Sulf loss influences N-, 2-O-, and 6-O-sulfation of multiple heparan sulfate proteoglycans and modulates fibroblast growth factor signaling
Sulf loss influences N-, 2-O-, and 6-O-sulfation of multiple heparan sulfate proteoglycans and modulates fibroblast growth factor signaling
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DOI：
10.1074/jbc.m802130200
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发表时间：
2008-10-10
影响因子：
4.8
通讯作者：
Dierks, Thomas
中科院分区：
文献类型：
--
作者：
Lamanna, William C.;Frese, Marc-Andre;Dierks, Thomas
Sulf 1和Sulf 2是两种硫酸乙酰肝素6-O-内硫酸酯酶，调节多种生长因子的活性，如成纤维细胞生长因子和Wnt，并且对哺乳动物的发育和生存至关重要。在这项研究中，哺乳动物Sulfs功能的特点是使用过表达细胞系，体外酶测定，并在体内Sulf敲除细胞模型。亚细胞Sulf定位的分析揭示了显着差异的酶分泌和洗涤剂的溶解度之间的人类亚型和他们以前的特点鹌鹑直系同源物。此外，在体外测定了Sulf对其天然硫酸乙酰肝素底物的活性，证明了对S结构域相关的6S二糖的有限特异性和不能修饰过渡区相关的UA-GlcNAc（6S）。硫酸乙酰肝素组合物的分析，从不同的细胞表面，脱落，糖基磷脂酰肌醇锚定和细胞外基质蛋白聚糖组分的Sulf敲除细胞系建立的差异影响Sulf 1和/或Sulf 2损失的非底物N-，2-O-和6-O-硫酸基团。这些研究结果表明硫缺乏对HS生物合成机制的动态影响。真实的时间PCR分析证实了在Sulf敲除细胞系中Hs 2st和Hs 6st硫酸乙酰肝素磺基转移酶的差异表达。在功能上，由硫损失引起的硫酸乙酰肝素硫酸化的变化显示出对成纤维细胞生长因子信号传导的显著影响。总之，这项研究表明，硫参与了一个潜在的细胞反馈机制，其中它们编辑多个硫酸乙酰肝素蛋白聚糖的硫酸化，从而调节细胞信号传导和调节硫酸乙酰肝素生物合成酶的表达。
Sulf1 and Sulf2 are two heparan sulfate 6-O-endosulfatases that regulate the activity of multiple growth factors, such as fibroblast growth factor and Wnt, and are essential for mammalian development and survival. In this study, the mammalian Sulfs were functionally characterized using overexpressing cell lines, in vitro enzyme assays, and in vivo Sulf knock-out cell models. Analysis of subcellular Sulf localization revealed significant differences in enzyme secretion and detergent solubility between the human isoforms and their previously characterized quail orthologs. Further, the activity of the Sulfs toward their native heparan sulfate substrates was determined in vitro, demonstrating restricted specificity for S-domain-associated 6S disaccharides and an inability to modify transition zone-associated UA-GlcNAc( 6S). Analysis of heparan sulfate composition from different cell surface, shed, glycosylphosphatidylinositol- anchored and extracellular matrix proteoglycan fractions of Sulf knock-out cell lines established differential effects of Sulf1 and/or Sulf2 loss on nonsubstrate N-, 2-O-, and 6-O-sulfate groups. These findings indicate a dynamic influence of Sulf deficiency on the HS biosynthetic machinery. Real time PCR analysis substantiated differential expression of the Hs2st and Hs6st heparan sulfate sulfotransferase enzymes in the Sulf knock-out cell lines. Functionally, the changes in heparan sulfate sulfation resulting from Sulf loss were shown to elicit significant effects on fibroblast growth factor signaling. Taken together, this study implicates that the Sulfs are involved in a potential cellular feed-back mechanism, in which they edit the sulfation of multiple heparan sulfate proteoglycans, thereby regulating cellular signaling and modulating the expression of heparan sulfate biosynthetic enzymes.