ACMG criteria and domain location. …

Expanding The Clinical And Molecular Spectrum Of Tubb2b Through Distinct Variants Identified Across Multiple Families, Haowei Du, Lauren E Westerfield, Chloe Munderloh, Divya Kalra, Yifan Wu, Yi Chen, Marie-Claude Gingras, Shalini N Jhangiani, Sarenur Yilmaz, Maha S Zaki, Daniel G Calame, Davut Pehlivan, Richard A Gibbs, Richard A Lewis, James R Lupski, Jennifer E Posey

Faculty, Staff and Students Publications

TUBB2B encodes a β-tubulin isotype essential for neuronal proliferation, migration, and organization during brain development. Pathogenic heterozygous variants in TUBB2B are associated with neurodevelopmental disorders including polymicrogyria and corpus callosum abnormalities. However, the phenotypic spectrum remains heterogeneous, most likely reflecting variant-specific effects on microtubule formation and stability. Homozygous TUBB2B variants are exceedingly rare, with one family reported to date. We describe five individuals in four families with rare TUBB2B variants. Four variants are described, including a previously reported de novo missense variant, c.292G>A (GenBank: NM_178012.5) (p.Gly98Arg), with potential phenotypic expansion including panhypopituitarism; a previously reported de novo missense variant, …

Expanding The Clinical And Molecular Spectrum Of Tubb2b Through Distinct Variants Identified Across Multiple Families, Haowei Du, Lauren E Westerfield, Chloe Munderloh, Divya Kalra, Yifan Wu, Yi Chen, Marie-Claude Gingras, Shalini N Jhangiani, Sarenur Yilmaz, Maha S Zaki, Daniel G Calame, Davut Pehlivan, Richard A Gibbs, Richard A Lewis, James R Lupski, Jennifer E Posey

Faculty, Staff and Students Publications

TUBB2B encodes a β-tubulin isotype essential for neuronal proliferation, migration, and organization during brain development. Pathogenic heterozygous variants in TUBB2B are associated with neurodevelopmental disorders including polymicrogyria and corpus callosum abnormalities. However, the phenotypic spectrum remains heterogeneous, most likely reflecting variant-specific effects on microtubule formation and stability. Homozygous TUBB2B variants are exceedingly rare, with one family reported to date. We describe five individuals in four families with rare TUBB2B variants. Four variants are described, including a previously reported de novo missense variant, c.292G>A (GenBank: NM_178012.5) (p.Gly98Arg), with potential phenotypic expansion including panhypopituitarism; a previously reported de novo missense variant, …

A Rare Recurring Gain-Of-Function Variant In Bmpr2 Causes Neurodevelopmental Phenotypes In Humans And Flies, Zhi Tan, Ariel Brautbar, Maria Descartes, Erica T Gray, Alejandro Iglesias, Scott D Mclean, Morgan Nutter, Olivia Daugherty, Amber Begtrup, Ingrid M Wentzensen, Yufeng Shen, Damian W Young, Martin M Matzuk, Oguz Kanca, Michael F Wangler, Hugo J Bellen, Wendy K Chung, Shinya Yamamoto

Faculty, Staff and Students Publications

Bone morphogenetic protein receptor type 2 (BMPR2) encodes an evolutionarily conserved serine/threonine kinase that phosphorylates type-1 BMP receptors to mediate intercellular communication upon ligand binding. Loss-of-function variants in BMPR2 are known to cause pulmonary arterial hypertension and other cardiovascular disorders. Here, we report a rare recurrent missense variant in BMPR2 (c.1126G>A [GenBank: NM_001204] [p.Glu376Lys]) identified in six individuals, who all present with neurodevelopmental phenotypes including autism spectrum disorder and global developmental delay. Trio analysis demonstrates de novo occurrence of this variant in at least five of the individuals. Using the fruit fly Drosophila melanogaster, we show that this variant …

Distinct Mutational Landscapes For Germline And Somatic Cancer Variants In Forty Tumor Suppressor Genes, Sharon E Plon

Faculty, Staff and Students Publications

Germline and somatic cancer variants in tumor suppressor genes (TSGs) share loss-of-function mechanisms, but studies of a few genes (DICER1 and CEBPA) have demonstrated differences in variant consequence and location. To systematically assess whether TSGs display distinct mutational patterns, we leveraged large public genetic databases and compared 32,941 high-quality pathogenic/likely pathogenic (P/LP) germline variants in ClinVar, with 12,907 oncogenic/likely oncogenic (O/LO) somatic tumor variants from cBioPortal across 40 TSGs. Only 3,863 (9.2%) variants were shared. Eighteen TSGs showed significantly different distributions of variant occurrences by molecular consequence, replicated with non-overlapping somatic data from the COSMIC database (chi-squared tests, false discovery …

Neurodevelopmental Phenotypes And Brain Anomalies In Individuals With Heterozygous Sema6a Variants, Evan Burchfiel, Xiaonan Zhao, Nichole M Owen, Tia Gordon, Mahshid S Azamian, Eric C Kao, Fan Xia, Xi Luo, Jill A Rosenfeld, Seema R Lalani, Allison P Ortega, Steven B Bleyl, Florence Petit, Sulekha Rajagopolan, Bénédicte Demeer, Meredith K Gillespie, Lijia Huang, Matthew Osmond, Kym M Boycott, Kyra E Stuurman, Marjon A Van Slegtenhorst, Haley Soller, Céline Jost, Aurore Garde, Hana Safraou, Laurence Faivre, Victor Faundes, Daryl A Scott

Faculty, Staff and Students Publications

SEMA6A is a transmembrane protein that plays a role in axon guidance and cell migration. Sema6a null mice have cerebral anatomical defects and altered social interactions and working memory. However, the phenotypes associated with loss of SEMA6A function have not been clearly defined in humans. Here we describe 11 individuals who are heterozygous for putatively damaging variants affecting SEMA6A. All of these individuals (100%) had neurodevelopmental phenotypes that included developmental delay, intellectual disability, and/or autism spectrum disorder. Abnormal behaviors were seen in 73% with oppositional defiant disorder being diagnosed in 27% and acting out, overeating, and tantrums each being described …

Massively Parallel Characterization Of Rna G-Quadruplex Stability And Molecular Recognition, Rebekah L Rothacher, Francisco F Cavazos, Yue Hu, Rhese D Thompson, Anthony M Mustoe, Maria M Aleman, Daniel Dominguez

Faculty, Staff and Students Publications

RNA G-quadruplexes (rG4s) have been implicated as important regulators of RNA metabolism and are promising targets for RNA-targeted therapeutics. rG4s typically require a canonical (G≥2N1-7)4 motif, but the sequence features that affect rG4 stability and recognition by RNA-binding proteins (RBPs) and rG4-binding ligands are not fully understood. To interrogate sequence-level drivers of rG4 folding, we applied a reverse-transcriptase stop sequencing strategy to a library of ∼3000 synthetic rG4s with varied G-tract lengths, loop lengths, and loop compositions, permitting massively parallel quantification of rG4 stability. Our data confirm known sequence-level features and characterize novel combinatorial impacts of these features. We also …

Targeting Oncogenic Flt3 Uncovers A Ferroptosis Vulnerability Through Selenocysteine Recoding In Acute Myeloid Leukaemia, Ken Furudate, Ayumi Kitano, Taotao Tan, Mengdie Feng, Jing Zhou, Tao Wang, Robert J Taylor, Alexandra M Stevens, Md Abul Hassan Samee, Jeffrey A Magee, Koichi Takahashi, Daisuke Nakada

Faculty, Staff and Students Publications

Ferroptosis, an iron-dependent form of cell death driven by lipid peroxidation, has emerged as a potential therapeutic strategy for therapy-resistant cancers. Glutathione peroxidase 4 and the selenoprotein biosynthesis pathway essential for its translation are key regulators of ferroptosis but lack effective therapeutic targeting. In a drug screening using a selenoprotein translation reporter, here we identify FMS-like tyrosine kinase 3 (FLT3) inhibitors as suppressors of selenoprotein translation that induce ferroptosis in FLT3-mutant acute myeloid leukaemia. Mechanistically, FLT3 inhibition disrupts selenocysteine recoding, in which a UGA stop codon is recoded as selenocysteine via the SECIS element and associated binding proteins. Notably, the …

Combining Menin And Mek Inhibition To Target Poor Prognosis Kmt2a-Rearranged Ras Pathway-Mutant Acute Myeloid Leukemia., Nastassja Scheidegger, Constanze Schneider, Gabriela Alexe, Yi-Cheng Wang, Todd A. Alonzo, Allen Basanthakumar, Wallace A. Bourgeois, Julia Dudkiewicz-Garbicz, Delan Khalid, Lucy A. Merickel, Jennifer A. Perry, Rhonda E. Ries, Silvi Salhotra, Audrey Taillon, Alan S. Gamis, Richard Aplenc, Marian H. Harris, Mark Wunderlich, Scott A. Armstrong, Jessica A. Pollard, Soheil Meshinchi, Yana Pikman, Kimberly Stegmaier

Manuscripts, Articles, Book Chapters and Other Papers

KMT2A-rearranged (KMT2A-r) acute leukemias are especially prevalent in the pediatric population. KMT2A-fusion proteins drive leukemogenic gene expression through an interaction with a chromatin complex that includes the scaffold protein menin, giving rise to aggressive acute leukemias. RAS pathway mutations are also common in pediatric leukemia. In a cohort of 1750 patients enrolled on Children's Oncology Group (COG) trials, we identified RAS pathway mutations in 43% of acute myeloid leukemia (AML) cases. The presence of RAS pathway mutations in KMT2A-r AML was associated with a lower complete remission rate, poor event-free survival and overall survival (OS), and early relapses. Given the …

Oncogenic Pik3ca Reprograms Glutamine Metabolism To Drive Bladder Cancer Progression, Jun Hyoung Park, Aaditya Krishna Arun, Sachin B Jorvekar, Yongchao Dou, Mohammed Khurshidul Hassan, Chandra Sekhar Amara, Abu Hena Mostafa Kamal, Chandra Shekar R Ambati, Jun Xu, Daniel Kraushaar, Adriana Langer Gramer, Allison C Bellman, Leomar Y Ballester, Yuan Xu, Robert Taylor Ripley, Seth P Lerner, Roni J Bollag, Martha K Terris, Chong-Xian Pan, Benny Abraham Kaipparettu, Yair Lotan, Nagireddy Putluri

Faculty, Staff and Students Publications

Background: Genomic analysis has revealed that approximately 40% of bladder cancer (BLCA) tumors harbor alterations in the PI3K/AKT pathway, with PIK3CA mutations occurring in 15-25% of cases. PIK3CA, which encodes the catalytic p110α subunit of PI3K, plays a critical role in regulating cell survival, proliferation, and metabolism. However, the metabolic and functional consequences of PIK3CA mutations in BLCA remain poorly defined.

Methods: To investigate the role of PIK3CA mutations in BLCA, we performed targeted sequencing on tumors from patients, identifying recurrent alterations. Using CRISPR/Cas9 knock-in models in SCaBER and UM-UC-3 cell lines, we introduced the PIK3CA E545K mutation to study …

Genetics Of Cerebrotendinous Xanthomatosis

Faculty, Staff and Students Publications

Cerebrotendinous xanthomatosis (CTX) is rare, autosomal recessive inborn error of metabolism caused by biallelic pathogenic variants in CYP27A1, which encodes sterile 27 hydroxylase, a key enzyme in bile acid biosynthesis. Enzyme deficiency results in reduced cholic and chenodeoxycholic acid synthesis with accumulation of cholestanol, bile acid intermediates, and bile alcohols, producing a progressive multisystem disorder characterized by chronic diarrhea, juvenile-onset cataracts, tendons xanthomas, and neurological dysfunction. Although CTX typically begins in childhood, diagnosis is frequently delayed until adulthood, limiting the benefit of effective disease modifying therapy with chenodeoxycholic acid. Since the identification of CYP27A1, more than 200 pathogenic variants have …

Single-Nucleus Profiling Reveals A Core Disease Signature And Cell Type-Specific Vulnerabilities In Early Rett Syndrome, Sih-Rong Wu, Jean-Pierre Revelli, Hu Chen, Zhandong Liu, Huda Y Zoghbi

Faculty, Staff and Students Publications

Rett syndrome (RTT) is an X-linked neurological disorder caused by MECP2 mutations, creating distinct cellular environments in females (mosaic) versus males (nonmosaic). Despite female patients representing most cases, how mosaicism contributes molecularly to RTT pathogenesis, particularly in presymptomatic stages, remains poorly understood. To address this question, we profiled hippocampal transcriptomes of young female and male RTT mice using bulk and single-nucleus RNA sequencing. We identified a core disease signature of consistently dysregulated genes only in MeCP2− cells across RTT models. Moreover, we uncovered non–cell autonomous effects exclusively in female MeCP2+ excitatory neurons, suggesting that these circuits are more vulnerable early …

Disease-Causing Mfn2 Mutants Impair Mitochondrial Fission Dynamics By Distinct Drp1 Dysregulation, Daniel Lagos, Pamela R. De Santiago, Nicolás Pérez-Bravo, Benjamín Cartes-Saavedra, Josefa Vial-Brizzi, Diego Troncoso-Chandía, Oliver Podmanicky, Rita Horvath, Verónica Eisner

Department of Pathology, Anatomy, and Cell Biology Faculty Papers

Mitochondria undergo fusion and fission. While DRP1 regulates fission, fusion is controlled by OPA1, MFN1, and MFN2. The balance between these processes and the crosstalk between machineries remains poorly understood. MFN2 mutations cause Charcot-Marie-Tooth disease type 2 A (CMT2A), affecting mitochondrial fusion and morphology. However, their role in fission is unclear. Using skin fibroblasts from CMT2A patients (L248H and M376V MFN2 mutations) and wild-type mouse embryonic fibroblasts expressing these variants, we studied how MFN2 mutations impact mitochondrial dynamics beyond fusion. We analyzed mitochondrial morphology and dynamics by live-cell confocal microscopy and tested fusion/fission protein levels, oxygen consumption rate (OCR), extracellular …

Phospho-Mimic Βiii-Tubulin Rescues Microtubule And Cardiac Defects In Duchenne Muscular Dystrophy Mice, Delong Zhou, Julie Nouet, Elam Mesa, Vasisht Yegneshwaran, Geovanni Geukgeuzian, Ifeanyichukwu D Adibemma, Edwin Ramirez, Nehal Kamal Ali, Sabine Hilfiker, Hong Li, Tong Liu, Ghassan Yehia, Peter J Romanienko, George G Rodney, Xander H T Wehrens, Paul D Lampe, Robert G Gourdie, Lai-Hua Xie, Diego Fraidenraich

Faculty, Staff and Students Publications

Duchenne muscular dystrophy (DMD) cardiomyopathy is caused by mutations in the dystrophin gene and characterized by profound cytoskeletal disorganization, particularly pathological remodeling of the cardiomyocyte microtubule network. While the role of dystrophin in maintaining sarcolemmal integrity is well-established, its specific association with microtubule dynamics through post-translational modifications (PTMs) remains underinvestigated. Here, we identified βIII-tubulin Ser172 phosphorylation as a specific regulatory residue in cardiac DMD microtubule pathology. Using CRISPR-generated mdx mice carrying a phospho-mimic S172E mutation in βIII-tubulin, we demonstrate rescue of microtubule organization defects, including normalization of orthogonal microtubule misalignment, and restoration of physiological density and polymerization kinetics. Microtubule stabilization …

Non-Isolated Dandy-Walker Malformation: Exome Sequencing Efficacy And Phenotypic Expansions, Seema R Lalani, Daryl A Scott

Faculty, Staff and Students Publications

Dandy-Walker malformation (DWM) is a rare congenital abnormality of the posterior fossa and the cerebellum and has an incidence of 1 in 10 000 to 30 000 births. Although DWM can present in isolation, it is often associated with other central nervous system (CNS) abnormalities or extra-CNS anomalies (DWM+). A molecular cause is not identified in the majority of individuals with DWM+. This is due, in part, to uncertainty regarding optimal testing strategies and an incomplete understanding of the genetic causes of DWM+. In this study, we analyzed clinical exome sequencing (cES) data from 91 individuals with DWM+ to determine …

Non-Isolated Dandy-Walker Malformation: Exome Sequencing Efficacy And Phenotypic Expansions, Sarah Araji, Xiaonan Zhao, Jill A Rosenfeld, Seema R Lalani, Daryl A Scott

Faculty, Staff and Students Publications

Dandy-Walker malformation (DWM) is a rare congenital abnormality of the posterior fossa and the cerebellum and has an incidence of 1 in 10 000 to 30 000 births. Although DWM can present in isolation, it is often associated with other central nervous system (CNS) abnormalities or extra-CNS anomalies (DWM+). A molecular cause is not identified in the majority of individuals with DWM+. This is due, in part, to uncertainty regarding optimal testing strategies and an incomplete understanding of the genetic causes of DWM+. In this study, we analyzed clinical exome sequencing (cES) data from 91 individuals with DWM+ to determine …

Integrated Proteogenomic And Metabolomic Profiling Of Acute Myeloid Leukemias To Identify Molecular Subtypes And Associated Therapy Targets, Jennifer E Kyle, Raghav Jain, Yamei Deng, Marina A Gritsenko, Leanne E Henry, Jonathan T Lei, Yongchao Dou, Bahar Tercan, Zhiao Shi, Mahnoor N Gondal, Chia-Feng Tsai, John M Elizarraras, Rosalie K Chu, Fengchao Yu, Sunil K Joshi, Xiaojun Jing, Daniel A Polasky, Karl K Weitz, Ginny Xiaohe Li, Vanessa L Paurus, Chaevien S Clendinen, Athena A Schepmoes, Priscila M Lalli, Josie G Eder, Javier E Flores, Kelly G Stratton, James C Pino, Camilo Posso, Vladislav A Petyuk, Tyler J Sagendorf, Yuanwei Xu, Omar M Ibrahim, Ronald J Moore, Rui Zhao, Jin Chen, Matthew E Monroe, Mathangi Thiagarajan, Galen Hostetter, Chelsea Newton, Eunkyung An, Ana I Robles, Xu Zhang, Nathan J Edwards, Yin Lu, Hui Zhang, Haitham Abdelhakim, Paul D Piehowski, Mehdi Mesri, Richard D Smith, Chandan Kumar-Sinha, Cristina E Tognon, Jennifer Dunlap, Elie Traer, Li Ding, Jeffrey W Tyner, Arul M Chinnaiyan, Gilbert S Omenn, Karin D Rodland, Saravana M Dhanasekaran, Sara J C Gosline, Alexey I Nesvizhiskii, Bing Zhang, Tao Liu, Marcin P Cieslik, Clinical Proteomic Tumor Analysis Consortium

Faculty, Staff and Students Publications

Acute myeloid leukemia (AML) is a genetically and phenotypically heterogeneous hematological malignancy. Here, to better define this clinically taxing and translationally challenging malignancy, we applied a multiomics approach, consisting of 13 modalities to analyze 173 treatment-naive individuals with AML. By integrating these 'omes', we identified distinct AML subtypes, genotype-phenotype associations, biomarkers and pathobiological mechanisms. Across the spectrum of primitive and committed AML, we found extensive metabolomic and lipidomic reprogramming driven by divergent MYC and mTOR activity. We linked metabolic changes to striking hyperacetylation of mitochondrial proteins in CEBPA-mutant AML. Protein-centric subtyping revealed a distinct NPM1-mutant subset characterized by outlier expression …

Coxfa4l2 Upregulation Preserves Residual Cytochrome C Oxidase Activity In Coxfa4-Related Leigh-Like Encephalopathy, Jiaze Gao, William L Macken, Chiara Pizzamiglio, Renata Kabiljo, Anna Lucia Francavilla, Pauline Gaignard, Antoine Pouzet, Jonathan Levy, Giulia Barcia, Jamie K Leighton, Efstathia Chronopoulou, Germaine Pierre, Riza Köksal Özgül, Ali Dursun, Rebecca Halligan, Helen Mundy, Javeria Raza Alvi, Tipu Sultan, William James Craigen, Lisa Emrick, Jill A Rosenfeld, Gehad Elmakkawy, Jihye Kim, Joseph J Gleeson, Aboulfazl Rad, Gabriela Oprea, Maqbool Hussain, Khalil Ur Rehman, Sadia Riaz, Robert W Taylor, Vincent Procaccio, Maha S Zaki, Erika Fernandez-Vizarra, Ciro Leonardo Pierri, Michael G Hanna, Henry Houlden, Reza Maroofian, Cristina Ugalde, Jan-Willem Taanman, Robert D S Pitceathly

Faculty, Staff and Students Publications

Primary mitochondrial diseases (PMDs) affect approximately 1 in 4300 individuals and cause early-onset neuromuscular and multisystem dysfunction with reduced lifespan. They result from pathogenic variants in mitochondrial or nuclear DNA that impair oxidative phosphorylation. Cytochrome c oxidase (COX; complex IV) deficiency is a well-established cause of PMD, leading to a broad spectrum of phenotypes. COXFA4 (cytochrome c oxidase subunit FA4), formerly NDUFA4, is a nuclear-encoded COX subunit, but its role in disease remains poorly defined. We report the largest genetically confirmed cohort of COXFA4-related PMD to date, comprising 13 individuals from 12 families with biallelic pathogenic COXFA4 variants. All present …

Natural History Of Ngly1 Deficiency: Motor Function & Clinical Features, William F Mueller, Pam Ventola, Regina Deck, Becky Schweighardt, Matt Wilsey, Kevin J Lee, Bernhard Suter

Faculty, Staff and Students Publications

N-glycanase 1 (NGLY1) Deficiency is an ultra-rare neurodevelopmental disorder caused by biallelic loss-of-function mutations in the NGLY1 gene, leading to severe impairments in neurocognitive and motor function abilities in the affected patient population. Its core clinical features include global developmental delay, hyperkinetic movement disorders, elevation of liver transaminases, (hypo)alacrima, and progressive sensorimotor neuropathy. Due to the range of phenotypes and severity within the confirmed patient population, ongoing characterization of the disease is critical. A prospective natural history study (NHS) was conducted to further elucidate disease phenotypes and examine any changes in neurocognitive or motor function over a one-year period. Fifteen …

Multi-Site Dms Probing Reveals Higher-Order Structure Of Rna-Protein Complexes In Living Cells, Anthony Hoang, Joshua Meehan, Ritwika Bose, David Mitchell, David H Price, Calla M Olson, Chase A Weidmann, Anthony M Mustoe

Faculty, Staff and Students Publications

Identifying tertiary structures and protein binding sites in RNA molecules remains a key challenge in RNA biology. We describe multi-site dimethyl sulfate (DMS)-mutational profiling (MaP) (msDMS-MaP), a strategy that enables simultaneous measurement of RNA secondary, tertiary, and quaternary structures via a single DMS chemical probing experiment. Optimized reverse transcription decodes typically invisible DMS N7-methylguanine (N7-G) modifications via a tautomer-induced mutational signature concurrent with N1 and N3 modifications. We show that N7-G reactivity reports on higher-order RNA structures, revealing key functional motifs such as pseudoknots and protein binding sites. Using msDMS-MaP, we find that E. coli ribosomal RNAs encode numerous independently …

Dolutegravir Developmental Toxicity Is Mitigated By Magnesium And Folate In Zebrafish Embryos, Katheryn A Neugebauer, Daniel A Gorelick

Faculty, Staff and Students Publications

Integrase strand transfer inhibitors have transformed human immunodeficiency virus (HIV) therapy, yet the widely prescribed drug dolutegravir (DTG) has been linked to developmental toxicity, and its teratogenic mechanism remains unclear. Here, we used zebrafish to dissect DTG toxicity during early vertebrate development. DTG exposure from 2-4 h post-fertilization (hpf) to 24 hpf produced high mortality and abnormal morphology. Co-treatment with folates partially restored normal morphology, whereas calcium had no effect. Strikingly, supplementation with magnesium (Mg) partially rescued DTG-exposed embryos, implicating Mg availability in protection. In competitive binding assays, Mg increased binding of folate to purified folate receptor (FOLR1) by 30% …

Expanding The Mutation Spectrum Of Non-Syndromic Retinitis Pigmentosa In Consanguineous Pakistani Families: Unraveling Novel Pathogenic Variants In Rp1, Pde6b, And Prcd Genes For Precision Diagnosis, Asad Ullah, Asfandyar Ahmad Khan, Yumei Li, Meng Wang, Raeesa Tehreem, Amtul Aziz, Kiran Afshan, Rui Chen, Sabika Firasat

Faculty, Staff and Students Publications

No abstract provided.

Rhesus Macaques With An Opa1 Mutation Demonstrate Features Of Autosomal Dominant Optic Atrophy, Brett D Story, Jun Wang, Rui Chen, Lucy G Moore, Leandro B C Teixeira, Jaeho Shim, Ana C Raposo, Maria Isabel Casanova, Sophie M Le, Sangwan Park, Laura J Young, Soohyun Kim, Karolina P Roszak, Vanessa Ureno, Paige M Karpinen, Nayeli Echeverria, Monica Ardon, Brian C Leonard, Marguerite Knipe, Eliza Bliss-Moreau, Brad Fortune, J Timothy Stout, Jeffrey Rogers, Nicholas Marsh-Armstrong, Sara M Thomasy

Faculty, Staff and Students Publications

Autosomal dominant optic atrophy (ADOA) is an inherited optic neuropathy primarily caused by mutations in OPA1. We identified and defined a spontaneous nonhuman primate (NHP) model of ADOA using rhesus macaques heterozygous for a missense mutation (OPA1A8S). With ocular examinations, ophthalmic imaging, electroretinography, histopathology, immunohistochemistry, and transmission electron microscopy (TEM), we documented retinal nerve fiber layer (RNFL) thinning, retinal ganglion cell (RGC) loss and dysfunction, OPA1 mislocalization, and reduced axonal mitochondrial density in affected macaques. Our investigation revealed substantial phenotypic variability among affected macaques, shedding light on the pathogenesis of ADOA. The retinas were evaluated using techniques …

Bi-Allelic Variants In Ap5z1 And Ap5b1 Lead To Retinal Degeneration, Behnoosh Tasharrofi, Yumei Li, Rebecca Lynn Clark, Emma Fale-Olsen, Grace Waldow, Mohammad Keramatipour, Mostafa Asadollahi, Mark E Pennesi, Rui Chen

Faculty, Staff and Students Publications

Inherited retinal diseases (IRDs) comprise a diverse group of disorders that frequently lead to progressive vision impairment and blindness. Despite advances in genetic testing, a significant number of IRD cases remain genetically unsolved, often due to unidentified disease-associated genes or variants. This study reports additional cases for the newly discovered IRD genes of the AP-5 complex. A comprehensive ophthalmological evaluation was performed for all patients, including retinal imaging (multimodal imaging), visual field testing, and electroretinogram (ERG) testing. Whole-genome and -exome sequencing (WGS and WES) were performed for clinically unsolved IRD patients, and data were analyzed to identify underlying causal variants. …

Contributing Factors Of Contralateral Breast Cancer In Patients With Brca Mutation (On-Brca Ii Study, Korea-Bsg 11), Sae Byul Lee, Hyung Seok Park, Eun-Shin Lee, Janghee Lee, Byeongju Kang, Sanghwa Kim, Eun Young Kim, Young-Joon Kang, Sun Young Min, Moohyun Lee, Se-Kang Kim, Hong-Kyu Kim, Chihwan David Cha

Faculty, Staff and Students Publications

Background: The cumulative risk of contralateral breast cancer (CBC) in BRCA1/2 carriers is remarkably higher than in non-carriers. This study aimed to investigate the clinicopathological predictors associated with the risk of metachronous CBC among breast cancer patients with BRCA1/2 mutations using correspondence analysis.

Methods: This retrospective study included patients who underwent BRCA testing between January 2008 and December 2018. The inclusion criterion was patients being between 20 and 80-years-old with invasive breast cancer (pT1-3, N0-3). We performed univariate and multivariate survival analyses to assess the risks associated with BRCA mutations, and conducted correspondence analysis to identify contributing factors for CBC …

Reduced Catalytic Activity Of Kpc Β-Lactamase Can Increase Ceftazidime Resistance, Shahlo O Solieva, Devin G Kelly, Zhizeng Sun, Gregory Bowman, Timothy Palzkill

Faculty, Staff and Students Publications

β-Lactamases confer bacterial resistance by hydrolyzing β-lactam antibiotics. Klebsiella pneumoniae carbapenemases (KPCs) are a major cause of resistance to carbapenems, but combinations of β-lactams with β-lactamase inhibitors, such as ceftazidime/avibactam, initially restored activity against these enzymes. However, treatment has driven the emergence of KPC variants that evade inhibition. Here we investigate the evolutionary and mechanistic consequences of three clinically observed substitutions -D179Y, T243M and H274Y- that together convert the broad-spectrum KPC-2 enzyme into a variant specialized for ceftazidime/avibactam resistance. Using steady-state and pre-steady-state kinetic analyses with ceftazidime, cephalothin and imipenem, we show that T243M enhances cephalosporin hydrolysis, whereas H274Y modulates …

Baseline Parp-1 Pet Imaging In Patients With Advanced Solid Tumors With Dna Damage Response Mutations, Tarek Daoud, Jiansong Chen, Peng Wei, Franklin Wong, Timothy A Yap

Faculty, Staff and Student Publications

Purpose: Inhibitors of poly(ADP-ribose) polymerase (PARP), an enzyme with numerous roles in DNA damage response signaling, represent a class of anti-cancer drugs approved for treating solid tumors with defects in the DNA damage response. However, additional methods for identifying patients who would benefit from the use of PARP inhibitors are urgently needed. We evaluated a novel radiotracer, 18F-FluorThanatrace ([18F]-FTT), to noninvasively assess PARP activity with PET/CT before treatment and determine associations between uptake, tumor mutation status, and prior receipt of therapy including PARP inhibitors.

Methods: Fifty-two patients with solid tumors underwent whole-body (skull base to thigh) [18F]-FTT PET/CT scans before …

Unraveling The Nexus: Tumor Mutational Burden, Pd-L1 Expression, And Oncogenic Alterations In Non-Small Cell Lung Cancer Cytology Specimens, Min Dai, Francis Anthony San Lucas, Hector Alvarez, Leomar Ballester, Hui Chen, Asif Rashid, Mark J Routbort, Gloria Sura, Keith Sweeney, Gokce Toruner, Peng Wei, Richard Yang, Hyvan Dang, Rajyalakshmi Luthra, Sinchita Roy-Chowdhuri

Faculty, Staff and Student Publications

Background: PD-L1 expression and tumor mutational burden (TMB) are biomarkers for immune checkpoint inhibitor (ICI) therapy in non-small cell lung cancer (NSCLC); however, patients harboring oncogenic alterations have limited benefit from ICIs. The impact of oncogenic alterations on TMB and PD-L1 tumor proportion score in lung cytology specimens is poorly understood. Herein, the association between oncogenic alterations, TMB, and PD-L1 in NSCLC cytology specimens is explored.

Methods: Next-generation sequencing results from 312 NSCLC cytology specimens were retrospectively reviewed that interrogate 610 genes and select immuno-oncology signatures. TMB and PD-L1 immunohistochemical expression across oncogenic alterations were analyzed to explore associations.

Results: …

Ins-17 Acts As A Nutrient Deprivation Signal To Mediate Adult Iis-Regulated Associative Behaviors In C. Elegans, Meredith I Williams, Rachana Rajupalem, Rachel N Arey

Faculty, Staff and Students Publications

Insulin/Insulin-like growth factor 1 (IGF-1) signaling (IIS) is a pleiotropic signaling pathway that functions across tissues to coordinate phenotypic changes in response to nutrient status. Thus, the ubiquity of the IIS pathway hinders efforts to elucidate the mechanisms driving specific IIS-related phenotypes. Previous research in the nematode worm C. elegans has demonstrated that loss of function of the IIS transmembrane receptor (IR) ortholog, DAF-2, results in a doubled lifespan and enhanced learning and memory behaviors in young and aged animals. However, these findings are the result of reducing DAF-2 receptor function rather than modulating ligand-receptor interactions. In the current study, …