Award details

Molecular analysis of Drosophila cryptochrome

Reference	BB/C003241/1
Principal Investigator / Supervisor	Professor Ezio Rosato
Co-Investigators / Co-Supervisors	Professor Charalambos Kyriacou
Institution	University of Leicester
Department	Biology
Funding type	Research
Value (£)	334,824
Status	Completed
Type	Research Grant
Start date	01/01/2005
End date	29/02/2008
Duration	38 months

Abstract

Circadian rhythms are universally based on transcription/translation feedback loops. In Drosophila the protein products of the period (per) and timeless (tim) genes negatively control their own transcription by inhibiting the positive factors CLOCK (CLK) and CYCLE (CYC), which bind per and tim promoters. Interlocked to this regulatory loop is a second negative feedback, which modulates the transcriptional cycling of Cik. Kinases such as DOUBLETIME (DBT), SHAGGY (SGG), CASEIN KINASE II (CKII), are also essential components of the inner clock machinery, introducing the delay necessary for the cycling of these molecular circuits. CRYPTOCROME (CRY) is a light signalling molecule, which plays a major role in the entrainment of the central pacemaker cells, and in peripheral organs, is also part of the core clock mechanism. However, to date many aspects of the biology of cryptochrome are still obscure in Drsophila; we propose to study several of the most compelling open questions with the general aims of understanding how CRY works in the fly, and how it signals the presence of light. In particular we will: 1) study the subcellular distribution and stability of CRY in different clusters of pacemaker neurons in wild type and (clock) mutant flies by confocal microscopy; 2) evaluate the genetic interactions between cry and other clock genes by behavioural analyses of double mutant flies in different environmental (light, temperature) conditions; 3) establish whether Drosophila CRY undergoes the same light-dependent phosphorylation events described for plant and mammalian CRYs with in vitro (autophosphorylation) and in vivo (other kinases) studies; 4) investigate potential functional motifs in the C-terminus of CRY by mutagenesis in vitro, generation of transgenic flies and behavioural, molecular and cellular analyses; 5) identify CRY interacting partners using coimmunoprecipitation and tandem mass spectrometry; 6) discover potential targets of light and/or CRY signalling by proteomics, using difference gel electrophoresis and mass spectrometry; 7) seek novel components involved in the process of CRY signalling to the clock by screening EMS mutagenised flies for loss of light-dependent degradation of CRY. We believe our work will contribute to elucidating CRY mode of action in light and darkness, CRY stability, activation and signalling and CRY dependent/independent modification of light targets.

Summary

unavailable

Committee	Closed Committee - Animal Sciences (AS)
Research Topics	X – not assigned to a current Research Topic
Research Priority	X – Research Priority information not available
Research Initiative	X - not in an Initiative
Funding Scheme	X – not Funded via a specific Funding Scheme

I accept the terms and conditions of use (opens in new window)

export PDF file

back to list new search