DIE NEUTRALIS and LATE BLOOMER 1 Contribute to Regulation of the Pea Circadian Clock

The DIE NEUTRALIS (DNE) locus in garden pea (Pisum sativum) was previously shown to inhibit flowering under noninductive short-day conditions and to affect a graft-transmissible flowering signal. In this study, we establish that DNE has a role in diurnal and/or circadian regulation of several clock genes, including the pea GIGANTEA (GI) ortholog LATE BLOOMER 1 (LATE1) and orthologs of the Arabidopsis thaliana genes LATE ELONGATED HYPOCOTYL and TIMING OF CHLOROPHYLL A/B BINDING PROTEIN EXPRESSION 1. We also confirm that LATE1 participates in the clock and provide evidence that DNE is the ortholog of Arabidopsis EARLY FLOWERING4 (ELF4). Circadian rhythms of clock gene expression in wild-type plants under constant light were weaker in pea than in Arabidopsis, and a number of differences were also seen in the effects of both DNE/ELF4 and LATE1/GI on clock gene expression. Grafting studies suggest that DNE controls flowering at least in part through a LATE1-dependent mobile stimulus, and dne mutants show elevated expression of a FLOWERING LOCUS T homolog under short-day conditions. However, the early flowering of the dne mutant is not associated with altered expression of a previously described CONSTANS-like gene. Collectively, our results characterize the clock system and reveal its importance for photoperiod responsiveness in a model legume.     

水稻ES1参与生物钟基因表达调控以及逆境胁迫响应

生物钟参与调控植物所有的生长阶段和发育活动。维持植物生物钟稳定的基因在这一过程中起着决定性作用。在克隆了ES1 (EARLY SENESCENCE 1)基因并证明该基因影响水稻(Oryza sativa)叶片失水的基础上, 以前期分离得到的水稻突变体es1-1作为研究对象, 对es1-1及其野生型(日本晴)苗期的地上部分和地下部分进行基因芯片分析。结果表明, es1-1主要的上调基因有42个, 下调基因有14个, 这些差异基因涉及24种代谢途径, 包括调节水稻生物钟的途径(4个)、甲烷代谢途径(3个)和苯基丙氨酸代谢途径(3个)等。进一步对水稻生物钟相关基因进行表达图谱分析, 结果表明, 与野生型相比, es1-1中生物钟相关基因出现了不同程度的差异表达。对es1-1和野生型进行冷胁迫处理, 结果表明es1-1表现更加耐冷, 且冷处理后生物钟基因在日本晴(NPB)和es1-1中都表现出不同程度的差异表达。此外, 在分蘖盛期接种白叶枯菌, 发现es1-1对特定的白叶枯菌具有一定的抗性。由此推测ES1基因参与调控水稻生物钟基因的表达以及响应水稻部分逆境胁迫, 这为更深入研究水稻生物钟基因提供了新线索。     

Human Intestinal Circadian Clock: Expression of Clock Genes in Colonocytes Lining the Crypt

Biological clock components have been detected in many epithelial tissues of the digestive tract of mammals (oral mucosa, pancreas, and liver), suggesting the existence of peripheral circadian clocks that may be entrainable by food. Our aim was to investigate the expression of main peripheral clock genes in colonocytes of healthy humans and in human colon carcinoma cell lines. The presence of clock components was investigated in single intact colonic crypts isolated by chelation from the biopsies of 25 patients (free of any sign of colonic lesions) undergoing routine colonoscopy and in cell lines of human colon carcinoma (Caco2 and HT29 clone 19A). Per‐1, per‐2, and clock mRNA were detected by real‐time RT‐PCR. The three‐dimensional distributions of PER‐1, PER‐2, CLOCK, and BMAL1 proteins were recorded along colonic crypts by immunofluorescent confocal imaging. We demonstrate the presence of per‐1, per‐2, and clock mRNA in samples prepared from colonic crypts of 5 patients and in all cell lines. We also demonstrate the presence of two circadian clock proteins, PER‐1 and CLOCK, in human colonocytes on crypts isolated from 20 patients (15 patients for PER‐1 and 6 for CLOCK) and in colon carcinoma cells. Establishing the presence of clock proteins in human colonic crypts is the first step toward the study of the regulation of the intestinal circadian clock by nutrients and feeding rhythms.     

Genome Array of Hair Follicle Genes in Lambskin with Different Patterns

Hu sheep lambskin comes from a specific breed of sheep of China. Hu sheep are considered a protected breed by the Chinese government. The hair follicles of these sheep have three types of waves, large, medium, and small. There are only few histological reports of Hu sheep lambskin, and there are no modern molecular or biological studies, so the molecular mechanisms underlying the formation of hair follicles with different patterns are not currently known. The aim of this article was to study the molecular mechanism of the formation of these types of hair follicles in Hu sheep. Histological and microscopic analysis indicated that the number of follicles with small waves was not significantly higher than the number of follicles with large waves (P>0.05). The diameters of primary and secondary small-wave follicles were significantly smaller than those of large-wave follicles (P<0.05; P<0.01). The ratio between the number primary follicles and the number of secondary follicles was significantly higher among small-wave follicles than among large-wave follicles (P<0.05). Differentially expressed genes in the skin tissue were screened using an Agilent gene chip and RT-PCR. Differential expression analysis revealed 3 groups of large waves and small waves; 1067, 2071, and 3879 differentially expressed genes; and 137 genes common to all 3 groups. Differentially expressed genes were classified using gene ontology. They were found to be mainly involved in cell differentiation, proliferation, apoptosis, growth, immune response, and ion transport. RT-PCR results of 4 differentially expressed genes were consistent with gene chip results. Combined with related literature, our results suggest that BMP7, MMP2, SNAI1, SFXN1, CDKNIC, MT3, and POU1F1 may have important effects on the formation of large-wave and small-wave hair follicles. This study may enrich knowledge of hair follicle development, and may identify the genes responsible for the formation of hair follicles with different patterns.     

Sirt1与生物钟的相互调控

生物钟调控机制广泛存在于各种类型的细胞中,控制着细胞代谢的节律性变化.最近的研究发现,NAD+依赖的组蛋白去乙酰化酶Sirt1参与了生物钟调控过程,对维持正常的生物钟节律具有重要作用;另一方面,Sirt1的表达也受到生物钟系统的调控,呈现出昼夜节律性的表达.因此Sirt1能与生物钟进行相互调控,并且这一作用机制很可能广泛参与了不同类型细胞内的信号转导和能量代谢过程.本文总结了Sirt1与生物钟之间相互调控的一些研究进展,对它们之间的分子调控机制进行了概述.     

Diurnal Oscillations of Soybean Circadian Clock and Drought Responsive Genes

Rhythms produced by the endogenous circadian clock play a critical role in allowing plants to respond and adapt to the environment. While there is a well-established regulatory link between the circadian clock and responses to abiotic stress in model plants, little is known of the circadian system in crop species like soybean. This study examines how drought impacts diurnal oscillation of both drought responsive and circadian clock genes in soybean. Drought stress induced marked changes in gene expression of several circadian clock-like components, such as LCL1-, GmELF4- and PRR-like genes, which had reduced expression in stressed plants. The same conditions produced a phase advance of expression for the GmTOC1-like, GmLUX-like and GmPRR7-like genes. Similarly, the rhythmic expression pattern of the soybean drought-responsive genes DREB-, bZIP-, GOLS-, RAB18- and Remorin-like changed significantly after plant exposure to drought. In silico analysis of promoter regions of these genes revealed the presence of cis-elements associated both with stress and circadian clock regulation. Furthermore, some soybean genes with upstream ABRE elements were responsive to abscisic acid treatment. Our results indicate that some connection between the drought response and the circadian clock may exist in soybean since (i) drought stress affects gene expression of circadian clock components and (ii) several stress responsive genes display diurnal oscillation in soybeans.     

Circadian Clock Regulation of the Cell Cycle in the Zebrafish Intestine

The circadian clock controls cell proliferation in a number of healthy tissues where cell renewal and regeneration are critical for normal physiological function. The intestine is an organ that typically undergoes regular cycles of cell division, differentiation and apoptosis as part of its role in digestion and nutrient absorption. The aim of this study was to explore circadian clock regulation of cell proliferation and cell cycle gene expression in the zebrafish intestine. Here we show that the zebrafish gut contains a directly light-entrainable circadian pacemaker, which regulates the daily timing of mitosis. Furthermore, this intestinal clock controls the expression of key cell cycle regulators, such as cdc2, wee1, p21, PCNA and cdk2, but only weakly influences cyclin B1, cyclin B2 and cyclin E1 expression. Interestingly, food deprivation has little impact on circadian clock function in the gut, but dramatically reduces cell proliferation, as well as cell cycle gene expression in this tissue. Timed feeding under constant dark conditions is able to drive rhythmic expression not only of circadian clock genes, but also of several cell cycle genes, suggesting that food can entrain the clock, as well as the cell cycle in the intestine. Rather surprisingly, we found that timed feeding is critical for high amplitude rhythms in cell cycle gene expression, even when zebrafish are maintained on a light-dark cycle. Together these results suggest that the intestinal clock integrates multiple rhythmic cues, including light and food, to function optimally.     

Intramolecular Regulation of Phosphorylation Status of the Circadian Clock Protein KaiC

Background

KaiC, a central clock protein in cyanobacteria, undergoes circadian oscillations between hypophosphorylated and hyperphosphorylated forms in vivo and in vitro. Structural analyses of KaiC crystals have identified threonine and serine residues in KaiC at three residues (T426, S431, and T432) as potential sites at which KaiC is phosphorylated; mutation of any of these three sites to alanine abolishes rhythmicity, revealing an essential clock role for each residue separately and for KaiC phosphorylation in general. Mass spectrometry studies confirmed that the S431 and T432 residues are key phosphorylation sites, however, the role of the threonine residue at position 426 was not clear from the mass spectrometry measurements.

Methodology and Principal Findings

Mutational approaches and biochemical analyses of KaiC support a key role for T426 in control of the KaiC phosphorylation status in vivo and in vitro and demonstrates that alternative amino acids at residue 426 dramatically affect KaiC''s properties in vivo and in vitro, especially genetic dominance/recessive relationships, KaiC dephosphorylation, and the formation of complexes of KaiC with KaiA and KaiB. These mutations alter key circadian properties, including period, amplitude, robustness, and temperature compensation. Crystallographic analyses indicate that the T426 site is phosphorylatible under some conditions, and in vitro phosphorylation assays of KaiC demonstrate labile phosphorylation of KaiC when the primary S431 and T432 sites are blocked.

Conclusions and Significance

T426 is a crucial site that regulates KaiC phosphorylation status in vivo and in vitro and these studies underscore the importance of KaiC phosphorylation status in the essential cyanobacterial circadian functions. The regulatory roles of these phosphorylation sites–including T426–within KaiC enhance our understanding of the molecular mechanism underlying circadian rhythm generation in cyanobacteria.     

Resveratrol Regulates Circadian Clock Genes in Rat-1 Fibroblast Cells

Circadian clocks, especially peripheral clocks, can be strongly entrained by daily feedings, but few papers have reported the effects of food components on circadian rhythm. The effects of resveratrol, a natural polyphenol, on circadian clocks of Rat-1 cells were analyzed. A dose of 100 μM resveratrol, which did not show cytotoxicity, regulated the expression of clock genes Per1, Per2, and Bmal1.     

昼夜生物钟、负反馈调节与翻译后修饰

到目前为止 ,几乎在所有类型的生物中发现了昼夜生物钟。它们以约 2 4小时的周期控制着众多的分子、生理和行为过程[1、2 ] 。一个典型的例子是 ,我们的睡眠、清醒过程受昼夜钟控制 ,并受光、温对相位的重拨 ,以及地球 2 4小时昼夜的约束。不仅如此 ,我们的中心体温、某些激素分泌、生化过程均受生物钟控制。研究发现 ,对某些药物而言 ,一天中某个时候给药会得到最好的效率。乘飞机的时差反应、三班倒都是生物钟系统对环境表现的不适应 ,尽管昼夜钟的相位能即时得到调整 ,但生物钟控制的过程需要一定的时间才能得到适应。生物钟的普遍性、基…