鞘氨酸 1-磷酸鹽接受器-1上酪胺酸硫酸化現象對於內皮細胞移行能力之研究

鞘氨酸 1-磷酸鹽（Sphingosine 1-phosphate, S1P）是一種小分子磷脂質類生長因子，主要由發炎反應過程中活化的血小板所分泌，可調控許多重要的細胞生理功能，如促使細胞增生、分化與移行。而許多癌細胞在生長的過程中，也會釋放出S1P調控腫瘤的生成，並促使血管新生。S1P會與細胞上內皮細胞分化基因（edg）族中的G蛋白接合型受器接合，藉此調節多種訊息傳遞路徑。已知內皮細胞會表現三種已知Edg受器－Edg1（S1P1）、Edg2、和Edg3（S1P3），其中S1P1對於細胞的爬行、血管的生成和哺乳類胚胎發育扮演著重要的角色。S1P1於N端具有可被硫酸化的酪胺酸，並有研究指出，S1P1的硫酸化可調控T細胞的增生與移行。然而，S1P1的酪胺酸硫酸化是否調控內皮細胞生理反應尚未清楚。在本研究中，利用可抑制酪胺酸硫酸化的抑制劑（sodium chlorate）及wild-type S1P1和mutant (Y19, 22F) S1P1表現質體，以了解細胞中S1P1的酪胺酸硫酸化現象是否調節其生理功能，如ICAM-1表現量、Src的磷酸化。結果發現抑制細胞硫酸化會減少S1P活化Src的情形，但不影響S1P調節ICAM-1在細胞膜上的表現量。另外，S1P所誘導的細胞移行能力、肌動蛋白的重組與層狀足的生成，也會經由S1P1的酪胺酸硫酸化現象來調控。而二維膠體電泳的結果中找到28個蛋白質會受S1P1的酪胺酸硫酸化現象所影響。此研究指出S1P1上第19與22個位置的酪胺酸硫酸化現象對於內皮細胞生理功能反應，可能具有重要的生理功能。

關鍵字

鞘氨酸 1-磷酸鹽；鞘氨酸 1-磷酸鹽接受器-1 ；酪胺酸硫酸化；內皮細胞；細胞爬行

並列摘要

Sphingosine 1-phosphate (S1P) is a lysophospholipid growth factor, which has been implicated in regulating various cellular behaviors. The cellular effects of S1P are mediated by binding to a family of G protein-coupled receptors (GPCRs), designated S1P1 through S1P5. By siRNA approaches, we have demonstrated that S1P1 mediated multiple important cellular functions, such as the phosphorylation of Src kinase, the expression of ICAM-1, IL-8 and MCP-1 in human umbilical vein endothelial cells (HUVECs). It has recently been shown that the O-sulfation of the tyrosine residues at N-terminal of S1P1 catalyzed by ATP-sulfurylase plays an important role for S1P1 mediated signaling. However, the molecular mechanisms of S1P1 sulfation-regulated cell functions remain unknown. In the present study, pre-incubation of 10 mM sodium chlorate, an inhibitor of ATP-sulfurylase, significantly inhibited S1P-stimulated Src phosphorylation but not ICAM-1 expression in HUVECs. In cell migration assay, S1P regulated cell migration, actin rearrangement and lamellipodia formation was affected by tyrosine sulfation of S1P1. Furthermore, by two dimensional SDS-PAGE of S1P- and sodium chlorate-treated HUVEC lysates, we have identified 28 proteins were affected by sulfated S1P1 through analysis using Image Master Software. These results imply that sulfation of the tyrosine residues at N-terminal of S1P1 might play important roles on S1P1 mediated signaling in HUVECs.

並列關鍵字

Sphingosine 1-phosphate ； S1P1 ； Tyrosine sulfation ； Endothelial Cells, Migration

參考文獻

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2. Bayless, K. J., and Davis, G. E. (2003) Sphingosine-1-phosphate markedly induces matrix metalloproteinase and integrin-dependent human endothelial cell invasion and lumen formation in three-dimensional collagen and fibrin matrices. Biochem. Biophys. Res. Commun. 312, 903–913.

3. Bettelheim, F. R. (1954) Tyrosine-O-sulfate in a peptide from fibrinogen. J. Am. Chem. Soc. 76, 2838–2839.

4. Chun, J., Goetzl, E., Hla, T., Ingarashi, Y., Lynch, K., Moolenaar, W., Pyne, S., and Tigyi, G. (2002) International Union of Pharmacology. XXXIV. Lysophospholipid receptor nomenclature. Pharmacol. Rev. 54, 265–269.

5. Coussens, L. M., and Werb, Z. (2002) Inflammation and cancer. Nature 420, 19–26.

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鞘氨酸 1-磷酸鹽接受器-1上酪胺酸硫酸化現象對於內皮細胞移行能力之研究

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