Cryptochromes

Hippocampal plasticity and mnemonic processing exhibit a striking time-of-day dependence and likely implicate a temporally structured replay of memory traces. Molecular mechanisms fulfilling the requirements of sensing time and capturing time-related information are coded in dynamics of so-called clock genes and their protein products, first discovered and described in the hypothalamic suprachiasmatic nucleus. Using real-time PCR and immunohistochemical analyses, we show that in wildtype mice core clock components (mPer1/PER1, mPer2/PER2, mCry1/CRY1, mCry2/CRY2, mClock/CLOCK, mBmal1/BMAL1) are expressed in neurons of all subregions of the hippocampus in a time-locked fashion over a 24-h (diurnal) day/night cycle. Temporal profiling of these transcriptional regulators reveals distinct and parallel peaks, at times when memory traces are usually formed and/or consolidated. The coordinated rhythmic expression of hippocampal clock gene expression is greatly disordered in mice deficient for the clock gene mPer1, a key player implicated in both, maintenance and adaptative plasticity of circadian clocks. Moreover, Per1-knockout animals are severely handicapped in a hippocampus-dependent long-term spatial learning paradigm. We propose that the dynamics of hippocampal clock gene expression imprint a temporal structure on memory processing and shape at the same time the efficacy of behavioral learning. © 2009 Wiley-Liss, Inc.

Glucocorticoid (GC) plays important roles in diverse physiological processes including metabolism and immune functions. While circadian control of GC synthesis and secretion is relatively well appreciated, circadian control of GC action within target tissues remains poorly understood. Here, we demonstrate that CLOCK/BMAL1, the core circadian clock components, reduces maximal GR transactivation (Amax) as well as efficacy (EC50) by a novel mechanism that requires binding to DNA and transactivation of target genes. Accordingly, we observe that PER1 and CRY1, the primary targets of CLOCK/BMAL1 action, reduce maximal GR transactivation while not affecting the efficacy. Moreover, we observe hyper-activations of GRE-dependent transcription in BMAL1- or PERs-deficient MEFs. In addition, endogenous GC target genes expression negatively correlates with the CLOCK/BMAL1 activity. Considering that GC sensitivity is widely implicated in human health and diseases, these results provide valuable in...

Light is one of the most important environmental factors regulating seed germination. It is known that light inhibits seed germination of some monocotyledonous species and that it is mostly related to the blue wavelength of the spectrum received by cryptochromes (cry). Research has also found that the red light (R) stimulates germination of dicotyledonous seeds and that this reaction involves mainly phytochromes (phy). Surprisingly, up to date, the role and the mechanism of action of blue light (BL) in seed biology of dicot plants is still very poorly understood and some questions are unexplained, e.g., whether BL plays a role in regulation of dicot seeds dormancy and/or germination? If, so what particular elements of light signaling pathway are involved in modulation of this(ese) process(es)? Also, is the BL action in regulation of dicot seeds dormancy and/or germination maybe due to changes of expression of genes related to metabolism and/or signaling of two phytohormones controlling seed-related events, such as gibberellins (GA) and abscisic acid (ABA)? To answer these intriguing questions, the combination of biological, transcriptomic, and genetic approaches was performed in this particular study. The germination tests show that freshly harvested wild type (WT) Arabidopsis thaliana Col-0 seeds are dormant and do not germinate in darkness (at 25 • C), while nondormant (after-ripened) seeds germinate well in these conditions. It is also proven that dormancy of seeds of this species is released in the presence of white and/or BL (λ = 447 nm) when placed at 25 • C. Presented here, novel results emphasize the role of BL in dormancy alleviation of dicot seeds, indicating that this wavelength of light spectrum received by phyB induces this process and that the sensitivity to this stimulus depends on the depth of seed dormancy. In addition, it is demonstrated that various elements of phy-mediated pathway can be used in response to the signal induced by BL in germinating dormant seeds of Arabidopsis. The quantitative real time PCR analysis supported by results of germination tests of WT, T-DNA insertion mutants (i.e., hy5, hfr1, and laf1) and overexpression transformants of Arabidopsis seeds (i.e., 35S:OE:HY5, 35S:OE:HYH, 35S:OE:HFR1, and 35S:OE:LAF1) revealed that the HY5 gene coding transcription factor is most probably responsible for the control of expression of genes involved in GA/ABA metabolism and/or signaling pathways during BL-dependent dormancy alleviation of Arabidopsis seeds, while biological functions of HYH and HFR1 are associated with regulation of germination. The model of BL action in regulation of dormancy alleviation and germination potential of Arabidopsis seeds is proposed.

Cryptochromes (Crys) are blue light receptors believed to have evolved from the DNA photolyase protein family, implying that light control and light protection share a common ancient origin. In this paper, we report the identification of five genes of the Cry/photolyase family (CPF) in two green algae of the Ostreococcus genus. Phylogenetic analyses were used to confidently assign three of these sequences to cyclobutane pyrimidine dimer (CPD) photolyases, one of them to a DASH-type Cry, and a third CPF gene has high homology with the recently described diatom CPF1 that displays a bifunctional activity. Both purified OtCPF1 and OtCPF2 proteins show non-covalent binding to flavin adenine dinucleotide (FAD), and additionally to 5,10-methenyl-tetrahydrofolate (MTHF) for OtCPF2. Expression analyses revealed that all five CPF members of Ostreococcus tauri are regulated by light. Furthermore, we show that OtCPF1 and OtCPF2 display photolyase activity and that OtCPF1 is able to interact with the CLOCK:BMAL heterodimer, transcription factors regulating circadian clock function in other organisms. Finally, we provide evidence for the involvement of OtCPF1 in the maintenance of the Ostreococcus circadian clock. This work improves our understanding of the evolutionary transition between photolyases and Crys.

The signaling pathways that couple light photoreception to entrainment of the circadian clock have yet to be deciphered. Two prominent groups of candidates for the circadian photoreceptors are opsins (e.g., melanopsin) and blue-light photoreceptors (e.g., cryptochromes). We have previously showed that the zebrafish is an ideal model organism in which to study circadian regulation and light response in peripheral tissues. Here, we used the light-responsive zebrafish cell line Z3 to dissect the response of the clock gene zPer2 to light. We show that the MAPK (mitogen-activated protein kinase) pathway is essential for this response, although other signaling pathways may also play a role. Moreover, action spectrum analyses of zPer2 transcriptional response to monochromatic light demonstrate the involvement of a blue-light photoreceptor. The Cry1b and Cry3 cryptochromes constitute attractive candidates as photoreceptors in this setting. Our results establish a link between blue-light photoreceptors, probably cryptochromes, and the MAPK pathway to elicit light-induced transcriptional activation of clock genes.

The Arabidopsis gene encoding the key flavonoid biosynthesis enzyme chalcone synthase (CHS) is regulated by several environmental and endogenous stimuli. Here we dissect the network of light signalling pathways that control CHS expression in mature leaves using cryptochrome (cry) and phytochrome (phy) deficient mutants. The UV-A/blue light induction of CHS is mediated principally by cry1, but neither cry1 nor cry2 is involved in UV-B induction or in the UV-A and blue light signalling pathways that interact synergistically with the UV-B pathway to enhance CHS expression. Moreover, these synergistic responses do not require phyA or phyB. Phytochrome is a positive regulator of the cry1 inductive pathway, mediating distinct potentiation and coaction effects. A red light pretreatment enhances subsequent cry1-mediated CHS induction. This potentiation is unaltered in phyA and phyB mutants but much reduced in a phyA phyB double mutant, indicating that it requires principally phyA or phyB. In contrast, the cry1-mediated induction of CHS, without pretreatment, is much reduced in phyB but not phyA mutants, indicating coaction between cry1 and phyB. Further experiments with phy-deficient mutants demonstrate that phyB is a negative regulator of the UV-B inductive pathway. We further show that phyB acts upstream of the points of interaction of the UV-A and blue synergism pathways with the UV-B pathway. We propose that phyB functions to balance flux through the cry1 and UV-B signalling pathways.