on 09-17-201202:41 AM - edited on 10-15-202111:00 AM by AnalyteGuru
Cargnello M, Tcherkezian J, Dorn JF, Huttlin EL, Maddox PS, Gygi SP, Roux PP Mol Cell Biol. 2012 Nov;32(22):4572-84. Processing Bodies (P-bodies or PBs) are cytoplasmic granules involved in mRNA storage and degradation that participate in the regulation of gene expression. PBs concentrate non-translated mRNAs and several factors involved in mRNA decay and translational repression, including the eIF4E-transporter (4E-T). 4E-T is required for PB assembly, but little is known about the molecular mechanisms that regulate its function. Here, we demonstrate that oxidative stress promotes multisite 4E-T phosphorylation. We show that the c-Jun N-terminal kinase (JNK) is targeted to PBs in response to oxidative stress and promotes the phosphorylation of 4E-T. Quantitative mass spectrometry analysis reveals that JNK phosphorylates 4E-T on six proline-directed sites that are required for the formation of 4E-T complex upon stress. We have developed an image-based computational method to quantify the size, number and density of PBs in cells and we find that while 4E-T is required for steady state PB assembly, its phosphorylation facilitates the formation of larger PBs upon oxidative stress. Using polysomal mRNA profiling we assessed global and specific mRNA translation, but did not find that 4E-T phosphorylation impacts upon translational control. Collectively, these data support a model whereby PB assembly is regulated by a two-step mechanism involving a 4E-T-dependent assembly stage in unstressed cells and a 4E-T phosphorylation-dependent aggregation stage in response to stress stimuli.