Studies of light perception in marine diatoms and discovery of a novel blue light cryptochrome photoreceptor

The research project has focused on the structural and functional characterization of a gene denoted PtCPF1 (Cryptochrome/Photolyase Family 1) that encodes a putative photoreceptor in the diatom Phaeodactylum tricomutum. Cryptochromes (cry) are blue light receptors that share sequence similarity with photolyases, flavoproteins that catalyze the repair of UV light-damaged DNA. In order to characterize the diatom PtCPF1 gene, expression at both transcriptional and translational levels have been performed in time course experiments designed to study circadian rhythmicity and acute light induction responses. From this analysis PtCPF1 was shown to be strongly induced under blue light and to be expressed diurnally. In order to understand better the function of the gene product, the protein has been expressed and purified in E. coli. Spectral and biochemical analyses of the purified protein have shown that PtCPF1 is a blue-light-absorbing protein with DNA repair activity. On the other hand, localization studies in diatom cells have evidenced the constitutive nuclear localization of the protein. Interestingly, comparative analysis of the diatom PtCPF1 protein has revealed it to be more similar to the animal cryptochromes than to plant counterparts. Since animal crys act as components of the circadian clock controlling daily physiological and behavioural rhythms and as photoreceptors that mediate entrainment of the circadian clock to light, it was important to elucidate the function of the PtCPF1 protein both in a heterologous system and in an in vivo system. Remarkably, transcription assays developed in mammalian cells have evidenced a repressor activity of the PtCPF1 protein within the clock machinery, mimicking the function of animal crys. Furthermore, gene expression studies of transgenic diatom lines overexpressing PtCPF1 have indicated that the protein acts as a blue light photoreceptor because it can modulate several blue light-dependent responses. Therefore, this research project has identified a novel protein that displays both blue light photoreceptor activity as well as DNA repair activity. This protein could, in fact, be considered the missing link in the evolutionary history of the Cryptochrome/Photolyase family