Sulfation of chondroitin sulfate proteoglycans is necessary for proper Indian hedgehog signaling in the developing growth plate

Abstract

In contrast to the functional role of heparan sulfate proteoglycans (HSPGs), the importance of chondroitin sulfate proteoglycans (CSPGs) in modulating signaling pathways involving hedgehog proteins, wingless-related proteins and fibroblast growth factors remains unclear. To elucidate the importance of sulfated CSPGs in signaling paradigms required for endochondral bone formation, the brachymorphic (bm) mouse was used as a model for undersulfated CSPGs. The bm mouse exhibits a postnatal chondrodysplasia caused by a mutation in the phosphoadenosine phosphosulfate (PAPS) synthetase (Papss2) gene, leading to reduced levels of PAPS and undersulfated proteoglycans. Biochemical analysis of the glycosaminoglycan (GAG) content in bm cartilage via sulfate labeling and fluorophore-assisted carbohydrate electrophoresis revealed preferential undersulfation of chondroitin chains (CS) and normal sulfation of heparan sulfate chains. In situ hybridization and immunohistochemical analysis of bm limb growth plates showed diminished Indian hedgehog (Ihh) signaling and abnormal Ihh protein distribution in the extracellular matrix. Consistent with the decrease in hedgehog signaling, BrdU incorporation exhibited a significant reduction in chondrocyte proliferation. Direct measurements of Ihh binding to defined GAG chains demonstrated that Ihh interacts with CS, particularly chondroitin-4-sulfate. Furthermore, co-immunoprecipitation experiments showed that Ihh binds to the major cartilage CSPG aggrecan via its CS chains. Overall, this study demonstrates an important function for CSPGs in modulating Ihh signaling in the developing growth plate, and highlights the importance of carbohydrate sulfation in regulating growth factor signaling.

ER Stress During the Pubertal Growth Spurt Results in Impaired Long-Bone Growth in Chondrocyte-Specific ERp57 Knockout Mice

“…In addition, other skeletal abnormalities, such as pseudoachondrodysplasia (PSACH) or multiple epiphyseal dysplasia (MED) are caused by mutations in cartilage ECM proteins like collagens II, IX, X, and XI, or aggrecan, COMP, and matrilin-3. (26)(27)(28)(29)34,(37)(38)(39)(40)(41)(42)(43) ER stress is thought to constitute the underlying common disease mechanism due to retention of abnormal, improperly folded proteins within the ER. The cells can be rescued after the ER stress-induced unfolded protein response, which arrests the cell cycle, reduces the general protein synthesis, and enhances the production of new chaperones facilitating additional folding processes.…”

Excesso de tiroxina materna associado ao hipertireoidismo pós-natal reduz o crescimento ósseo e o perfil proliferativo e angiogênico das cartilagens de crescimento de ratos

“…A vasculatura fornece oxigênio, nutrientes e fatores de crescimento às células e aos tecidos (Karaca et al, 2015). Para que isso ocorra, a matriz extracelular das cartilagens de crescimento desempenha função importante na difusão de substâncias, particularmente das moléculas de sinalização e dos fatores de crescimento (Cortes et al, 2009). Desta forma, é possível que a redução dos fatores pró-angiogênicos observados nas cartilagens de crescimento de ratos hipertireoideos possa impedir a difusão de moléculas pela matriz extracelular, alterando a composição da matriz cartilaginosa.…”

Epiphyseal growth plate and secondary peripheral chondrosarcoma: the neighbours matter

“…Albeit the genetic correlation between mutations in EXT1/EXT2 and osteochondromas, the mechanism by which alterations in heparan sulphate biosynthesis leads to osteochondroma is not entirely understood. As heparan sulphate acts as a coreceptor for fibroblast growth factors and BMPs [49], and regulates the diffusion of Ihh [19] and members of the Wnt family [26], improper elongation of heparan sulphate chains may result in a variety of growth factor signalling defects and impaired cell-matrix interactions, which ultimately may result in osteochondroma formation ( Figure 2).…”

“…Throughout the ECM, the distribution of morphogens in a gradient fashion is shown to be established by proteoglycans [5,18,19]. Proteoglycans are found in the ECM and attached to the cell membrane in virtually all types of tissue.…”

“…Therefore, proteoglycans regulate the distribution of morphogens across the ECM [17]. Proteoglycans, such as heparan and chondroitin sulphate, function in concert to establish an Indian hedgehog (Ihh) gradient, either through affecting its diffusion or by protecting it from degradation [19]. Ihh is a key regulator of the epiphyseal growth plate.…”