Functional analysis of the rodent tau mutation in a marine unicellular alga

ABSTRACT: Casein Kinase 1 (CK1) is one of few proteins known to affect cellular timekeeping across metazoans, and the naturally occurring CK1tau mutation shortens circadian period in mammals. Functional conservation of a timekeeping function for CK1 in the green lineage was recently identified in the green marine unicell Ostreococcus tauri, in spite of the absence of CK1's transcriptional targets known from other species. The short-period phenotype of CK1tau mutant in mammals depends specifically on increased CK1 activity against PERIOD proteins. To understand how CK1 acts differently upon the algal clock, we analysed the effect of CK1tau overexpression on the phospho-preoteome of O. tauri using label-free quantification compared to the parent line. Ostreococcus tauri cells were harvested, trypsinised and phosphopeptide enrichment was performed using TiO2 columns followed by LC-MS separation (140minute gradient) on a Orbitrap XL. All those steps are described previously (Le Bihan et al J Proteomics. 2011 Sep 6;74(10):2060-70). MSMS data was searched using MASCOT Version 2.4 (Matrix Science Ltd, UK) against the Ostreococcus tauri subset of the NCBI protein database (12/01/2011; 8,726 sequences) using a maximum missed-cut value of 2., variable oxidation (M), N-terminal protein acetylation, phosphorylation (STY) and fixed carbamidomethylation (C); precursor mass tolerance was 7 ppm and MSMS tolerance 0.4 amu. The significance threshold (p) was set below 0.05 (MudPIT scoring).

INSTRUMENT(S): orbitrap

ORGANISM(S): Ostreococcus Tauri

TISSUE(S): Cell Culture

SUBMITTER: Thierry Le Bihan  

LAB HEAD: Thierry Le Bihan

PROVIDER: PXD000378 | Pride | 2013-10-18

REPOSITORIES: Pride