Osteoporosis and skeletal dysplasia caused by pathogenic variants in SGMS2
Minna Pekkinen, Paulien A. Terhal, Lorenzo D. Botto, Petra Henning, Riikka E. Mäkitie, Paul Roschger, Amrita Jain, Matthijs Kol, Matti A. Kjellberg, Eleftherios P. Paschalis, Koen van Gassen, Mary Murray, Pinar Bayrak-Toydemir, Maria K. Magnusson, Judith Jans, Mehran Kausar, John C. Carey, Pentti Somerharju, Ulf H. Lerner, Vesa M. Olkkonen, Klaus Klaushofer, Joost C.M. Holthuis, Outi Mäkitie
Minna Pekkinen, Paulien A. Terhal, Lorenzo D. Botto, Petra Henning, Riikka E. Mäkitie, Paul Roschger, Amrita Jain, Matthijs Kol, Matti A. Kjellberg, Eleftherios P. Paschalis, Koen van Gassen, Mary Murray, Pinar Bayrak-Toydemir, Maria K. Magnusson, Judith Jans, Mehran Kausar, John C. Carey, Pentti Somerharju, Ulf H. Lerner, Vesa M. Olkkonen, Klaus Klaushofer, Joost C.M. Holthuis, Outi Mäkitie
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Research Article Endocrinology Genetics

Osteoporosis and skeletal dysplasia caused by pathogenic variants in SGMS2

Abstract

Mechanisms leading to osteoporosis are incompletely understood. Genetic disorders with skeletal fragility provide insight into metabolic pathways contributing to bone strength. We evaluated 6 families with rare skeletal phenotypes and osteoporosis by next-generation sequencing. In all the families, we identified a heterozygous variant in SGMS2, a gene prominently expressed in cortical bone and encoding the plasma membrane–resident sphingomyelin synthase SMS2. Four unrelated families shared the same nonsense variant, c.148C>T (p.Arg50*), whereas the other families had a missense variant, c.185T>G (p.Ile62Ser) or c.191T>G (p.Met64Arg). Subjects with p.Arg50* presented with childhood-onset osteoporosis with or without cranial sclerosis. Patients with p.Ile62Ser or p.Met64Arg had a more severe presentation, with neonatal fractures, severe short stature, and spondylometaphyseal dysplasia. Several subjects had experienced peripheral facial nerve palsy or other neurological manifestations. Bone biopsies showed markedly altered bone material characteristics, including defective bone mineralization. Osteoclast formation and function in vitro was normal. While the p.Arg50* mutation yielded a catalytically inactive enzyme, p.Ile62Ser and p.Met64Arg each enhanced the rate of de novo sphingomyelin production by blocking export of a functional enzyme from the endoplasmic reticulum. SGMS2 pathogenic variants underlie a spectrum of skeletal conditions, ranging from isolated osteoporosis to complex skeletal dysplasia, suggesting a critical role for plasma membrane–bound sphingomyelin metabolism in skeletal homeostasis.

Authors

Minna Pekkinen, Paulien A. Terhal, Lorenzo D. Botto, Petra Henning, Riikka E. Mäkitie, Paul Roschger, Amrita Jain, Matthijs Kol, Matti A. Kjellberg, Eleftherios P. Paschalis, Koen van Gassen, Mary Murray, Pinar Bayrak-Toydemir, Maria K. Magnusson, Judith Jans, Mehran Kausar, John C. Carey, Pentti Somerharju, Ulf H. Lerner, Vesa M. Olkkonen, Klaus Klaushofer, Joost C.M. Holthuis, Outi Mäkitie

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Figure 7

Effect of SGMS2 pathogenic variants on SMS2 catalytic activity and sphingomyelin biosynthesis.

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Effect of SGMS2 pathogenic variants on SMS2 catalytic activity and sphin...
(A) Top: TLC analysis of reaction products formed when lysates of yeast strains expressing WT or mutant versions of V5-tagged human SMS2 were incubated with C6-NBD-ceramide for 20 minutes at 37°C. Note that SMS2I62S and SMS2M64R produce similar amounts of NBD-sphingomyelin (NBD-SM) as WT-SMS2, whereas SMS2R50X and the active site mutants SMS2D276E and SMS2D276A lack sphingomyelin synthase activity. EV denotes yeast lysate from strain transfected with empty vector. Bottom: SMS2 levels in yeast lysates were analyzed by immunoblotting using anti-V5 antibody. (B) Total membranes of control (WT) or patient-derived fibroblasts carrying heterozygous p.Arg50*, p.Ile62Ser, or p.Met64Arg nonsense and missense variants were subjected to immunoblot analysis using anti-SMS2 and anti-calnexin antibodies. Immunoblot of total membranes from yeast transfected with empty vector (EV) or SMS2-encoding plasmid was stained with anti-SMS2 antibody and served as control. (C) TLC analysis of lipid extracts from control (WT) or patient-derived fibroblasts carrying heterozygous p.Ile62Ser or p.Met64Arg missense variants grown for 4 hours in medium supplemented with 14C-choline. Incorporation of 14C-choline into phosphatidylcholine (PC) and sphingomyelin (SM) was analyzed by autoradiography (top). Lipids were stained with iodine vapor to verify that total lipid content between extracts was comparable (bottom). (D) Quantitation of 14C-choline incorporation into sphingomyelin in control or patient-derived fibroblasts treated as in C. Data are mean ± SD; n ≥ 3. (E) Sphingomyelin, phosphatidylcholine, and triacylglycerol (TAG) levels in total lipid extracts of control or patient-derived fibroblasts were determined by LC-MS/MS and expressed as mole percent of total phospholipid analyzed. Data are mean ± SD; n = 3.