Isolation and characterization of an Arabidopsis biotin carboxylase gene and its promoter.
In the plastids of most plants, acetyl-CoA carboxylase (ACCase; EC 6.4.1.2) is a multisubunit complex consisting of biotin carboxylase (BC), biotin-carboxyl carrier protien (BCCP), and carboxytransferase (alpha-CT, beta-CT) subunits. To better understand the regulation of this enzyme, we have isolated and sequenced a BC genomic clone from Arabidopsis and partially characterized its promoter. Fifteen introns were identified. The deduced amino acid sequence of the mature BC protein is highly conserved between Arabidopsis and tobacco (92.6% identity). BC expression was evaluated using northern blots and BC/GUS fusion constructs in transgenic Arabidopsis. GUS activity in the BC/GUS transgenics as well as transcript level of the native gene were both found to be higher in silique and flower than in root and leaf. Analysis of tobacco suspension cells transformed with truncated BC promoter/GUS gene fusions indicated the region from -140 to +147 contained necessary promoter elements which supported basal gene expression. A positive regulatory region was found to be located between -2100 and -140, whereas a negative element was possibly located in the first intron. In addition, several conserved regulatory elements were identified in the BC promoter. Surprisingly, although BC is a low-abundance protein, the expression of BC/GUS fusion constructs was similar to 35S/GUS constructs.
Bao
X
X
Department of Botany and Pathology, Michigan State University, East Lansing 48824, USA.
Shorrosh
B S
BS
Ohlrogge
J B
JB
D000595:Amino Acid Sequence; D017360:Arabidopsis / Q000201:enzymology / Q000235:genetics*; D001483:Base Sequence; D019731:Carbon-Nitrogen Ligases / Q000235:genetics*; D003001:Cloning, Molecular; D018506:Gene Expression Regulation, Plant / Q000235:genetics; D017343:Genes, Plant / Q000235:genetics*; D017930:Genes, Reporter; D005966:Glucuronidase / Q000235:genetics / Q000378:metabolism; D007438:Introns; D008969:Molecular Sequence Data; D030821:Plants, Genetically Modified; D011401:Promoter Regions, Genetic / Q000235:genetics*; D012333:RNA, Messenger / Q000032:analysis; D018749:RNA, Plant / Q000032:analysis; D011993:Recombinant Fusion Proteins; D017422:Sequence Analysis, DNA; D017386:Sequence Homology, Amino Acid
D012333:RNA, Messenger; D018749:RNA, Plant; D011993:Recombinant Fusion Proteins; D005966:Glucuronidase; D019731:Carbon-Nitrogen Ligases; C019791:biotin carboxylase