Plant and Cell Physiology, 2002, Vol. 43, No. 6 660-667
© 2002 Oxford University Press
CO2 Response Element and Corresponding trans-acting Factor of the Promoter for Ribulose-1,5-bisphosphate Carboxylase/oxygenase Genes in Synechococcus sp. PCC7002 Found by an Improved Electrophoretic Mobility Shift Assay
1 Biological Sciences Department, Toray Research Center, Inc., 1111 Tebiro, Kamakura, Kanagawa, 248-8555 Japan
2 Technical Research Center, Kansai Electric Power Co., 3-11-20 Nakoji, Amagasaki, Hyogo, 661-0974 Japan
We analyzed the promoter of the genes encoding the ribulose-1,5-bisphosphate carboxylase/oxygenase (rbc) in the cyanobacterium Synechococcus sp. PCC7002 and localized the CO2-regulatory element. Cyanobacterial transformants were constructed with several DNA segments of the rbc promoter fused to the chloramphenicol acetyltransferase (CAT) gene, and their acetyltransferase activities were analyzed under 0.03% and 1% CO2 conditions. We found that the AT-rich element localized from –262 to –291 relative to the rbc translation-starting site was required for CO2-dependent repression. Fluorescent-labeled oligonucleotide probes of identical sequence to the AT-rich element were reacted with protein extracts from cells cultured under conditions of low and high CO2 atmospheric content. We detected a gel retardation complex of a strong signal intensity in extracts from cells cultured under 15% CO2, but only a weak signal from cells cultured under 1% CO2. Moreover, a DNA affinity precipitation assay identified a 16-kDa protein that bound to nucleotide sequences within the AT-rich element. The partial amino acid sequence of the protein was similar to the deduced protein sequences of ORF129 and ORF155 from Synechocystis 6803. Our findings suggest that the AT-rich element plays a role as a negative CO2-regulatory element and its trans-acting factor possibly regulates the rbc transcription in response to CO2 levels.
3 Corresponding author: E-mail, Takuo_Onizuka@trc.toray.co.jp; Fax, +81-467-320414.
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