Novel mutations in exon 2 ofMATN3affect residues within the α-helices of the A-domain and can result in the intracellular retention of mutant matrilin-3

Abstract

Multiple epiphyseal dysplasia (MED) is a clinically variable and genetically heterogeneous chondrodysplasia characterized by mild to moderate short stature and early onset osteoarthritis. Some forms of MED result from mutations in the gene encoding the cartilage structural protein matrilin-3 (MATN3). The majority of MATN3 mutations affect conserved residues within the β-sheet of the single A-domain of matrilin-3. These mutations cause the protein to misfold and prevent its secretion from the rER, both in vitro and in vivo. More recently a single mutation (p.Phe105Ser) has been identified within the α1 -helix of the Adomain, but its affect on the structure and/or function of matrilin-3 is unknown. In this paper we describe the characterization of two additional α-helical mutations (p.Ala173Asp and p.Lys231Asn) and show that both p.Phe105Ser and pAla173Asp prevent the secretion of A-domain in vitro. In contrast, p.Lys231Asn does not prevent the secretion of matrilin-3 Adomain, nor does it disrupt the structure of this domain or inhibit its binding to type II or type IX collagen. Therefore, despite extensive biochemical analysis the disease mechanism of p.Lys231Asn remains unresolved and care should be taken in counseling for these types of mutation in MATN3.

A matrilin-3 mutation associated with osteoarthritis does not affect collagen affinity but promotes the formation of wider cartilage collagen fibrils

“…It is still uncertain if there is a correlation between the domain localization of the mutation and the disease phenotype. In most cases chondrodysplasia-causing mutations in matrilin-3 interfere with protein trafficking and lead to an intracellular protein accumulation [Cotterill et al, 2005;Fresquet et al, 2008;Otten et al, 2005]. Protein retention is a common phenomenon in the pathology of skeletal diseases, and has also been described for disease-causing mutations in COMP and both collagen II and IX [Bönnemann et al, 2000; Dinser et al, Figure 3.…”

“…Mutant COMP was found in enlarged cisternae of the ER together with other coretained matrix molecules, and elicits a cell stress response and chondrocyte apoptosis [Dinser et al, 2002;Hashimoto et al, 2003;Hecht et al, 2004;Merritt et al, 2007;Schmitz et al, 2006Schmitz et al, , 2008. However, for both COMP and matrilin-3 MED-causing mutations have been described that do not prevent protein secretion [Fresquet et al, 2008;Schmitz et al, 2006;Spitznagel et al, 2004]. Furthermore, the OA-associated mutation p.T303M does not hinder protein trafficking [Otten et al, 2005].…”

“…To date, 17 disease-causing amino acid substitutions have been identified. The majority of the mutations affect conserved residues within the VWA-like domain and cause different forms of MED [Chapman et al, 2001;Cotterill et al, 2005;Fresquet et al, 2008;Jackson et al, 2004;Mabuchi et al, 2004;Mostert et al, 2003]. Mutations within the central b-sheet are more frequent than mutations in the a-helical regions [Fresquet et al, 2008].…”

Armet/Manf and Creld2 are components of a specialized ER stress response provoked by inappropriate formation of disulphide bonds: implications for genetic skeletal diseases

“…The A-domain is arranged into a classical Rossman fold and contains a single intra-chain disulphide bond. To date, all MED causing mutations in matrilin-3 are located within the single A-domain and primarily affect residues that comprise the central β-sheet (2,18–21). A murine model of MED with the Matn3 V194D mutation develops a progressive short-limb dysplasia resulting from decreased chondrocyte proliferation and dysregulated apoptosis.…”

Matrilin-3 Chondrodysplasia Mutations Cause Attenuated Chondrogenesis, Premature Hypertrophy and Aberrant Response to TGF-β in Chondroprogenitor Cells

“…Currently there are more than 13 known MED associated autosomal dominant missense mutations have been mapped to MATN3 ’s vWFA protein domain [14,15,16,17,18,19]. A single tryptophan to arginine (R121W) point mutation in the β-strand of the vWFA domain has been identified in approximately one third of all MED patients, making it the most common MATN3 mutation to cause this disorder [1,17].…”