chondroitin-sulfates and Dwarfism

TMEM165 deficiency leads to skeletal disorder characterized by major skeletal dysplasia and pronounced dwarfism. However, the molecular mechanisms involved have not been fully understood. Here, we uncover that TMEM165 deficiency impairs the synthesis of proteoglycans by producing a blockage in the elongation of chondroitin-and heparan-sulfate glycosaminoglycan chains leading to the synthesis of proteoglycans with shorter glycosaminoglycan chains. We demonstrated that the blockage in elongation of glycosaminoglycan chains is not due to defect in the Golgi elongating enzymes but rather to availability of the co-factor Mn

Topics: Animals; Antiporters; Case-Control Studies; Cation Transport Proteins; Cell Differentiation; Cell Line, Tumor; Chondrocytes; Chondrogenesis; Chondroitin Sulfates; Dwarfism; Fibroblasts; Gene Knockout Techniques; Glycosylation; HEK293 Cells; Heparan Sulfate Proteoglycans; Humans; Hypertrophy; Mice; Signal Transduction; Transfection

Galnt3, UDP-N-acetyl-α-D-galactosamine:polypeptide N-acetylgalactosaminyltransferase 3, transfers N-acetyl-D-galactosamine to serine and threonine residues, initiating mucin type O-glycosylation of proteins. We searched the target genes of Runx2, which is an essential transcription factor for chondrocyte maturation, in chondrocytes and found that Galnt3 expression was up-regulated by Runx2 and severely reduced in Runx2(-/-) cartilaginous skeletons. To investigate the function of Galnt3 in chondrocytes, we generated Galnt3(-/-) mice and chondrocyte-specific Galnt3 transgenic mice under the control of the Col2a1 promoter-enhancer. Galnt3(-/-) mice showed a delay in endochondral ossification and shortened limbs at embryonic day 16.5, suggesting that Galnt3 is involved in chondrocyte maturation. Galnt3 transgenic mice presented dwarfism, the chondrocyte maturation was retarded, the cell cycle in chondrocytes was accelerated, premature chondrocyte apoptosis occurred, and the growth plates were disorganized. The binding of Vicia villosa agglutinin, which recognizes the Tn antigen (GalNAc-O-Ser/Thr), was drastically increased in chondrocytes, and aggrecan (Acan) was highly enriched with Tn antigen. However, safranin O staining, which recognizes glycosaminoglycans (GAGs), and Acan were severely reduced. Chondroitin sulfate was reduced in amount, but the elongation of chondroitin sulfate chains had not been severely disturbed in the isolated GAGs. These findings indicate that overexpression of Galnt3 in chondrocytes caused dwarfism due to the increase of mucin-type O-glycans and the reduction of GAGs, probably through competition with xylosyltransferases, which initiate GAG chains by attaching O-linked xylose to serine residues, suggesting a negative effect of Galnt family proteins on Acan deposition in addition to the positive effect of Galnt3 on chondrocyte maturation.

Topics: Aggrecans; Animals; Apoptosis; Cartilage; Cell Proliferation; Cells, Cultured; Chondrocytes; Chondroitin Sulfates; Core Binding Factor Alpha 1 Subunit; Dwarfism; Female; Gene Expression; Glycosylation; Growth Plate; Male; Mice, 129 Strain; Mice, Inbred C57BL; Mice, Knockout; N-Acetylgalactosaminyltransferases; Osteogenesis; Polypeptide N-acetylgalactosaminyltransferase; Protein Processing, Post-Translational

The brachymorphic (bm/bm) mouse is a disproportionate dwarf with a disturbance of the endochondral growth of the skeleton. Rib cartilage from 25-day-old affected animals and their normal siblings was analyzed for its contents and composition of proteoglycans. In addition to the previously reported undersulfation of chondroitin sulfate, it was demonstrated that one of the two types of aggregating proteoglycan and possibly the small ones are decreased in bm/bm costal cartilage, both in the growth region and in the remaining part. The molecular defect of the bm/bm condition is known to affect the synthesis of 3-phosphoadenosine 5-phosphosulfate (Sugahara and Schwartz, Proc. Natl. Acad. Sci. USA 76: 6615-6618, 1979). The above finding therefore suggests the existence of feedback mechanisms for the regulation of proteoglycan synthesis, whereby the undersulfation of glycosaminoglycans would result in decreased synthesis or increased turnover of certain proteoglycan subpopulations. Analysis of the glycosaminoglycan side chains indicated that mouse rib cartilage contains small amounts of keratan sulfate of extremely small size. The affected and control tissues, however, seemed to contain equal amounts of both glucosamine and galactosamine.

Topics: Animals; Cartilage; Chondroitin Sulfates; Dwarfism; Feedback; Glycosaminoglycans; Hexosamines; Keratan Sulfate; Mice; Mice, Mutant Strains; Molecular Weight; Phosphoadenosine Phosphosulfate; Proteoglycans; Rodent Diseases

Cartilage from patients with pseudoachondroplasia is characterized by unique inclusions in the cisternae of the endoplasmic reticulum and proteoglycan abnormalities have been suggested in this form of dwarfism. To elucidate the nature of the proteoglycan defect, we determined the amount of the individual glycosaminoglycans present in iliac-crest cartilage of three patients and extracted the proteoglycan monomers from one of the samples. Sections of iliac-crest cartilage and proximal fibular growth plates were examined by electron microscopy and also stained with hematoxylin and eosin, safranin O-fast green, and alcian blue in the presence of increasing concentrations of magnesium chloride (zero to one molar). The chondrocytes of the iliac crest and fibular physes were arranged in clusters more than in columns and contained characteristic endoplasmic reticulum inclusions, which were particularly large in the hypertrophic cells. The cartilage stained very poorly with hematoxylin and eosin and with safranin O-fast green. The alcian-blue stain was abolished from perilacunar areas and from longitudinal septa by magnesium chloride concentrations that were lower than those required by normal tissue. The proteoglycans of iliac-crest cartilage were found to be significantly enriched in keratan sulphate and had a below-normal ratio of chondroitin-4-sulphate to chondroitin-6-sulphate, although the amount of the two isomeric chondroitin sulphates combined was within normal limits. The urinary excretion of glycosaminoglycan by the three patients was normal. Pseudoachondroplasia appears to be a generalized cartilage disorder involving abnormalities of proteoglycans, probably related to the core protein or to enzymes that are responsible for the formation of the glycosaminoglycan chains.(ABSTRACT TRUNCATED AT 250 WORDS)

Topics: Achondroplasia; Adolescent; Cartilage; Child; Chondroitin Sulfates; Dwarfism; Endoplasmic Reticulum; Glycosaminoglycans; Histocytochemistry; Humans; Keratan Sulfate; Proteoglycans

A 14-year-old boy was found to excrete excessive amounts of acidic glycosaminoglycans which were predominantly chondroitin 4-sulfate and chondroitin 6-sulfate. Clinical features included dwarfism, mental retardation, coarse facies, deformities of the spine, hip joints and thorax, and granulations in leucocytes. The clinical and biochemical features found in this boy were compared with the known types of mucopolysaccharidosis and it has been concluded that this case is a new type of mucopolysacchariduria.

Topics: Adolescent; Chondroitin; Chondroitin Sulfates; Dwarfism; Glycosaminoglycans; Humans; Intellectual Disability; Male; Mucopolysaccharidoses

Incorporation of 35SO42- into adenosine 5'-phosphosulfate (APS), 3'-phosphoadenosine 5'-phosphosulfate (PAPS), and chondroitin sulfate was simultaneously assessed with extracts prepared from epiphyseal cartilage of neonatal normal or homozygous brachymorphic mice. Radioactivity measured in APS, PAPS, and chondroitin sulfate of extracts from brachymorphic cartilage was approximately 300%, 9%, and 13% of the normal levels, respectively. Even though more APS accumulated in the mutant cartilage extracts, APS actually synthesized (total 35SO42- incorporated into APS, PAPS, and macromolecular products) was only 17% of that in the normal. However, of the amount synthesized, 90% and 55% of newly synthesized APS were converted to PAPS by cartilage extracts of normal and brachymorphic mice, respectively. Specific assays for ATP sulfurylase (sulfate adenylyltransferase; ATP:sulfate adenylyltransferase, EC 2.7.7.4) and APS kinase (adenylylsulfate kinase; ATP:adenylylsulfate 3'-phosphotransferase, EC 2.7.1.25) showed that the sulfurylase enzyme activity is reduced to approximately 1/2 and the kinase to approxomately 1/14 in brachymorphic mice. These results suggest that the production of an undersulfated proteoglycan in brachymorphic cartilage results from a defective conversion of APS to PAPS.

Topics: Adenine Nucleotides; Adenosine Phosphosulfate; Animals; Cartilage; Chondroitin; Chondroitin Sulfates; Dwarfism; Mice; Phosphoadenosine Phosphosulfate; Phosphotransferases; Sulfates; Sulfurtransferases

Topics: Animals; Cartilage; Centrifugation, Density Gradient; Chick Embryo; Chondroitin Sulfates; Dwarfism; Proteoglycans

Topics: Animals; Cattle Diseases; Chondroitin Sulfates; Dwarfism; Glycosaminoglycans; Hydrolases; Mucopolysaccharidosis I; Research; Spectrophotometry; Urine