Involvement of N-terminal-extended form of sphingosine kinase 2 in serum-dependent regulation of cell proliferation and apoptosis.
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.
Okada
Taro
T
Division of Biochemistry, Department of Molecular and Cellular Biology, Kobe University Graduate School of Medicine, Kobe 650-0017, Japan.
Ding
Guo
G
Sonoda
Hirofumi
H
Kajimoto
Taketoshi
T
Haga
Yuki
Y
Khosrowbeygi
Ali
A
Gao
Sanyang
S
Miwa
Noriko
N
Jahangeer
Saleem
S
Nakamura
Shun-Ichi
S
D000595:Amino Acid Sequence; D000818:Animals; D017209:Apoptosis; D001483:Base Sequence; D002460:Cell Line; D002467:Cell Nucleus / Q000378:metabolism; D049109:Cell Proliferation; D003001:Cloning, Molecular; D004247:DNA / Q000378:metabolism; D018076:DNA, Complementary / Q000378:metabolism; D004305:Dose-Response Relationship, Drug; D005091:Exons; D006367:HeLa Cells; D006801:Humans; D007150:Immunohistochemistry; D051379:Mice; D008969:Molecular Sequence Data; D017853:Phosphotransferases (Alcohol Group Acceptor) / Q000737:chemistry* / Q000502:physiology; D011485:Protein Binding; D020033:Protein Isoforms; D017434:Protein Structure, Tertiary; D012333:RNA, Messenger / Q000378:metabolism; D034741:RNA, Small Interfering / Q000378:metabolism; D020133:Reverse Transcriptase Polymerase Chain Reaction; D013045:Species Specificity; D014018:Tissue Distribution; D000099157:Sphingosine Kinase
D004247:DNA; D018076:DNA, Complementary; D017853:Phosphotransferases (Alcohol Group Acceptor); D020033:Protein Isoforms; D012333:RNA, Messenger; D034741:RNA, Small Interfering; D000099157:Sphingosine Kinase