Latent TGF-β–binding protein 4 modifies muscular dystrophy in mice
Ahlke Heydemann, … , Abraham A. Palmer, Elizabeth M. McNally
Ahlke Heydemann, … , Abraham A. Palmer, Elizabeth M. McNally
Published November 2, 2009
Citation Information: J Clin Invest. 2009;119(12):3703-3712. https://doi.org/10.1172/JCI39845.
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Research Article

Latent TGF-β–binding protein 4 modifies muscular dystrophy in mice

Abstract

Most single-gene diseases, including muscular dystrophy, display a nonuniform phenotype. Phenotypic variability arises, in part, due to the presence of genetic modifiers that enhance or suppress the disease process. We employed an unbiased mapping approach to search for genes that modify muscular dystrophy in mice. In a genome-wide scan, we identified a single strong locus on chromosome 7 that influenced two pathological features of muscular dystrophy, muscle membrane permeability and muscle fibrosis. Within this genomic interval, an insertion/deletion polymorphism of 36 bp in the coding region of the latent TGF-β–binding protein 4 gene (Ltbp4) was found. Ltbp4 encodes a latent TGF-β–binding protein that sequesters TGF-β and regulates its availability for binding to the TGF-β receptor. Insertion of 12 amino acids into the proline-rich region of LTBP4 reduced proteolytic cleavage and was associated with reduced TGF-β signaling, decreased fibrosis, and improved muscle pathology in a mouse model of muscular dystrophy. In contrast, a 12-amino-acid deletion in LTBP4 was associated with increased proteolysis, SMAD signaling, and fibrosis. These data identify Ltbp4 as a target gene to regulate TGF-β signaling and modify outcomes in muscular dystrophy.

Authors

Ahlke Heydemann, Ermelinda Ceco, Jackie E. Lim, Michele Hadhazy, Pearl Ryder, Jennifer L. Moran, David R. Beier, Abraham A. Palmer, Elizabeth M. McNally

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Figure 5

The 12-amino-acid deletion increases LTBP4 protease susceptibility.

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The 12-amino-acid deletion increases LTBP4 protease susceptibility. (A a...
(A and B) Total protein was extracted from D2 and 129 fibroblasts with and without the Sgcg-null allele, digested, and immunoblotted with an antibody directed toward the amino terminus of LTBP4. (A) Digests with 12 μg of pancreatin. Each pair represents Sgcg mutant, then wild-type. (B) Digestion with 25 μg of plasmin. Each set of 4 represents 3 mutants, then a wild-type. LTBP4 from the D2 strain was digested more easily than LTBP4 from the 129 strain. The percent digested refers to the proteolyzed lower product. On average, plasmin digested 38% of 129-derived LTBP4 versus 57% of D2-derived LTBP4. (C and D) In vitro expression constructs. D2- and 129-derived LTBP4 proline-rich sequences were expressed in vitro. The LTBP4 sequence is underlined; the insertion is shown in gray; and the deletion is represented by dashed lines. The position of the cysteine residues in the expressed sequences is indicated by the asterisks. (E) The constructs were expressed in vitro with [35S]cysteine and then exposed to plasmin, a protease implicated in cleavage of LTBP4 (15). The D2-derived sequences are digested at low levels of plasmin, while only the highest level of plasmin can begin to produce the fully digested product from the 129-derived sequence. The slower migration of the 12-amino-acid-deleted proline-rich region, prior to digestion, is consistent with an altered conformational state that is more susceptible to proteolysis. The arrow indicates the cleavage product.