Regulation of the expression of intracellular beta-carbonic anhydrase in response to CO2 and light in the marine diatom Phaeodactylum tricornutum.

Abstract

Cells of the marine diatom Phaeodactylum tricornutum Bohlin (UTEX 642) grown in 5% CO(2) were transferred to air-level CO(2) in the light or dark and allowed to acclimate to air. No accumulation of the transcript of the P. tricornutum beta-carbonic anhydrase 1 (ptca1) was detected in 5% CO(2)-grown cells, but ptca1 mRNA accumulated and reached a peak after 6 h acclimation to air but decreased over the next 18 h. A similar accumulation time course was observed in cells air-acclimated in the dark, except that levels of mRNA were <50% those in the light. These results suggest that air-level [CO(2)] is required to trigger the transcription of ptca1 and that light affects the extent of acclimation. During acclimation to air for 120 h in the light, levels of ptca1 mRNA exhibited a periodic oscillation with a cycle of about 24 h, which, however, was not reflected in protein accumulation levels. A 5'-upstream region from the transcription-start site toward -1,292 bp of ptca1 was cloned by inverse polymerase chain reaction, and 5'-truncations were carried out on this fragment. The truncated promoter regions were fused with the beta-glucuronidase gene (uidA) and introduced into P. tricornutum. The promoter fragments, truncated at positions -1,292, -824, -484, -225, and -70 bp, conferred on transformants clear CO(2)-responsive beta-glucuronidase expressions. In contrast, the CO(2)-responsive regulation was severely impaired or completely abolished by truncations, respectively, at position -50 or -30 bp. These results indicate that critical cis-elements required for CO(2)-responsive transcription of ptca1 may be located between -70 and -30 bp relative to the transcription start site.