Cloning and characterization of beta-carotene ketolase gene promoter in Haematococcus pluvialis.
The unicellular green alga Haematococcus pluvialis accumulates a highly valuable ketocarotenoid, astaxanthin, under various environmental stresses. beta-carotene ketolase (BKT) plays a key role in astaxanthin biosynthesis in H. pluvialis. In this paper, an approximate 700 bp 5'-flanking region of the bkt gene containing a putative promoter was cloned through walking upstream. The results of the sequence analysis showed that this bkt 5'-flanking region might have cis-acting elements such as sterol regulatory element (SRE-1)-like motifs, the C-repeat/dehydration responsive element (DRE) and al-3 proximal element (APE)-like motifs, except for typical TATA and CCAAT boxes. The results of the beta-galactosidase assay and the transient expression of lacZ driven by a series of sequential deletions revealed that a minimal promoter-like region might exist from -630 to -408 bp, and the highest promoter activity was observed to span the positions from -630 to -308 bp. The results of the site-directed mutagenesis of a C-repeat/DRE and two APE-like motifs in a promoter-like region (-630 to -308 bp) suggested that two APE-like motifs might be essential for transcriptional control of the bkt gene.
Meng
Chun-Xiao
CX
Institute of Oceanology, Chinese Academy of Sciences, Qingdao 266071, China.
Teng
Chang-Ying
CY
Jiang
Peng
P
Qin
Song
S
Tseng
Cheng-Kui
CK
D001426:Bacterial Proteins / Q000235:genetics* / Q000378:metabolism*; D001483:Base Sequence; D000460:Chlorophyta / Q000201:enzymology* / Q000235:genetics*; D003001:Cloning, Molecular / Q000379:methods*; D008969:Molecular Sequence Data; D010105:Oxygenases / Q000235:genetics* / Q000378:metabolism*; D011401:Promoter Regions, Genetic / Q000235:genetics*; D011994:Recombinant Proteins / Q000378:metabolism
D001426:Bacterial Proteins; D011994:Recombinant Proteins; D010105:Oxygenases; C093259:beta-carotene ketolase