Characterization of an Arabidopsis cDNA encoding a subunit of serine palmitoyltransferase, the initial enzyme in sphingolipid biosynthesis.
Serine palmitoyltransferase (SPT; EC 2.3.1.50) catalyzes the condensation of serine with palmitoyl-CoA to form 3-ketosphinganine in the first step of de novo sphingolipid biosynthesis. In this study, we describe the cloning and functional characterization of a cDNA from Arabidopsis thaliana encoding the LCB2 subunit of SPT. The Arabidopsis LCB2 (AtLCB2) cDNA contains an open reading frame of 1,467 nucleotides, encoding 489 amino acids. The predicted polypeptide contains three transmembrane helices and a highly conserved motif involved in pyridoxal phosphate binding. Expression of this open reading frame in the Saccharomyces cerevisiae mutant strains defective in SPT activity resulted in the expression of a significant level of sphinganine, suggesting that AtLCB2 cDNA encodes SPT. Southern blot analysis and inspection of the complete Arabidopsis genome sequence database suggest that there is a second LCB2-like gene in Arabidopsis. Expression of a green fluorescent protein (GFP) fusion product in suspension-cultured tobacco BY-2 cells showed that AtLCB2 is localized to the endoplasmic reticulum. AtLCB2 cDNA may be used to study how sphingolipid synthesis is regulated in higher plants.
Tamura
K
K
Department of Biology, Graduate School of Natural Science, Konan University, Kobe, 658-8501 Japan.
Mitsuhashi
N
N
Hara-Nishimura
I
I
Imai
H
H
D000217:Acyltransferases / Q000737:chemistry / Q000235:genetics* / Q000378:metabolism; D000595:Amino Acid Sequence; D017360:Arabidopsis / Q000201:enzymology / Q000235:genetics*; D018076:DNA, Complementary; D018744:DNA, Plant; D004721:Endoplasmic Reticulum / Q000378:metabolism; D015964:Gene Expression Regulation, Bacterial; D008969:Molecular Sequence Data; D016296:Mutagenesis; D016366:Open Reading Frames; D010171:Palmitoyl Coenzyme A / Q000737:chemistry / Q000378:metabolism; D011487:Protein Conformation; D012441:Saccharomyces cerevisiae; D012694:Serine / Q000378:metabolism; D051102:Serine C-Palmitoyltransferase; D013107:Sphingolipids / Q000096:biosynthesis* / Q000737:chemistry / Q000378:metabolism; D013110:Sphingosine / Q000031:analogs & derivatives* / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D014026:Nicotiana / Q000235:genetics / Q000378:metabolism
D018076:DNA, Complementary; D018744:DNA, Plant; D013107:Sphingolipids; D010171:Palmitoyl Coenzyme A; D012694:Serine; D000217:Acyltransferases; D051102:Serine C-Palmitoyltransferase; D013110:Sphingosine; C005682:safingol