LTBP-1 is a large extracellular glycoprotein that is a component of the large latent TGF-β complex. The extracellular sequestration of latent TGF-β in the extracellular matrix (ECM) is fundamental to the regulation of TGF-β bioavailability and activity. LTBP-1 is described to contribute to the regulation of TGF-β bioavailability through mediating the extracellular sequestration of newly secreted latent TGF-β with fibrillin microfibrils in the ECM. However it is not well understood how LTBP-1, and thus latent TGF-β, becomes deposited into the ECM. Previous work by our group suggested that LTBP-1 interactions with the glycosaminoglycan heparan sulphate (HS) at the cell-matrix interface might facilitate the association of LTBP-1 with fibrillin microfibrils. Using recombinant LTBP-1 fragments and mutants, LTBP-1 interaction with HS have been fine-mapped. Deposition of a LTBP-1 HS-binding mutant, and of LTBP-1 when HS was depleted, was studied in cell cultures; findings presented here demonstrate that HS may not be critical for the deposition of LTBP-1 into the ECM. Contributions of fibrillin and fibronectin to LTBP-1 deposition were investigated, and data presented here support published findings that fibrillin is not always required for LTBP-1 deposition. In addition, the dependency of LTBP-1 deposition upon fibronectin was suggested to differ between different cell types (epithelial and mesenchymal). How LTBP-1 may be stabilised within the ECM through crosslinking by tissue transglutaminase was investigated using recombinant fragments and cell culture studies. Tissue transglutaminase was found to promote the extracellular incorporation of LTBP-1, and novel cross-links within LTBP-1, and between LTBP-1 and fibrillin-1, but not LTBP-1 and fibronectin, were identified. Additionally, results indicated that LTBP-1 was present in extremely high molecular weight assemblies in the ECM of cultured fibroblasts.Collectively, these results have contributed to current knowledge of how LTBP-1 becomes deposited into the ECM. They indicate that the deposition of LTBP-1 is not underpinned by HS, may be cell type-specific, and that LTBP-1 may potentially self-assemble extracellularly into homotypic structures that may associate with fibrillin microfibrils.
|Date of Award||1 Aug 2014|
- The University of Manchester
|Supervisor||Catherine Kielty (Supervisor) & Anthony Day (Supervisor)|
Investigations of the extracellular deposition of latent TGF-beta binding protein-1 (LTBP-1)
Steer, R. (Author). 1 Aug 2014
Student thesis: Phd