Period (PER) proteins are essential components of the mammalian circadian clock. They form complexes with cryptochromes (CRY), which negatively regulate CLOCK/BMAL1-dependent transactivation of clock and clock-controlled genes. To define the roles of mammalian CRY/PER complexes in the circadian clock, we have determined the crystal structure of a complex comprising the photolyase homology region of mouse CRY1 (mCRY1) and a C-terminal mouse PER2 (mPER2) fragment. mPER2 winds around the helical mCRY1 domain covering the binding sites of FBXL3 and CLOCK/BMAL1, but not the FAD binding pocket. Our structure revealed an unexpected zinc ion in one interface, which stabilizes mCRY1-mPER2 interactions in vivo. We provide evidence that mCRY1/mPER2 complex formation is modulated by an interplay of zinc binding and mCRY1 disulfide bond formation, which may be influenced by the redox state of the cell. Our studies may allow for the development of circadian and metabolic modulators.

The majority of transplants are derived from donors who suffered from brain injury. There is evidence that brain death causes inflammatory changes in the donor. To define the impact of brain death, we evaluated the gene expression of cytokines in human brain dead and ideal living donors and compared these data to organ function following transplantation.Hepatic tissues from brain dead (n = 32) and living donors (n = 26) were collected at the time of donor laparotomy. Additional biopsies were performed before organ preservation, at the time of transplantation and one hour after reperfusion. Cytokines were assessed by real-time reverse transcriptase-polymerase chain reaction (RT-PCR) and cytometric bead array. Additionally, immunohistological analysis of tissue specimens was performed. Inflammatory cytokines including IL-6, IL-10, TNF-a , TGF-b and MIP-1a were significantly higher in brain dead donors immediately after laparotomy compared to living donors. Cellular infiltrates significantly increased in parallel to the soluble cytokines IL-6 and IL-10. Enhanced immune activation in brain dead donors was reflected by a deteriorated I/R injury proven by elevated alanin-aminotransferase (ALT), aspartat-amino-transferase (AST) and bilirubin levels, increased rates of acute rejection and primary nonfunction. Based on our clinical data, we demonstrate that brain death and the events that †Authors contributed equally to this manuscript.precede it are associated with a significant upregulation of inflammatory cytokines and lead to a worse ischemia/reperfusion injury after transplantation.

Circadian clocks govern a wide range of cellular and physiological functions in various organisms. Recent evidence suggests distinct functions of local clocks in peripheral mammalian tissues such as immune responses and cell cycle control. However, studying circadian action in peripheral tissues has been limited so far to mouse models, leaving the implication for human systems widely elusive. In particular, circadian rhythms in human skin, which is naturally exposed to strong daytime-dependent changes in the environment, have not been investigated to date on a molecular level. Here, we present a comprehensive analysis of circadian gene expression in human epidermis. Whole-genome microarray analysis of suction-blister epidermis obtained throughout the day revealed a functional circadian clock in epidermal keratinocytes with hundreds of transcripts regulated in a daytime-dependent manner. Among those, we identified a circadian transcription factor, Krüp-pel-like factor 9 (Klf9), that is substantially up-regulated in a cortisol and differentiation-state-dependent manner. Gain-and loss-offunction experiments showed strong antiproliferative effects of Klf9. Putative Klf9 target genes include proliferation/differentiation markers that also show circadian expression in vivo, suggesting that Klf9 affects keratinocyte proliferation/differentiation by controlling the expression of target genes in a daytime-dependent manner.glucocorticoids | skin cancer B iological rhythms regulate cellular and physiological processes ranging from milliseconds to years. A well-studied timing system is the circadian (∼24-h) clockwork that allows organisms to anticipate diurnal variations in environmental conditions such as light, food availability, oxidative stress, pathogen exposure, or temperature. In mammals, the central circadian pacemaker resides in the suprachiasmatic nucleus (SCN), located in the anterior hypothalamus. Oscillations of SCN neurons are cell-autonomous, self-sustained, and synchronized to external time cues (Zeitgebers) such as light (1). The SCN, in turn, synchronizes peripheral clocks by systemic time cues such as neuronal input, hormonal signaling (e.g., cortisol), body temperature, and possibly many others (2). Interestingly, most cells in peripheral tissues also possess cell-autonomous clockworks with a similar molecular makeup to SCN neurons. These peripheral clocks are thought to generate or amplify daytime-dependent physiological and metabolic functions in a tissue-specific manner by circadian regulation of clock-controlled genes (3, 4).On a molecular level, circadian oscillation is generated by interlocked transcriptional-translational feedback loops. The transcription factor dimer CLOCK/BMAL1 drives expression of target genes such as Periods (Per1-3) and Cryptochromes (Cry1-2) by binding to E-box elements in their promoters. The negative feedback is formed by PER/CRY protein complexes that shuttle back into the nucleus, where they block CLOCK/BMAL1-mediated transactivation, thereby inhibiting their own transcr...

Heme oxygenase-1 is a regulator of vascular function in hypertension via determining the phenotype of inflammatory circulating and infiltrating monocytes with possible implications for all-cause mortality.

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