Involvement and abnormality of lysosphingolipid metabolic enzymes in malignant diseases

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The gene expression of human sphingosine 1-phosphate lyase (SPL) catalyzing sphingosine 1-phosphate (S1P) remains to be determined. Among 5 human lung cancer cell lines examined, SPL protein levels paralleled the respective mRNA and enzyme activities. Between H1155 and H1299 cells used for further experiments, lower cellular S1P was observed in H1155 with higher SPL activity compared with H1299 with lower SPL activity. Previous reprots by others have shown that GATA-4 affect SPL transcription in Dictyostelium discoideum. GATA-4 was observed in H1155 but not in other cell lines. Promoter analysis of human SPL revealed that the most important region was located between -136 bp and -88 bp from the first exon, where 2 Sp1 sites exist but no GATA site. Electrophoresis mobility shift assay (EMSA), chromatin immunoprecipitation (ChIP) assay, reporter assay using reporter with mutated binding motif suggest the major role of Sp1 in SPL transcription and no direct binding of GATA-4 with this 5' promoter region. The collive role of GATA-4 wasproved by showing co-immunoprecipitation of Sp1 and GATA-4. Thus, high SPL transcription of H1155 cells was regulated by Sp1 and GATA-4/Sp1 complex formation, both of which bind to Sp1 sites of the 5' -SPL promoter.