Two low-temperature-inducible Chlorella genes for delta12 and omega-3 fatty acid desaturase (FAD): isolation of delta12 and omega-3 fad cDNA clones, expression of delta12 fad in Saccharomyces cerevisiae, and expression of omega-3 fad in Nicotiana tabacum.
In an attempt to clarify the involvement of fatty acid desaturases (FADs) in the freezing tolerance of Chlorella vulgaris IAM C-27, developed by hardening, we have isolated cDNA clones for two types of FADs from the Chlorella strain, based on the sequence information of genes for delta12 and omega-3 FADs, respectively desaturating oleic acid (18:1) to linoleic acid (18:2) and linoleic acid (18:2) to linolenic acid (18:3). The deduced amino acid sequence of the first clone, designated CvFad2, showed about 66% similarity to the microsomal delta12 FADs from several higher plants and this gene had delta12 FAD activity when expressed in Saccharomyces cerevisiae. The predicted protein encoded by a second gene, designated CvFad3, showed about 60% similarity to the microsomal and plastidial omega-3 FADs from several higher plants. The features of the amino acid sequences of the C- and N-terminal regions of CvFAD3 and fatty acid analysis of polar lipids in transgenic tobacco plant expressing the CvFad3 gene suggested that this gene encodes the microsomal omega-3 FAD. Southern blot analysis showed that both genes were single-copy genes in the genome of the Chlorella strain. Different transcriptional patterns were observed with the two genes during hardening in Northern blot analysis.
Suga
Koushirou
K
Division of Bioresource and Bioenvironmental Sciences, Graduate School, Kyushu University, Fukuoka, Japan.
Honjoh
Ken-ichi
K
Furuya
Naoki
N
Shimizu
Hideyuki
H
Nishi
Koutarou
K
Shinohara
Fuminori
F
Hirabaru
Yoshie
Y
Maruyama
Isao
I
Miyamoto
Takahisa
T
Hatano
Shoji
S
Iio
Masayoshi
M
D000595:Amino Acid Sequence; D001483:Base Sequence; D015152:Blotting, Northern; D015139:Blotting, Southern; D002708:Chlorella / Q000201:enzymology* / Q000235:genetics*; D002849:Chromatography, Gas; D003001:Cloning, Molecular; D018076:DNA, Complementary / Q000235:genetics*; D044943:Fatty Acid Desaturases / Q000737:chemistry / Q000235:genetics* / Q000378:metabolism; D005227:Fatty Acids / Q000032:analysis; D015870:Gene Expression; D008969:Molecular Sequence Data; D010802:Phylogeny; D030821:Plants, Genetically Modified; D012333:RNA, Messenger / Q000235:genetics / Q000378:metabolism; D011994:Recombinant Proteins / Q000235:genetics; D012441:Saccharomyces cerevisiae / Q000737:chemistry / Q000235:genetics*; D013696:Temperature; D014026:Nicotiana / Q000737:chemistry / Q000235:genetics*
D018076:DNA, Complementary; D005227:Fatty Acids; D012333:RNA, Messenger; D011994:Recombinant Proteins; D044943:Fatty Acid Desaturases