The Circadian Clocks Central Mechanism - Assignment Example

?Circadian Clocks A circadian clock also known as a circadian oscillator is a biochemical system in living organisms that oscillates within a period of 24 hours. It is normally coordinated with the day- night cycle. The circadian clocks are main the central mechanisms which drive circadian rhythms. Circadian rhythms are physical, mental and behavioral changes that follow an approximate 24-hour cycle, responding mainly to light and darkness in an organism’s environment. They are found in most living things, including animals, plants and many bacteria (Lowrey PL) The most compelling demonstration of the circadian clock’s utility has been made by using cyanobacteria strains with different clock properties growing in competition with each other. In cyanobacteria, strains with a clock period length that matched the environmental photoperiod outgrow strains with no clock or with an out of phase period length in competitive culture conditions (Woefle et al., 2004). This does gesture to the probability of the clock being an important evolutionary step that allowed survival of early organisms. Circadian rhythms can influence sleep-wake cycles, hormone release, body temperature and other important intrinsic functions. Organization and function Function: The main principle of circadian clocks is oscillatory gene activation. The initial gene activation is regulated by the last one in the sequence, making up an auto-regulatory feedback loop that lasts about 24 hours. That is, feedback loops of transcription and translation, whereby the protein product of a clock gene will indirectly shut off its own expression. Such feedback loops are controlled and probably lengthened by post-translational modifications of most proteins involved (Harms et al., 2004) Organization: They have three major components: 1) A central oscillating mechanism with a period of about 24 hours. 2) Several output pathways associated with distinct phases of the oscillator. These control the activities of the organism. 3) Several input pathways to this central oscillator to allow programming of the circadian clock. Metazoans possess circadian clocks in most cells of the body. Each of these clocks is autonomous (Welsh et al., 1995). This means that each of the cells has to be entrained autonomously as well. But, since the oscillations of the clocks are roughly 24 hours in length, there has to be a form of synchronization. In the case of direct light entrained organisms, such as the drosophila, this entrainment occurs in cells independently (Plautz et al.,1997) (Whitmore et al., 2000). In drosophila, the Drosophila ring gland and Malpighian tubules show some sort of rhythms. This autonomous cell entrainment may also be seen in higher organisms where the parasympathetic system is responsible for entrainment (Ishida et al., 2005). It is itself entrained by the central master clock. In higher organisms, the process is more complex and requires what is known as hierarchical entrainment. This is discussed below. In higher organisms such as man, the entrainment of the clock is hierarchical. This is similar to most other vertebrates. In mammals, the suprachiasmic nucleus acts as the main/master clock. It is a paired neuronal structure located at the base of the hypothalamus. It is just above the optic chiasm/bifurcation (Klein et al.1991). It facilitates entrainment and synchrony of all other tissues by direct and indirect methods. These include use of hormones, temperature regulation, feeding regulation and metabolism. The suprachiasmic nucleus undergoes entrainment by relation with the optic apparatus via optic tract. This is the retionohypothalamic tract (Ben-Schlomo and Kyriacou, 2002). The SCN consists of a mixed population of neuronal and glial cells, but which types of cells might be capable of acting as circadian oscillators is presently unknown . Since they are autonomous in function, several theories have been proposed as to their synchronization. One is the synchrony of circadian rhythm due to production of melatonin by the pineal gland. It should normally feed back onto SCN cells to keep them synchronized. Another theory is the production of NO that acts as a synchronizing factor (McArthur et al.1991) The first figure is simple entrainment, while the second is hierarchical entrainment. (Image borrowed from University of Zurich chrono-biology of sleep.) Below are diagrams of the timing loops of man, D.melanogaster , N.Crassa and plants In Man The CLOCK proteins (yellow) and BMAL1 proteins (purple) undergo heterodimerisation and drive the expression of the Per, Cry and Rev-erb alpha (dark green) genes within the nucleus. The PER proteins in red, and CRY proteins (shown as cyan) interact in the nucleus to inhibit CLOCK/BMAL1 action. This is an unknown mechanism. This leads to their down regulation. When the REV-ERB alpha protein is absent, BMAL1 is expressed and thus transcribed to produce more CLOCK/BMAL1 transcription factors that reinitiate a new circadian cycle. CLOCK proteins undergo post translational modifications. For example, the (CKI) phosphorylates PER. Increased phosphorylation of PER promotes its degradation (Albrecht) 12 In figure one, Clk–Cyc heterodimers bind to E-boxes and initiate transcription of Per and Tim. As Per is produced it is phosphorylated by the Drosophila feedback loop 1 by Dbt and CK2. Tim binds to, and stabilizes, phosphorylated Per, which remains bound to Dbt. Furthermore, Per is also stabilized by PP2a, which removes phosphation products that were added to Per. The Tim–Per–Dbt complexes are phosphorylated by the Sgg which, in conjunction with phosphorylation by CK2, facilitates their transport into the nucleus. Tim–Per–Dbt complexes then bind to Clk–Cyc, thereby removing Clk–Cyc from the E-box and lead to inhibiting Per and Tim transcription. Per and Clk are thus destabilized, via Dbt phosphorylation, and degraded, whereas Tim degradation is triggered by tyrosine phosphorylation. The accumulation of non-phosphorylated Clk leads to heterodimerization with Cyc and another cycle of Per and Tim transcription. In figure 2 The Clk–Cyc heterodimers bind to the E-boxes as in the fig 1, but instead activate Vri and Pdp1epsilon transcription. Vri then accumulates in parallel with its forming mRNA, binds to V/P boxes and inhibits Clk transcription. PDP1 epsilon accumulates slowly and displaces Vri from V/P box to derepress Clk transcription. A cicardian clock independent activator (Act) constitutively activates Clk transcription in the absence of Vri, which would therefore explain the high levels of Clk mRNA in the absence of Clk or Cyc. Accumulation of non-phosphorylated Clk leads to heterodimerization with the Cyc and another cycle of Vri and Pdp1epsilon transcription. (Hardin P, 2005) In the fungi, Neurosporra crassa In the fungi, the effects of light were measured by periodicity in the conidiation of the fungi both in space and on earth. This was done to eliminate the other periodic factors that may have affected the results. The experiment showed that the N.crasssa had a very adaptable circadian system. Studies on the N.crassa have shown that the core circadian oscillator of Neurospora consists of a negative feedback loop in which FRQ, FRH, WHITE COLLAR 1 (WC-1), and WC-2 are the core components. (The FRH is a helicase) (Heintzen) WC-1 and WC-2 form a heterodimer (D-WCC) which binds to the Clock box in the frq promoter. It leads to frq transcription (Cheng). FRQ protein dimerises with itself and forms a complex with FRH (FFC). In the nucleus, FFC then inhibits D-WCC activity, resulting in a decrease in frq mRNA levels. When FRQ levels drop below a certain level, D-WCC is no longer inhibited by FFC, and thus the frq transcription is reactivated to start a new cycle. Double functions of FRQ proteins allow them to participate in multiple clock-related feedback loops in the Neurospora clock (green). The WC-1 and WC-2 proteins form a WCC compound that activates gene expression in the dark from frq and also from clock-controlled genes known as ccgs. These are associated with output and mediate light-induced transcription from frq, ccgs, and wc-1. These are shown in golden arrows. The N.crassa RNA is translated to make FRQ proteins which have two roles: (i) FRQ feeds back into the nucleus to quickly block the positive effect of the WCC in driving the frq transcription. (ii) The FRQ acts to facilitate the synthesis of more WC-1. Phosphorylation of FRQ complex triggers its turnover. This is a major determinant of period length in the circadian clock. (Lee K) In plants In plants, the circadian rhythm regulation is obtained by light. The Arabidopsis is the one most used for the tests and experiments. Plant circadian rhythms control when the flower flowers or bears fruits. Examples of circadian controlled rhythms are the germination, growth, leaf movement, flower opening and stomata cycles. Features of how these rhythms are maintained include two interacting transcription-translation feedback loops. Some proteins containing PAS domains, (which facilitate protein-protein interactions) and several photoreceptors that tune the clock to different light conditions are required for the feedback loops to function. Phytochromes and cryptochromes as well are required for the entrapment of light. The phytochromes absorb blue and red light. There are three main phytochromes (ABE). The phyA is found in seedlings grown in dark, and the phy B in seedlings grown in light. There is cryptochrome 1 and 2 that bind blue and UV-A. In plants, the central oscillator generates an autonomous rhythm and is driven by two interacting feedback loops that are active at different times of day. The morning loop has CCA1 gene (Circadian and Clock Associated 1) and LHY gene (Late Elongated Hypocotyl), which encode closely related MYB transcription factors. These regulate circadian rhythms in the aforementioned Arabidopsis, as well as PRR 7 and 9 (Pseudo-Response Regulators). The evening loop consists of the GI (Gigantea) and ELF4, that are both both involved in the regulation of flowering time genes (Kolmos). When the CCA1 and LHY are overexpressed, plants lose rhythm, and mRNA signals reduce, contributing to a negative feedback loop. The CCA1 and LHY repress TOC1 and over-expressed TOC1 is a positive regulator of CCA1 and LHY. The TOC1 is a gene that acts as a repressor of cca1, lhy, and prr7 and 9 during the morning, but also of gi and elf4 (McClung). TOC1 gene expression peaks in the early evening. 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