The olfactory bulb (OB) receives odor information from the olfactory epithelium and relays this to the olfactory cortex. Using a mouse model, we found that development and maturation of OB interneurons depends on the zinc finger homeodomain factor teashirt zinc finger family member 1 (TSHZ1). In mice lacking TSHZ1, neuroblasts exhibited a normal tangential migration to the OB; however, upon arrival to the OB, the neuroblasts were distributed aberrantly within the radial dimension, and many immature neuroblasts failed to exit the rostral migratory stream. Conditional deletion of
Daniela Ragancokova, Elena Rocca, Anne M.M. Oonk, Herbert Schulz, Elvira Rohde, Jan Bednarsch, Ilse Feenstra, Ronald J.E. Pennings, Hagen Wende, Alistair N. Garratt
Aniridia is a congenital and progressive panocular condition with poor visual prognosis that is associated with brain, olfactory, and pancreatic abnormalities. Development of aniridia is linked with nonsense mutations that result in paired box 6 (
Cheryl Y. Gregory-Evans, Xia Wang, Kishor M. Wasan, Jinying Zhao, Andrew L. Metcalfe, Kevin Gregory-Evans
A systems pharmacological approach that capitalizes on the characterization of intracellular signaling networks can transform our understanding of human diseases and lead to therapy development. Here, we applied this strategy to identify pharmacological targets for the treatment of Stargardt disease, a severe juvenile form of macular degeneration. Diverse GPCRs have previously been implicated in neuronal cell survival, and crosstalk between GPCR signaling pathways represents an unexplored avenue for pharmacological intervention. We focused on this receptor family for potential therapeutic interventions in macular disease. Complete transcriptomes of mouse and human samples were analyzed to assess the expression of GPCRs in the retina. Focusing on adrenergic (AR) and serotonin (5-HT) receptors, we found that adrenoceptor α 2C (
Yu Chen, Grazyna Palczewska, Debarshi Mustafi, Marcin Golczak, Zhiqian Dong, Osamu Sawada, Tadao Maeda, Akiko Maeda, Krzysztof Palczewski
Recombinant adeno-associated virus (rAAV) vectors have shown promise for the treatment of several diseases; however, immune-mediated elimination of transduced cells has been suggested to limit and account for a loss of efficacy. To determine whether rAAV vector expression can persist long term, we administered rAAV vectors expressing normal, M-type α-1 antitrypsin (M-AAT) to AAT-deficient subjects at various doses by multiple i.m. injections. M-specific AAT expression was observed in all subjects in a dose-dependent manner and was sustained for more than 1 year in the absence of immune suppression. Muscle biopsies at 1 year had sustained AAT expression and a reduction of inflammatory cells compared with 3 month biopsies. Deep sequencing of the TCR Vβ region from muscle biopsies demonstrated a limited number of T cell clones that emerged at 3 months after vector administration and persisted for 1 year. In situ immunophenotyping revealed a substantial Treg population in muscle biopsy samples containing AAT-expressing myofibers. Approximately 10% of all T cells in muscle were natural Tregs, which were activated in response to AAV capsid. These results suggest that i.m. delivery of rAAV type 1–AAT (rAAV1-AAT) induces a T regulatory response that allows ongoing transgene expression and indicates that immunomodulatory treatments may not be necessary for rAAV-mediated gene therapy.
Christian Mueller, Jeffrey D. Chulay, Bruce C. Trapnell, Margaret Humphries, Brenna Carey, Robert A. Sandhaus, Noel G. McElvaney, Louis Messina, Qiushi Tang, Farshid N. Rouhani, Martha Campbell-Thompson, Ann Dongtao Fu, Anthony Yachnis, David R. Knop, Guo-jie Ye, Mark Brantly, Roberto Calcedo, Suryanarayan Somanathan, Lee P. Richman, Robert H. Vonderheide, Maigan A. Hulme, Todd M. Brusko, James M. Wilson, Terence R. Flotte
Nuclear factor-κB (NF-κB) regulates cellular responses to inflammation and aging, and alterations in NF-κB signaling underlie the pathogenesis of multiple human diseases. Effective clinical therapeutics targeting this pathway remain unavailable. In primary human keratinocytes, we found that hypochlorite (HOCl) reversibly inhibited the expression of
Thomas H. Leung, Lillian F. Zhang, Jing Wang, Shoucheng Ning, Susan J. Knox, Seung K. Kim
Approximately 90% of patients with isolated agammaglobulinemia and failure of B cell development have mutations in genes required for signaling through the pre–B cell and B cell receptors. The nature of the gene defect in the majority of remaining patients is unknown. We recently identified 4 patients with agammaglobulinemia and markedly decreased numbers of peripheral B cells. The B cells that could be detected had an unusual phenotype characterized by the increased expression of CD19 but the absence of a B cell receptor. Genetic studies demonstrated that all 4 patients had the exact same de novo mutation in the broadly expressed transcription factor E47. The mutant protein (E555K) was stable in patient-derived EBV-transformed cell lines and cell lines transfected with expression vectors. E555K in the transfected cells localized normally to the nucleus and resulted in a dominant negative effect when bound to DNA as a homodimer with wild-type E47. Mutant E47 did permit DNA binding by a tissue-specific heterodimeric DNA-binding partner, myogenic differentiation 1 (MYOD). These findings document a mutational hot-spot in E47 and represent an autosomal dominant form of agammaglobulinemia. Further, they indicate that E47 plays a critical role in enforcing the block in development of B cell precursors that lack functional antigen receptors.
Bertrand Boisson, Yong-Dong Wang, Amma Bosompem, Cindy S. Ma, Annick Lim, Tatiana Kochetkov, Stuart G. Tangye, Jean-Laurent Casanova, Mary Ellen Conley
Lumbar disc degeneration (LDD) is associated with both genetic and environmental factors and affects many people worldwide. A hallmark of LDD is loss of proteoglycan and water content in the nucleus pulposus of intervertebral discs. While some genetic determinants have been reported, the etiology of LDD is largely unknown. Here we report the findings from linkage and association studies on a total of 32,642 subjects consisting of 4,043 LDD cases and 28,599 control subjects. We identified carbohydrate sulfotransferase 3 (
You-Qiang Song, Tatsuki Karasugi, Kenneth M.C. Cheung, Kazuhiro Chiba, Daniel W.H. Ho, Atsushi Miyake, Patrick Y.P. Kao, Kit Ling Sze, Anita Yee, Atsushi Takahashi, Yoshiharu Kawaguchi, Yasuo Mikami, Morio Matsumoto, Daisuke Togawa, Masahiro Kanayama, Dongquan Shi, Jin Dai, Qing Jiang, Chengai Wu, Wei Tian, Na Wang, John C.Y. Leong, Keith K.D. Luk, Shea-ping Yip, Stacey S. Cherny, Junwen Wang, Stefan Mundlos, Anthi Kelempisioti, Pasi J. Eskola, Minna Männikkö, Pirkka Mäkelä, Jaro Karppinen, Marjo-Riitta Järvelin, Paul F. O’Reilly, Michiaki Kubo, Tomoatsu Kimura, Toshikazu Kubo, Yoshiaki Toyama, Hiroshi Mizuta, Kathryn S.E. Cheah, Tatsuhiko Tsunoda, Pak-Chung Sham, Shiro Ikegawa, Danny Chan
Anorexia nervosa and bulimia nervosa are common and severe eating disorders (EDs) of unknown etiology. Although genetic factors have been implicated in the psychopathology of EDs, a clear biological pathway has not been delineated. DNA from two large families affected by EDs was collected, and mutations segregating with illness were identified by whole-genome sequencing following linkage mapping or by whole-exome sequencing. In the first family, analysis of twenty members across three generations identified a rare missense mutation in the estrogen-related receptor α (
Huxing Cui, Jarrette Moore, Sunbola S. Ashimi, Brittany L. Mason, Jordan N. Drawbridge, Shizhong Han, Benjamin Hing, Abigail Matthews, Carrie J. McAdams, Benjamin W. Darbro, Andrew A. Pieper, David A. Waller, Chao Xing, Michael Lutter
The autosomal dominantly inherited east Texas bleeding disorder is linked to an
Lisa M. Vincent, Sinh Tran, Ruzica Livaja, Tracy A. Bensend, Dianna M. Milewicz, Björn Dahlbäck
Progressive osseous heteroplasia (POH) is a rare developmental disorder of heterotopic ossification (HO) caused by heterozygous inactivating germline mutations in the paternal allele of the
Dana M. Cairns, Robert J. Pignolo, Tomoya Uchimura, Tracy A. Brennan, Carter M. Lindborg, Meiqi Xu, Frederick S. Kaplan, Eileen M. Shore, Li Zeng
For most lysosomal storage diseases (LSDs) affecting the CNS, there is currently no cure. The BBB, which limits the bioavailability of drugs administered systemically, and the short half-life of lysosomal enzymes, hamper the development of effective therapies. Mucopolysaccharidosis type IIIA (MPS IIIA) is an autosomic recessive LSD caused by a deficiency in sulfamidase, a sulfatase involved in the stepwise degradation of glycosaminoglycan (GAG) heparan sulfate. Here, we demonstrate that intracerebrospinal fluid (intra-CSF) administration of serotype 9 adenoassociated viral vectors (AAV9s) encoding sulfamidase corrects both CNS and somatic pathology in MPS IIIA mice. Following vector administration, enzymatic activity increased throughout the brain and in serum, leading to whole body correction of GAG accumulation and lysosomal pathology, normalization of behavioral deficits, and prolonged survival. To test this strategy in a larger animal, we treated beagle dogs using intracisternal or intracerebroventricular delivery. Administration of sulfamidase-encoding AAV9 resulted in transgenic expression throughout the CNS and liver and increased sulfamidase activity in CSF. High-titer serum antibodies against AAV9 only partially blocked CSF-mediated gene transfer to the brains of dogs. Consistently, anti-AAV antibody titers were lower in CSF than in serum collected from healthy and MPS IIIA–affected children. These results support the clinical translation of this approach for the treatment of MPS IIIA and other LSDs with CNS involvement.
Virginia Haurigot, Sara Marcó, Albert Ribera, Miguel Garcia, Albert Ruzo, Pilar Villacampa, Eduard Ayuso, Sònia Añor, Anna Andaluz, Mercedes Pineda, Gemma García-Fructuoso, Maria Molas, Luca Maggioni, Sergio Muñoz, Sandra Motas, Jesús Ruberte, Federico Mingozzi, Martí Pumarola, Fatima Bosch
Single-minded 1 (SIM1) is a basic helix-loop-helix transcription factor involved in the development and function of the paraventricular nucleus of the hypothalamus. Obesity has been reported in
Shwetha Ramachandrappa, Anne Raimondo, Anna M.G. Cali, Julia M. Keogh, Elana Henning, Sadia Saeed, Amanda Thompson, Sumedha Garg, Elena G. Bochukova, Soren Brage, Victoria Trowse, Eleanor Wheeler, Adrienne E. Sullivan, Mehul Dattani, Peter E. Clayton, Vippan Datta, John B. Bruning, Nick J. Wareham, Stephen O’Rahilly, Daniel J. Peet, Ines Barroso, Murray L. Whitelaw, I. Sadaf Farooqi
The DNA-dependent protein kinase catalytic subunit (DNA-PKcs; encoded by
Lisa Woodbine, Jessica A. Neal, Nanda-Kumar Sasi, Mayuko Shimada, Karen Deem, Helen Coleman, William B. Dobyns, Tomoo Ogi, Katheryn Meek, E. Graham Davies, Penny A. Jeggo
The identification of a gain-of-function mutation in
Kapil V. Ramachandran, Jessica A. Hennessey, Adam S. Barnett, Xinhe Yin, Harriett A. Stadt, Erika Foster, Raj A. Shah, Masayuki Yazawa, Ricardo E. Dolmetsch, Margaret L. Kirby, Geoffrey S. Pitt
Adeno-associated virus (AAV) vectors are attractive for gene delivery-based therapeutics, but data from recent clinical trials have indicated that AAV capsids induce a cytotoxic T lymphocyte (CTL) response that eliminates transduced cells. In this study, we used traditional pharmacological agents and AAV mutants to elucidate the pathway of capsid cross-presentation in AAV-permissive cells. Endosomal acidification inhibitors blocked AAV2 antigen presentation by over 90%, while proteasome inhibitors completely abrogated antigen presentation. Using mutant viruses that are defective for nuclear entry, we observed a 90% decrease in capsid antigen presentation. Different antigen presentation efficiencies were achieved by selectively mutating virion nuclear localization signals. Low antigen presentation was demonstrated with basic region 1 (BR1) mutants, despite relatively high transduction efficiency, whereas there was no difference in antigen presentation between BR2 and BR3 mutants defective for transduction, as compared with wild-type AAV2. These results suggest that effective AAV2 capsid antigen presentation is dependent on AAV virion escape from the endosome/lysosome for antigen degradation by proteasomes, but is independent of nuclear uncoating. These results should facilitate the design of effective strategies to evade capsid-specific CTL-mediated elimination of AAV-transduced target cells in future clinical trials.
Chengwen Li, Yi He, Sarah Nicolson, Matt Hirsch, Marc S. Weinberg, Ping Zhang, Tal Kafri, R. Jude Samulski
Ataxia-telangiectasia (A-T) is an autosomal recessive neurodegenerative disorder characterized by radiosensitivity, genomic instability, and predisposition to cancer. A-T is caused by biallelic mutations in the ataxia-telangiectasia mutated (
Andrea Prodosmo, Andrea De Amicis, Cecilia Nisticò, Mario Gabriele, Giuliana Di Rocco, Laura Monteonofrio, Maria Piane, Enrico Cundari, Luciana Chessa, Silvia Soddu
Increased mucus production is a common cause of morbidity and mortality in inflammatory airway diseases, including asthma, chronic obstructive pulmonary disease (COPD), and cystic fibrosis. However, the precise molecular mechanisms for pathogenic mucus production are largely undetermined. Accordingly, there are no specific and effective anti-mucus therapeutics. Here, we define a signaling pathway from chloride channel calcium-activated 1 (CLCA1) to MAPK13 that is responsible for IL-13–driven mucus production in human airway epithelial cells. The same pathway was also highly activated in the lungs of humans with excess mucus production due to COPD. We further validated the pathway by using structure-based drug design to develop a series of novel MAPK13 inhibitors with nanomolar potency that effectively reduced mucus production in human airway epithelial cells. These results uncover and validate a new pathway for regulating mucus production as well as a corresponding therapeutic approach to mucus overproduction in inflammatory airway diseases.
Yael G. Alevy, Anand C. Patel, Arthur G. Romero, Dhara A. Patel, Jennifer Tucker, William T. Roswit, Chantel A. Miller, Richard F. Heier, Derek E. Byers, Tom J. Brett, Michael J. Holtzman
Congenital diarrheal disorders (CDDs) are a collection of rare, heterogeneous enteropathies with early onset and often severe outcomes. Here, we report a family of Ashkenazi Jewish descent, with 2 out of 3 children affected by CDD. Both affected children presented 3 days after birth with severe, intractable diarrhea. One child died from complications at age 17 months. The second child showed marked improvement, with resolution of most symptoms at 10 to 12 months of age. Using exome sequencing, we identified a rare splice site mutation in the DGAT1 gene and found that both affected children were homozygous carriers. Molecular analysis of the mutant allele indicated a total loss of function, with no detectable DGAT1 protein or activity produced. The precise cause of diarrhea is unknown, but we speculate that it relates to abnormal fat absorption and buildup of DGAT substrates in the intestinal mucosa. Our results identify DGAT1 loss-of-function mutations as a rare cause of CDDs. These findings prompt concern for DGAT1 inhibition in humans, which is being assessed for treating metabolic and other diseases.
Joel T. Haas, Harland S. Winter, Elaine Lim, Andrew Kirby, Brendan Blumenstiel, Matthew DeFelice, Stacey Gabriel, David Branski, Carrie A. Grueter, Mauro S. Toporovski, Tobias C. Walther, Mark J. Daly, Robert V. Farese Jr.
The HELLP syndrome is a pregnancy-associated disease inducing hemolysis, elevated liver enzymes, and low platelets in the mother. Although the HELLP symptoms occur in the third trimester in the mother, the origin of the disease can be found in the first trimester fetal placenta. A locus for the HELLP syndrome is present on chromosome 12q23 near PAH. Here, by multipoint nonparametric linkage, pedigree structure allele sharing, and haplotype association analysis of affected sisters and cousins, we demonstrate that the HELLP locus is in an intergenic region on 12q23.2 between PMCH and IGF1. We identified a novel long intergenic noncoding RNA (lincRNA) transcript of 205,012 bases with (peri)nuclear expression in the extravillous trophoblast using strand-specific RT-PCR complemented with RACE and FISH. siRNA-mediated knockdown followed by RNA-sequencing, revealed that the HELLP lincRNA activated a large set of genes that are involved in the cell cycle. Furthermore, blocking potential mutation sites identified in HELLP families decreased the invasion capacity of extravillous trophoblasts. This is the first large noncoding gene to be linked to a Mendelian disorder with autosomal-recessive inheritance.
Marie van Dijk, Hari K. Thulluru, Joyce Mulders, Omar J. Michel, Ankie Poutsma, Sandra Windhorst, Gunilla Kleiverda, Daoud Sie, Augusta M.A. Lachmeijer, Cees B.M. Oudejans
Translocations are chromosomal rearrangements that are frequently associated with a variety of disease states and developmental disorders. We identified 2 families with brachydactyly type E (BDE) resulting from different translocations affecting chromosome 12p. Both translocations caused downregulation of the parathyroid hormone-like hormone (PTHLH) gene by disrupting the cis-regulatory landscape. Using chromosome conformation capturing, we identified a regulator on chromosome 12q that interacts in cis with PTHLH over a 24.4-megabase distance and in trans with the sex-determining region Y-box 9 (SOX9) gene on chromosome 17q. The element also harbored a long noncoding RNA (lncRNA). Silencing of the lncRNA, PTHLH, or SOX9 revealed a feedback mechanism involving an expression-dependent network in humans. In the BDE patients, the human lncRNA was upregulated by the disrupted chromosomal association. Moreover, the lncRNA occupancy at the PTHLH locus was reduced. Our results document what we believe to be a novel in cis– and in trans–acting DNA and lncRNA regulatory feedback element that is reciprocally regulated by coding genes. Furthermore, our findings provide a systematic and combinatorial view of how enhancers encoding lncRNAs may affect gene expression in normal development.
Philipp G. Maass, Andreas Rump, Herbert Schulz, Sigmar Stricker, Lisanne Schulze, Konrad Platzer, Atakan Aydin, Sigrid Tinschert, Mary B. Goldring, Friedrich C. Luft, Sylvia Bähring