title: Biochemical consequences of sedlin mutations that cause spondyloepiphyseal dysplasia tarda.
abstract: SEDT (spondyloepiphyseal dysplasia tarda) is a late-onset X-linked recessive skeletal dysplasia caused by mutations in the gene SEDL coding for sedlin. In the present paper, we investigated four missense mutations observed in SEDT and compare biochemical and cellular characteristics relative to the wild-type protein to address the mechanism of disease and to gain insight into the function of the sedlin protein. In situ hybridization and immunohistochemical experiments in mouse growth plates revealed sedlin to be predominantly expressed in proliferating and hypertrophic chondrocytes. Cell culture studies showed that the wild-type protein localized predominantly in the vicinity of the nucleus and the Golgi, with further localization around the cytoplasm, whereas mutation resulted in mislocalization. The D47Y mutant was expressed similarly to the wild-type, but the S73L, F83S and V130D mutants showed particularly low levels of expression that were rescued in the presence of the proteasome inhibitor MG132 (benzyloxycarbonyl-leucylleucylleucinal). Furthermore, whereas the D47Y mutant folded similarly and had similar stability to the wild-type sedlin as shown by CD and fluorescence, the S73L, F83S and V130D mutants all misfolded during expression. Two independent assays showed that the D47Y mutation resulted in an increased affinity for the transport protein particle component Bet3 compared with the wild-type sedlin. Our results suggest that the sedlin mutations S73L, F83S and V130D cause SEDT by sedlin misfolding, whereas the D47Y mutation may influence normal TRAPP (transport protein particle) dynamics.
authors: Department of Biochemistry, The University of Hong Kong, Pokfulam, Hong Kong, PR China.
authors: Mei Y
authors: MY
authors: Choi
authors: Caleb C Y
authors: CC
authors: Chan
authors: Danny
authors: D
authors: Chan
authors: Keith D K
authors: KD
authors: Luk
authors: Kathryn S E
authors: KS
authors: Cheah
authors: Julian A
authors: JA
authors: Tanner
mesh_terms: D019943:Amino Acid Substitution / Q000235:genetics / Q000502:physiology; D000818:Animals; D019556:COS Cells; D002478:Cells, Cultured; D002522:Chlorocebus aethiops; D019902:Chondrocytes / Q000378:metabolism; D006132:Growth Plate / Q000378:metabolism; D026901:Membrane Transport Proteins / Q000737:chemistry / Q000235:genetics* / Q000378:metabolism* / Q000502:physiology; D051379:Mice; D008958:Models, Molecular; D009154:Mutation / Q000502:physiology*; D010009:Osteochondrodysplasias / Q000235:genetics*; D020641:Polymorphism, Single Nucleotide / Q000502:physiology; D046988:Proteasome Endopeptidase Complex / Q000378:metabolism; D017510:Protein Folding; D011499:Protein Processing, Post-Translational / Q000235:genetics; D021381:Protein Transport / Q000235:genetics; D014018:Tissue Distribution; D014157:Transcription Factors / Q000737:chemistry / Q000235:genetics* / Q000378:metabolism* / Q000502:physiology
chemical_list: D026901:Membrane Transport Proteins; C120759:TRAPPC2 protein, human; D014157:Transcription Factors; C544123:sedlin protein, mouse; D046988:Proteasome Endopeptidase Complex