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Site-1 protease deficiency causes human skeletal dysplasia due to defective inter-organelle protein trafficking
Yuji Kondo, … , Patrick M. Gaffney, Lijun Xia
Yuji Kondo, … , Patrick M. Gaffney, Lijun Xia
Published July 26, 2018
Citation Information: JCI Insight. 2018;3(14):e121596. https://doi.org/10.1172/jci.insight.121596.
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Research Article Cell biology Genetics

Site-1 protease deficiency causes human skeletal dysplasia due to defective inter-organelle protein trafficking

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Abstract

Site-1 protease (S1P), encoded by MBTPS1, is a serine protease in the Golgi. S1P regulates lipogenesis, endoplasmic reticulum (ER) function, and lysosome biogenesis in mice and in cultured cells. However, how S1P differentially regulates these diverse functions in humans has been unclear. In addition, no human disease with S1P deficiency has been identified. Here, we report a pediatric patient with an amorphic and a severely hypomorphic mutation in MBTPS1. The unique combination of these mutations results in a frequency of functional MBTPS1 transcripts of approximately 1%, a finding that is associated with skeletal dysplasia and elevated blood lysosomal enzymes. We found that the residually expressed S1P is sufficient for lipid homeostasis but not for ER and lysosomal functions, especially in chondrocytes. The defective S1P function specifically impairs activation of the ER stress transducer BBF2H7, leading to ER retention of collagen in chondrocytes. S1P deficiency also causes abnormal secretion of lysosomal enzymes due to partial impairment of mannose-6-phosphate–dependent delivery to lysosomes. Collectively, these abnormalities lead to apoptosis of chondrocytes and lysosomal enzyme–mediated degradation of the bone matrix. Correction of an MBTPS1 variant or reduction of ER stress mitigated collagen-trafficking defects. These results define a new congenital human skeletal disorder and, more importantly, reveal that S1P is particularly required for skeletal development in humans. Our findings may also lead to new therapies for other genetic skeletal diseases, as ER dysfunction is common in these disorders.

Authors

Yuji Kondo, Jianxin Fu, Hua Wang, Christopher Hoover, J. Michael McDaniel, Richard Steet, Debabrata Patra, Jianhua Song, Laura Pollard, Sara Cathey, Tadayuki Yago, Graham Wiley, Susan Macwana, Joel Guthridge, Samuel McGee, Shibo Li, Courtney Griffin, Koichi Furukawa, Judith A. James, Changgeng Ruan, Rodger P. McEver, Klaas J. Wierenga, Patrick M. Gaffney, Lijun Xia

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Figure 6

Improving ER function or antisense morpholino oligo splicing therapy mitigates collagen retention in the ER of MBTPS1 mutant cells.

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Improving ER function or antisense morpholino oligo splicing therapy mit...
(A) Structure of MBTPS1 pre-mRNA in abnormal and corrected maternal transcripts. The antisense morpholino oligo (AMO) was designed to block the maternal alternative MBTPS1 splicing site. The red box indicates AMO designed to block cryptic splice site. The asterisk indicates the premature termination. AMO hampers pathogenic alternative splicing and promote correct mRNA splicing. (B) Maternal (blue) and patient (red) fibroblasts were treated with AMO for 12, 24, and 48 hours, and MBTPS1 expression was analyzed by qRT-PCR normalized to HPRT expression. Numbers indicate fold change compared with time 0. For total MBTPS1, a forward primer designed in exon 19 and a reverse primer designed in exon 21 of MBTPS1 were used. For MBTPS1 with Δ41 bp, a forward primer designed in the boundary of exon 7–8 and a reverse primer designed in the boundary of exon 9–10 of MBTPS1 were used. (C) Western blotting of endogenous BBF2H7 in AMO or control oligo-treated patient fibroblasts. PF, S1P inhibitor PF-429242. Defective S1P-dependent cleavage of BBF2H7 and improved cleavage by AMO treatment in patient fibroblasts. (D) Secreted procollagen I from maternal and patient fibroblasts, treated as indicated, was measured by ELISA. AMO treatment increased collagen I secretion in patient fibroblasts. (E) Immunofluorescence images of collagen I in fibroblasts treated with or without the PF-429242 or the PBA for 48 hours. TO-PRO, nuclear staining. Scale bar: 5 μm. Quantification of mean fluorescent intensity (MFI) of intracellular collagen I staining. A dramatic decrease in collagen I in PBA-treated patient fibroblasts was seen. (F) A model showing how S1P deficiency causes compound defects in the ER and lysosome, leading to skeletal dysplasia. All data are from at least 3 independent experiments. Mean ± SEM. *P < 0.05; **P < 0.01, Student’s t test.

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