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Extracellular signal-regulated kinase-2 phosphorylates RORalpha4 in vitro
Adriane Lechtken1, Michael Hörnig, Oliver Werz
1Institute of Pharmaceutical Chemistry/ZAFES, Johann Wolfgang Goethe University Frankfurt, Max-von-Laue-Strasse 9, 60438 Frankfurt am Main, Germany.
Biochemical and Biophysical Research Communications
|May 22, 2007
Summary
Extracellular signal-regulated kinase 2 (ERK-2) phosphorylates RORalpha4, enhancing its DNA binding and transcriptional activity. This phosphorylation impacts circadian rhythm and gene regulation.
Area of Science:
- Molecular Biology
- Gene Regulation
- Chronobiology
Background:
- Retinoic acid related orphan receptor alpha (RORalpha) is crucial for cerebellar development, neuroprotection, inflammation control, and circadian rhythm.
- RORalpha plays a pivotal role in the mammalian core-clock mechanism.
Purpose of the Study:
- To identify the kinase responsible for RORalpha4 phosphorylation.
- To investigate the functional consequences of RORalpha4 phosphorylation by ERK-2.
Main Methods:
- In vitro kinase assays to identify RORalpha4 phosphorylating kinase.
- Site-directed mutagenesis to create RORalpha4-T128A mutant.
- Analysis of DNA-binding affinity and transcriptional activity.
Main Results:
- Extracellular signal-regulated kinase 2 (ERK-2) was identified as a RORalpha4 phosphorylating kinase in vitro.
- Mutation of Thr-128 to Alanine (RORalpha4-T128A) prevented ERK-2 mediated phosphorylation.
- The RORalpha4-T128A mutant showed increased DNA-binding affinity and transcriptional activity compared to wild-type RORalpha4 (RORalpha4-WT).
- RORalpha4-T128A acted as a stronger competitor against RevErbalpha at ROR response elements.
Conclusions:
- ERK-2 mediated phosphorylation of RORalpha4 at Thr-128 modulates its activity.
- Phosphorylation by ERK-2 influences RORalpha4's role in gene regulation and circadian rhythm control.
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