CLOCK GENES IN

DROSOPHILA

 

Drosophila is probably the most extensively studied organism in circadian clock research. Clock research in Drosophila was started by the cloning of the clock gene period (dper) after screening for Drosophila mutants with altered period lengths. This was followed by the cloning of the gene timeless (dtim). 

Transcription of both per and tim starts low at the star of the day but starts to increase at about midday. The peaks levels of mRNA occur approximately 4 hours after dusk (Green 1998). Like in Neurospora there is a delay between the peak mRNA levels and the peak levels of the proteins dPER and dTIM due to post-transcriptional delays. These proteins accumulate in the cytoplasm where they form heterodimers by interacting at their PAS domains. dPER and dTIM appear to have no other functional domains (Harmer et al 2001). 

The Drosophila clock also contains 2 other genes called clock (dclk) and cycle (cyc). Both dclk and cyc contain basic helix-loop-helix domains (bHLH), which are DNA binding domains, and PAS domains. dclk is transcribed in a cyclic manner, (Bae et al. 1998) with the peak levels of dclk mRNA occurring at around dawn (Bae et al 2000). cyc on the other hand does not cycle (contrary to its name), either as mRNA or as a protein (Bae et al 2000). In the absence of cyc and dclk (or in mutants of these genes), the levels of dper and dtim mRNA, and dPER and dTIM protein are reduced and do not cycle. (Harmer et al 2001). This is because dclk and cyc are transcription factors that regulate transcription of dper and dtim (this is indicated by the presence of the bHLH domains in cyc and dclk). It is thought that dCLK and CYC act by forming heterodimers (using the PAS domains as with WC-1 and WC-2 in Neurospora). These then bind to E-box elements in the promoters of dper and dtim. This increases the expression of these genes (as can be seen by increased expression of luciferase reporter genes that are tied to these promoters (Hogenesch 1998)). CYC and dCLK therefore act as positive elements in the feedback loop. The negative regulation of the cycle comes from dPER and dTIM, which bind to CYC and dCLK. This prevents the two proteins from binding to the DNA and up regulating the transcription of dper and dtim (Harmer et al. 2001). 

A second feedback loop also exists in Drosophila. The presence of dCLK and CYC repress the transcription of dclk. The mechanism for this is unknown.

The Drosophila clock. Proteins with positive actions are in yellow, negative components are in blue. Protein-protein interactions are indicated with dotted black arrows. Green arrows indicate a positive effect on some process or promoter, and red lines indicate an inhibitory action.  The question mark indicates that CYC and dCLK have not been shown to bind the promoters of output genes.

 (From Harmer 2001)

 

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