Maintenance of chondroitin sulfation balance by chondroitin-4-sulfotransferase 1 is required for chondrocyte development and growth factor signaling during cartilage morphogenesis

Abstract

Glycosaminoglycans (GAGs) such as heparan sulfate and chondroitin sulfate are polysaccharide chains that are attached to core proteins to form proteoglycans. The biosynthesis of GAGs is a multistep process that includes the attachment of sulfate groups to specific positions of the polysaccharide chains by sulfotransferases. Heparan-sulfate and heparan sulfate-sulfotransferases play important roles in growth factor signaling and animal development. However, the biological importance of chondroitin sulfation during mammalian development and growth factor signaling is poorly understood. We show that a gene trap mutation in the BMP-induced chondroitin-4-sulfotransferase 1 (C4st1) gene (also called carbohydrate sulfotransferase 11 – Chst11), which encodes an enzyme specific for the transfer of sulfate groups to the 4-O-position in chondroitin, causes severe chondrodysplasia characterized by a disorganized cartilage growth plate as well as specific alterations in the orientation of chondrocyte columns. This phenotype is associated with a chondroitin sulfation imbalance, mislocalization of chondroitin sulfate in the growth plate and an imbalance of apoptotic signals. Analysis of several growth factor signaling pathways that are important in cartilage growth plate development showed that the C4st1gt/gt mutation led to strong upregulation of TGFβ signaling with concomitant downregulation of BMP signaling, while Indian hedgehog (Ihh) signaling was unaffected. These results show that chondroitin 4-O-sulfation by C4st1 is required for proper chondroitin sulfate localization, modulation of distinct signaling pathways and cartilage growth plate morphogenesis. Our study demonstrates an important biological role of differential chondroitin sulfation in mammalian development.

C4ST-1/CHST11-controlled chondroitin sulfation interferes with oncogenic HRAS signaling in Costello syndrome

“…24 Thus, senescence appears to be associated with both gain-of-function of HRAS and loss-of-function of C4ST-1, consistent with our model that loss of C4ST-1 function is important for phenotypic manifestation of oncogenic HRAS signaling. In addition to Costello syndrome, a group of multiple congenital anomaly syndromes collectively named RASopathies have been shown to be associated with germline mutations in various genes of the RAS/ MAPK pathway.…”

“…The analysis of proliferation rates by BrdU incorporation was performed as previously described. 24 Western blot analysis was performed by separating cell lysates in TNTE-based lysis buffer on 10% SDS-PAGE gels, followed by transfer to nitrocellulose membranes and incubation with mouse a-C4ST-1 (Abcam, 1:100) and mouse a-beta-tubulin (Abcam, 1:1000) antibodies.…”

“…21 We have recently generated a loss-of-function mutation in C4ST-1 and demonstrated severely decreased C4S levels and severe cartilage defects in homozygous mutant mouse embryos. 24,32 Although other laboratories have shown that RAS signaling can affect the sulfation status and expression of CS proteoglycans in vitro, 33,34 we have established previously that cellular defects of fibroblasts derived from Costello syndrome patients can be reversed by degrading CSs in these cultures. 35 We also demonstrated an altered chondroitin sulfation balance with decreased C4S levels in the heart tissue of Costello syndrome patients.…”

“…[20][21][22][23] The balance of chondroitin sulfation is tightly controlled through spatial and temporal expression of distinct chondroitin sulfotransferase genes. 21,24,25 Moreover, this tightly controlled chondroitin sulfation balance has been shown to be disturbed in human disease and malignancies. [26][27][28][29][30][31] Carbohydrate sulfotransferase 11/Chondroitin-4-sulfotransferase-1 (CHST11/C4ST-1; subsequently named C4ST-1) catalyzes the transfer of sulfate to the C-4 position of CS disaccharides, thus creating chondroitin-4-sulfate (C4S) as one of its products.…”

Chondroitin beta-1,4-N-acetylgalactosaminyltransferase-1 missense mutations are associated with neuropathies

“…9 Recently, it has been reported that a broad spectrum of skeletal dysplasias result from mutations causing undersulfation of chondroitin sulfate chains in humans and in mice. [10][11][12][13][14] In contrast, no association between CSPGs and human peripheral neuropathies has been reported to date.…”