Involvement of N-terminal-extended form of sphingosine kinase 2 in serum-dependent regulation of cell proliferation and apoptosis
Involvement of N-terminal-extended form of sphingosine kinase 2 in serum-dependent regulation of cell proliferation and apoptosis
复制标题
DOI：
10.1074/jbc.m504507200
复制
发表时间：
2005-10-28
影响因子：
4.8
通讯作者：
Nakamura, S
中科院分区：
文献类型：
--
作者：
Okada, T;Ding, G;Nakamura, S
鞘氨醇激酶（SPHK）1通过催化重要的生物活性信使鞘氨醇1-磷酸的形成而参与细胞增殖和抗凋亡过程的调节。与SPHK 1的增殖作用不同，另一种同工酶SPHK 2已被证明具有抗增殖或促凋亡作用。然而，SPHK 2作用的分子机制在很大程度上是未知的。本研究通过与最初报道的SPHK 2-S的比较来表征SPHK 2的N-末端延伸形式（SPHK 2-L）。实时荧光定量PCR分析显示SPHK 2-L mRNA在多种人类细胞系和组织中以表达为主。从序列分析得出结论，SPHK 2-L是一种物种特异性的同种型，在人类中表达，但在小鼠中不表达。在蛋白质水平上，免疫沉淀研究已经证明SPHK 2-L是人肝癌HepG 2细胞中的主要同种型。SPHK 2-L在人胚肾（HEK）293细胞中表达时，在血清存在下不显示任何DNA合成抑制，而在不存在血清时显示显著抑制。此外，血清剥夺导致SPHK 2-L易位到细胞核中。此外，血清剥夺诱导SPHK 2-L在HEK 293细胞中的表达。此外，通过小干扰RNA处理抑制SPHK 2可以防止HEK 293细胞中血清剥夺或药物诱导的细胞凋亡。两者合计，这些结果表明，SPHK 2剪接变体，SPHK 2-L，在人类细胞中的主要形式不抑制DNA合成在正常条件下，和SPHK 2- L的积累在细胞核中诱导的血清剥夺可能参与细胞增殖或凋亡的停止取决于细胞类型。
Sphingosine kinase (SPHK) 1 is implicated in the regulation of cell proliferation and anti-apoptotic processes by catalyzing the formation of an important bioactive messenger, sphingosine 1-phosphate. Unlike the proliferative action of SPHK1, another isozyme, SPHK2, has been shown to possess anti-proliferative or pro-apoptotic action. Molecular mechanisms of SPHK2 action, however, are largely unknown. The present studies were undertaken to characterize the N-terminal-extended form of SPHK2 (SPHK2-L) by comparing it with the originally reported form, SPHK2-S. Real-time quantitative PCR analysis revealed that SPHK2-L mRNA is the major form in several human cell lines and tissues. From sequence analyses it was concluded that SPHK2-L is a species-specific isoform that is expressed in human but not in mouse. At the protein level it has been demonstrated by immunoprecipitation studies that SPHK2-L is the major isoform in human hepatoma HepG2 cells. SPHK2-L, when expressed in human embryonic kidney (HEK) 293 cells, did not show any inhibition of DNA synthesis in the presence of serum, whereas it showed marked inhibition in the absence of serum. Moreover, serum deprivation resulted in the translocation of SPHK2-L into the nuclei. In addition, serum deprivation induced SPHK2-L expression in HEK293 cells. Furthermore, suppression of SPHK2 by small interfering RNA treatment prevented serum deprivation- or drug-induced apoptosis in HEK293 cells. Taken together, these results indicate that a major form of SPHK2 splice variant, SPHK2-L, in human cells does not inhibit DNA synthesis under normal conditions and that SPHK2- L accumulation in the nucleus induced by serum deprivation may be involved in the cessation of cell proliferation or apoptosis depending on the cell type.