Latent transforming growth factor binding protein 4 regulates transforming growth factor beta receptor stability

Mutations in the gene for the latent transforming growth factor beta binding protein 4 (LTBP4) cause autosomal recessive cutis laxa type 1C. To understand the molecular disease mechanisms of this disease, we investigated the impact of LTBP4 loss on transforming growth factor beta (TGF beta) signaling. Despite elevated extracellular TGF beta activity, downstream signaling molecules of the TGF beta pathway, including pSMAD2 and pERK, were down-regulated in LTBP4 mutant human dermal fibroblasts. In addition, TGF beta receptors 1 and 2 (TGFBR1 and TGFBR2) were reduced at the protein but not at the ribonucleic acid level. Treatment with exogenous TGF beta 1 led to an initially rapid increase in SMAD2 phosphorylation followed by a sustained depression of phosphorylation and receptor abundance. In mutant cells TGFBR1 was co-localized with lysosomes. Treatment with a TGFBR1 kinase inhibitor, endocytosis inhibitors or a lysosome inhibitor, normalized the levels of TGFBR1 and TGFBR2. Co-immunoprecipitation demonstrated a molecular interaction between LTBP4 and TGFBR2. Knockdown of LTBP4 reduced TGF beta receptor abundance and signaling in normal cells and supplementation of recombinant LTBP4 enhanced these measures in mutant cells. In a mouse model of Ltbp4 deficiency, reduced TGF beta signaling and receptor levels were normalized upon TGFBR1 kinase inhibitor treatment. Our results show that LTBP4 interacts with TGFBR2 and stabilizes TGF beta receptors by preventing their endocytosis and lysosomal degradation in a ligand-dependent and receptor kinase activity-dependent manner. These findings identify LTBP4 as a key molecule required for the stability of the TGF beta receptor complex, and a new mechanism by which the extracellular matrix regulates cytokine receptor signaling.