An ADAMTSL2 Founder Mutation Causes Musladin-Lueke Syndrome, a Heritable Disorder of Beagle Dogs, Featuring Stiff Skin and Joint Contractures Hannah L. Bader 1. , Alison L. Ruhe 2. , Lauren W. Wang 1 , Aaron K. Wong 2¤a , Kari F. Walsh 2 , Rebecca A. Packer 3 , Jonathan Mitelman 4 , Kathryn R. Robertson 2 , Dennis P. O’Brien 5 , Karl W. Broman 6 , G. Diane Shelton 7 , Suneel S. Apte 1 , Mark W. Neff 2 * ¤b¤c 1 Department of Biomedical Engineering, Lerner Research Institute, Cleveland Clinic, Cleveland, Ohio, United States of America, 2 Veterinary Genetics Laboratory, University of California Davis, Davis, California, United States of America, 3 Department of Veterinary Clinical Sciences, Purdue University, West Lafayette, Indiana, United States of America, 4 Kingston Road Animal Hospital, Toronto, Ontario, Canada, 5 Department of Veterinary Medicine and Surgery, University of Missouri, Columbia, Missouri, United States of America, 6 Department of Biostatistics and Medical Informatics, University of Wisconsin, Madison, Wisconsin, United States of America, 7 Department of Pathology, School of Medicine, University of California San Diego, La Jolla, California, United States of America Abstract Background: Musladin-Lueke Syndrome (MLS) is a hereditary disorder affecting Beagle dogs that manifests with extensive fibrosis of the skin and joints. In this respect, it resembles human stiff skin syndrome and the Tight skin mouse, each of which is caused by gene defects affecting fibrillin-1, a major component of tissue microfibrils. The objective of this work was to determine the genetic basis of MLS and the molecular consequence of the identified mutation. Methodology and Principal Findings: We mapped the locus for MLS by genome-wide association to a 3.05 Mb haplotype on canine chromosome 9 (CFA9 (50.11–54.26; p raw ,10 27 )), which was homozygous and identical-by-descent among all affected dogs, consistent with recessive inheritance of a founder mutation. Sequence analysis of a candidate gene at this locus, ADAMTSL2, which is responsible for the human TGFb dysregulation syndrome, Geleophysic Dysplasia (GD), uncovered a mutation in exon 7 (c.660C.T; p.R221C) perfectly associated with MLS (p-value = 10 212 ). Murine ADAMTSL2 containing the p.R221C mutation formed anomalous disulfide-bonded dimers when transiently expressed in COS-1, HEK293F and CHO cells, and was present in the medium of these cells at lower levels than wild-type ADAMTSL2 expressed in parallel. Conclusions/Significance: The genetic basis of MLS is a founder mutation in ADAMTSL2, previously shown to interact with latent TGF-b binding protein, which binds fibrillin-1. The molecular effect of the founder mutation on ADAMTSL2 is formation of disulfide-bonded dimers. Although caused by a distinct mutation, and having a milder phenotype than human GD, MLS nevertheless offers a new animal model for study of GD, and for prospective insights on mechanisms and pathways of skin fibrosis and joint contractures. Citation: Bader HL, Ruhe AL, Wang LW, Wong AK, Walsh KF, et al. (2010) An ADAMTSL2 Founder Mutation Causes Musladin-Lueke Syndrome, a Heritable Disorder of Beagle Dogs, Featuring Stiff Skin and Joint Contractures. PLoS ONE 5(9): e12817. doi:10.1371/journal.pone.0012817 Editor: Gregory S. Barsh, Stanford University, United States of America Received July 30, 2010; Accepted August 24, 2010; Published September 17, 2010 Copyright: ß 2010 Bader et al. This is an open-access article distributed under the terms of the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are credited. Funding: This work was supported by intramural funds from the Veterinary Genetics Laboratory of UC Davis and by National Institutes of Health award AR53890 (S. Apte) and T32HL007914-08 (Training Program in Vascular Biology and Pathology, Principal Investigator: Edward Plow) awarded to H. Bader. The funders had no role in study design, data collection and analysis, decision to publish, or preparation of the manuscript. Competing Interests: The authors have declared that no competing interests exist. * E-mail: mark.neff@vai.org . These authors contributed equally to this work. ¤a Current address: Department of Computer Science, Princeton University, Princeton, New Jersey, United States of America ¤b Current address: Translational Genomics Research Institute, Phoenix, Arizona, United States of America ¤c Current address: The Van Andel Research Institute, Grand Rapids, Michigan, United States of America Introduction The critical role of extracellular matrix (ECM) in sequestration and regulated release or activation of growth factors is highly relevant to common medical problems such as fibrosis. An excellent example showcasing this role is regulation of TGF-b by tissue microfibrils, which contain the glycoprotein fibrillin-1 as a major constituent. The three TGFb isoforms exist as disulfide- bonded dimers [1]. Their propeptides are processed intracellularly by furin, but remain associated with the active dimer (the latency- associated peptide) [1] as the small latent complex (SLC), which in turn, binds to latent TGFb–binding protein (LTBP)-1, -3, and -4, to form a large latent complex (LLC) [1,2,3]. The LLC is sequestered in the ECM through binding of LTBP1 to the ECM glycoproteins fibrillin-1 and fibronectin [4,5,6,7]. TGFb activity appears to be regulated primarily at the level of activation in ECM, which occurs either by a non-proteolytic mechanism involving integrins and thrombospondin-1, or via proteolysis of the PLoS ONE | www.plosone.org 1 September 2010 | Volume 5 | Issue 9 | e12817