AtCRCT1 and AtCRCT2, homologs of CO2 responsive CCT protein in <i>Arabidopsis thaliana</i> , are involved in the regulation of starch synthesis-related gene expression

CO2-Responsive CCT Protein (CRCT) is a transcription factor that regulates the expression of starch synthesis-related genes, and consequently starch content in the vegetative organs of rice. Two CRCT homologues, AtCRCT1 and AtCRCT2, were identified in the genome of Arabidopsis thaliana. AtCRCT1 and AtCRCT2 were expressed in most organs of the plant, and AtCRCT2 in particular was expressed in their vascular bundles. The expression of these genes was markedly upregulated by sucrose treatment and tended to increase under elevated CO2 condition. The overexpression or knockout of these genes did not have a notable effect on the growth of Arabidopsis. On the other hand, the expression of some starch synthesis-related genes, such as ADP-glucose pyrophosphorylase large subunit3 (AtAPL3) and glucose 6-phosphate/Pi transporter2 (AtGPT2) was significantly upregulated in the overexpression lines. Accordingly, the expression of AtAPL3 and AtGPT2 in the double knockout line was significantly lower than that in wild-type (Col-0) under sucrose treatment. In addition, the starch content of the double knockout line at the end of the night was slightly lower than that of Col-0. Yeast two-hybrid and BiFC analyses demonstrated that both AtCRCT1 and AtCRCT2 interact with Growth Regulation Factor 7, a 14-3-3 protein. These results suggest that AtCRCT1 and AtCRCT2 have similar molecular and physiological functions to CRCT in rice and regulate the expression of some starch synthesis-related genes. However, their effects are limited and cannot markedly affect starch content, unlike CRCT in rice.