PARENT SESSION
Posters P4C Controling CO₂: Stomates and carbon concentrating mechanisms. Abstracts (631-642)

CO₂-responsive-promoter region of intracellular -carbonic anhydrase gene in the marine diatom Phaeodactylum tricornutum. Hisashi Harada^*,1, Maki Ishida¹, Yusuke Matsuda¹, ¹ Graduate School of Science, Kwansei-Gakuin Univ., 2-1 Gakuen, Sanda, Hyogo, Japan

ABSTRACT- Diatoms are one of the major photoautotrophic organisms in the hydrosphere. They are considered to play a key role in global primary production and hence in global cycles of carbon and other major inorganic nutrients. Similarly to many other microscopic algae, some of marine and freshwater diatoms so far studied have been reported to possess "inorganic carbon concentrating mechanism (CCM)", which confers cells to do high-affinity photosynthesis for dissolved inorganic carbon (DIC). It was reported that the marine diatom, Phaeodactylum tricornutum, possesses fairly efficient HCO₃^- transport and that intracellular -type carbonic anhydrase (CA) is essential to acquire the high-affinity photosynthesis. The expression of CCM and CA are reported to be regulated primarily by [CO₂] in the bulk medium rather than [HCO₃^-] or [CO₃^2-]. In this study, we investigated the expression mechanism of -CA as a model molecule which is regulated by [CO₂]. 1.3 kbp of 5′-upstream region of P. tricornutum -CA (PtCA) gene was cloned by inverse PCR. Various intermediate length of 5′-upstream region from 30 bp to 1.3 kbp was fused with GUS gene as a reporter and these chimeric constructs were introduced into P. tricornutum cells by microparticle bombardment. 10-100 Zeocin-resistant colonies were recovered from 10⁸ cells and GUS activity was detected in about 50% these transformants. DNA-gel blot analysis indicated that these transformants possess 1-4 copies of GUS gene. The GUS activity exhibited in air-grown transformants was 15-25 fold that in 5% CO₂-grown transformants. Such CO₂-responsible GUS expression was observed in transformants that contain the insertion of 5′-upstream region of 70 bp at the shortest. The critical cis-elements of -CA promoter which are required for CO₂ responsibility of -CA gene expression were further investigated.

KEY WORDS: carbon concentrating mechanism, transcriptional regulation, Marine Diatom, carbonic anhydrase