Stereoselectivity of induction of the retinoblastoma gene product (pRb) dephosphorylation by D-erythro-sphingosine supports a role for pRb in growth suppression by sphingosine.
Sphingosine has been shown to inhibit cell growth in many cell lines although the mechanism of this effect remains obscure. More recently, D-erythro-sphingosine has been shown to act as an early inducer of dephosphorylation of the retinoblastoma gene product (pRb) in the lymphoblastic leukemia cell line MOLT-4 [Chao, R., Khan, W., & Hannun, Y.A. (1992) J. Biol. Chem., 267, 23459-23462]. In the current study, the role of the natural D-erythro-sphingosine in regulation of cell growth and pRb dephosphorylation was evaluated using chemically synthesized pure isomers of sphingosine. Of the four possible stereoisomers of sphingosine, D-erythro-sphingosine was most active in inducing dephosphorylation of pRb protein with an EC50% of 0.6 microM whereas its enantiomer L-erythro-sphingosine was 8-fold less potent with an EC50% of 5 microM. The dose responses for inhibition of cell growth were nearly identical to the EC50% for pRb dephosphorylation with D-erythro-sphingosine causing 50% inhibition at 0.6 microM whereas L-erythro-sphingosine was 5-6-fold less potent. All of the stereoisomers were taken up by the cells, and the greater potency of D-erythro-sphingosine was not due to differences in cellular uptake. The metabolism of D-erythro-sphingosine was also studied to evaluate the possible role of sphingosine metabolites on regulation of retinoblastoma protein. Evidence is provided against a role for ceramide or sphingosine 1-phosphate as mediators of the effects of sphingosine on pRb dephosphorylation.(ABSTRACT TRUNCATED AT 250 WORDS)
Department of Medicine, Duke University Medical Center, Durham, North Carolina 27710.
M
M
Pushkareva
R
R
Chao
A
A
Bielawska
A H
AH
Merrill
H M
HM
Crane
B
B
Lagu
D
D
Liotta
Y A
YA
Hannun
D016923:Cell Death / Q000187:drug effects; D002455:Cell Division / Q000187:drug effects*; D004357:Drug Synergism; D037341:Fumonisins; D007700:Kinetics; D008246:Lysophospholipids; D009183:Mycotoxins / Q000494:pharmacology; D010766:Phosphorylation; D054198:Precursor Cell Lymphoblastic Leukemia-Lymphoma; D016160:Retinoblastoma Protein / Q000378:metabolism*; D013110:Sphingosine / Q000031:analogs & derivatives / Q000737:chemistry / Q000378:metabolism / Q000494:pharmacology*; D013237:Stereoisomerism; D014407:Tumor Cells, Cultured
D037341:Fumonisins; D008246:Lysophospholipids; D009183:Mycotoxins; D016160:Retinoblastoma Protein; C060506:sphingosine 1-phosphate; C056933:fumonisin B1; D013110:Sphingosine