Research
 
 
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Universität Bielefeld > Fakultät für Biologie > Chair of Molecular Cell Physiology > Research
  

RNA-binding proteins and post-transcriptional control in the circadian system

Circadian rhythms

Organisms have evolved endogenous circadian clocks with a period of about 24 h that allow synchronization with the periodic changes in light and temperature in the outside world. Plants with a functional circadian clock have an enhanced performance and fitness because they correctly phase their daily life to the light/dark cycle in the environment.
Underlying these endogenous circadian rhythms like photosynthetic activity, growth or hormonal levels, is a transcriptional network under control of the circadian clock: Almost 90% of transcripts in Arabidopsis accumulate at defined times of the day.

The circadian transcriptome

The core clockwork comprises transcriptional feedback loops with positively and negatively acting proteins that regulate expression of their own genes and thus generate a self-sustained 24 h oscillation.
Our research focuses on RNA-binding proteins and posttranscriptional regulation in the circadian system.

In situ hybridisation of the glycine-rich RNA-binding protein SaGRP in Sinapis alba stem (Heintzen et al. 1994).

 

The RNA-binding protein AtGRP7

The RNA-binding protein AtGRP7 (Arabidopsis thaliana glycine-rich RNA-binding protein 7) is part of a molecular slave oscillator downstream of the Arabidopsis core clockwork. AtGRP7 negatively autoregulates oscillations of its own mRNA by alternative splicing coupled to Nonsense-mediated decay. AtGRP8 encoding a related RRM-protein is not only an AtGRP7 target but in turn also negatively autoregulates and reciprocally cross-regulates AtGRP7. Thus, we identify an interlocked feedback loop through which two RNA-binding proteins autoregulate and reciprocally crossregulate by coupling unproductive splicing to NMD. It appears that the two RNA-binding proteins fine-tune their expression through this cross-regulation and effectively buffer against disturbances.

Negative autoregulation and reciprocal regulation of AtGRP7 and AtGRP8 via AS-NMD.

 

Transcript profiling of transgenic plants ectopically overexpressing AtGRP7 compared to wt plants has identified candidate AtGRP7 target transcripts.
In collaboration with the Department of Laser Physics we have used Fluorescence Correlation Spectroscopy to study the interaction of AtGRP7 with its RNA target and conformational dynamics of the RNA substrate. Photoelectron transfer (PET) based fluorescence was used to assay conformational dynamics of the RNA binding substrates.

AtGRP7 preferentially binds to its 3’ UTR in an extended conformation.

 

In collaboration with the Department of Biophysics and Nanotechnology we have adapted Atomic Force Microscopy to study RNA-protein interaction at the single molecule level.

Identification of different binding modes

 

AtGRP7 promotes the transition to flowering in Arabidopsis.

Contrary to the expectation for a clock-controlled protein, AtGRP7 presumably is not involved in mediating the photoperiodic response to inductive long days. Rather, it appears to be part of the autonomous pathway, pointing to extensive crosstalk between the circadian timing system and the floral network.

AtGRP7-ox plants flower early and have reduced levels of the floral repressor FLC.