Consequences of disease-causing mutations on lubricin protein synthesis, secretion, and post-translational processing

被引:51
|
作者
Rhee, DK
Marcelino, J
Al-Mayouf, S
Schelling, DK
Bartels, CF
Cui, YJ
Laxer, R
Goldbach-Mansky, R
Warman, ML
机构
[1] Case Western Reserve Univ, Dept Genet, Sch Med, Cleveland, OH 44106 USA
[2] Case Western Reserve Univ, Ctr Human Genet, Sch Med, Cleveland, OH 44106 USA
[3] Univ Hosp Cleveland, Cleveland, OH 44106 USA
[4] King Faisal Specialist Hosp & Res Ctr, Dept Pediat, Riyadh 11211, Saudi Arabia
[5] Hosp Sick Children, Dept Pediat, Toronto, ON M5G 1X8, Canada
[6] Hosp Sick Children, Div Rheumatol, Toronto, ON M5G 1X8, Canada
[7] Univ Toronto, Toronto, ON M5G 1X8, Canada
[8] NIAMS, NIH, Bethesda, MD 20892 USA
关键词
D O I
10.1074/jbc.M505401200
中图分类号
Q5 [生物化学]; Q7 [分子生物学];
学科分类号
071010 ; 081704 ;
摘要
Lubricin, a protein product of the gene PRG4, is a secreted mucin-like proteoglycan that is a major lubricant in articulating joints. Mutations in PRG4 cause the autosomal recessive, human disorder camptodactyly-arthropathycoxa vara-pericarditis syndrome. We developed rabbit polyclonal antibodies against human lubricin to determine the consequence of disease-causing mutations at the protein level and to study the protein's normal post-translational processing. Antiserum generated against an epitope in the amino-terminal portion of lubricin detected protein in wild-type synovial fluid and in conditioned media from wild-type cultured synoviocytes. However, the antiserum did not detect lubricin in synovial fluid or cultured synoviocytes from several patients with frameshift or nonsense mutations in PRG4. Antiserum generated against an epitope in the protein's carboxyl-terminal, hemopexin-like domain identified a post-translational cleavage event in wild-type lubricin, mediated by a subtilisin-like proprotein convertase (SPC). Interestingly, in contrast to wild-type lubricin, one disease-causing mutation that removes the last 8 amino acids of the protein, including a conserved cysteine residue, was not cleaved within the hemopexin-like domain when expressed in COS-7 cells. This suggests that formation of an intrachain disulfide bond is required for SPC-mediated cleavage and that SPC-mediated cleavage is essential to protein function.
引用
收藏
页码:31325 / 31332
页数:8
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