A MAMMALIAN HOMOLOG OF THE YEAST LCB1 ENCODES A COMPONENT OF SERINE PALMITOYLTRANSFERASE, THE ENZYME CATALYZING THE FIRST STEP IN SPHINGOLIPID SYNTHESIS

Serine palmitoyltransferase (SPT; EC 2.3.1.50) catalyzes the initial s tep dedicated to sphingolipid biosynthesis and is thought to be a key enzyme for regulating cellular sphingolipid content. For SPT activity, the yeast Saccharomyces cerevisiae requires two genes, LCB1 and LCB2. We isolated mammalian LCB1 cDNA homologs from mouse and Chinese hamst er ovary (CHO) cells and an LCB2 cDNA homolog from CHO cells. The mamm alian LCB1 proteins are predicted to have about 35% amino acid identit y to the yeast Lcb1 protein, whereas the CHO LCB2 protein is predicted to have about 40% amino acid identity to the yeast Lcb2 protein. Nort hern blot analysis of mRNA isolated from various mouse tissues reveale d that the tissue distribution of both LCB1 and LCB2 messengers follow ed a similar pattern. Transfection of an SPT-defective CHO mutant stra in with a CHO LCB1-expressing plasmid restored both SPT activity and d e novo sphingolipid synthesis to the wild type levels, whereas transfe ction of the mutant strain with a CHO LCB2-expressing plasmid did not exhibit any recovery effects, indicating that the SPT defect in the mu tant cells is specifically complemented by the CHO LCB1 homolog. Furth ermore, when the SPT-defective mutant cells were transfected with a pl asmid encoding a His(6)-tagged CHO LCB1 protein, SPT activity bound to a Ni2+-immobilized resin. These results indicate that the CHO LCB1 ho molog encodes a component of SPT.