Defective O-Glycosylation due to a Novel Homozygous S129P Mutation Is Associated with Lack of Fibroblast Growth Factor 23 Secretion and Tumoral Calcinosis
Defective O-Glycosylation due to a Novel Homozygous S129P Mutation Is Associated with Lack of Fibroblast Growth Factor 23 Secretion and Tumoral Calcinosis
复制标题
DOI：
10.1210/jc.2009-0961
复制
发表时间：
2009-11-01
影响因子：
5.8
通讯作者：
Jueppner, Harald
中科院分区：
文献类型：
--
作者：
Bergwitz, Clemens;Banerjee, Santanu;Jueppner, Harald
背景资料：成纤维细胞生长因子（FGF 23）的纯合突变最近被描述为一种形式的高磷血症肿瘤钙质沉着症（HFTC）的遗传原因。然而，目前尚不清楚这些突变如何导致循环中生物活性FGF 23的丧失。FGF 23外显子2中的385 T> C，其在先前描述的受HFTC影响的个体中将密码子129从丝氨酸改变为脯氨酸（S129 P）。将S129 P突变以及两种已知的FGF 23突变S71 G和S129 F引入编码野生型（wt）人（h）FGF 23的表达载体中，以产生[P129] hFGF 23、[F129] hFGF 23和[G71] hFGF 23;完整裂解物，糖蛋白级分，使用亲和纯化的山羊抗-hFGF 23（51-69）和抗-hFGF 23（51-69）对来自表达这些构建体的HEK 293和COS-7细胞的条件培养基进行Western印迹分析。结果：我们在表达wt-hFGF 23的HEK 293细胞的总裂解物中检测到25-和32-kDa蛋白质种类。代表O-糖基化hFGF 23的32-kDa条带在来自表达[P129] hFGF 23的HEK 293细胞的裂解物的糖蛋白级分中检测不到，并且与wt-FGF 23相比，仅少量的[P129] hFGF 23分泌到培养基中。[G71] hFGF 23和[F129] hFGF 23表达细胞的结果相似。结论：我们的数据首次直接表明HFTC相关的FGF 23突变在体外损害了O-糖基化，导致突变激素分泌不良，从而解释了体内纯合子携带者的特征性高磷血症表型。（临床内分泌代谢杂志94：4267-4274，2009）
Background: Homozygous mutations in fibroblast growth factor (FGF23) have recently been described as the genetic cause of one form of hyperphosphatemic tumoral calcinosis (HFTC). However, it remained unclear to date how these mutations lead to loss of biologically active FGF23 in the circulation.Methods: We here report a novel homozygous mutation, c. 385T > C in FGF23 exon 2, which changes codon 129 from serine to proline (S129P) in a previously described individual affected by HFTC. The S129P mutation as well as two known FGF23 mutations, S71G and S129F, were introduced into an expression vector encoding wild-type (wt) human (h) FGF23 to yield [P129]hFGF23, [F129]hFGF23, and [G71] hFGF23; whole lysates, glycoprotein fractions, and conditioned media from HEK293 and COS-7 cells expressing these constructs were subjected to Western blot analysis using affinity-purified goat anti-hFGF23(51-69) and anti-hFGF23(206-222) antibodies.Results: We detected 25-and 32-kDa protein species in total lysates of HEK293 cells expressing wt-hFGF23. The 32-kDa band, representing O-glycosylated hFGF23, was not detectable in the glycoprotein fraction of lysates from HEK293 cells expressing [P129] hFGF23, and in comparison with wt-FGF23 only small amounts of [P129] hFGF23 were secreted into the medium. Similar results were obtained for cells expressing [G71] hFGF23 and [F129] hFGF23.Conclusion: Our data for the first time directly show that FGF23 mutations associated with HFTC impair O-glycosylation in vitro resulting in poor secretion of the mutant hormone thereby explaining the characteristic hyperphosphatemic phenotype of homozygous carriers in vivo. (J Clin Endocrinol Metab 94: 4267-4274, 2009)