Processing body and Stress granule rhythms in cell cultures

MALCOLM, MELISA; PENAZZI, LAURA GABRIELA; GARBARINO-PICO, EDUARDO

Congreso; XVI Latin American Symposium on Chronobiology; 2021

PROCESSING BODY AND STRESS GRANULE RHYTHMS IN CELL CULTURESMalcolm M, Pennazi LG, Garbarino-Pico E.1,21Universidad Nacional de Córdoba, Facultad de Ciencias Químicas, Departamento de Química Biológica Ranwel Caputto, Córdoba, Argentina.2CONICET, Universidad Nacional de Córdoba, Centro de Investigaciones en Química Biológica de Córdoba (CIQUIBIC), Córdoba, Argentina.Stress granules (SGs) and processing bodies (PBs) are cytoplasmic membraneless organelles that are formed by liquid-liquid phase transitions. They are involved in the regulation of translation, stability and storage of mRNA and contain mRNA and several RNA binding proteins. SGs form in response to different stress stimuli, typically through phosphorylation of the eIF2alpha. PBs are constitutively present but increase in number under stress conditions. Since stress response has been shown to be circadianly regulated, we wonder whether SGs and PBs oscillate. NIH/3T3 and N2a cell cultures were synchronized with dexamethasone and harvested every 4 h for 68 h. We induced the formation of SGs with sodium arsenite (oxidative stress). We performed a double immunolabeling of SGs (eIF3 and G3BP1) and PB (GE-1/HEDLS and DDX6) by immunocytochemistry. We studied the phosphorylation temporal profile of eIF2alpha and eIF3 levels and we found no differences over time. We observed that NIH/3T3 and N2A cells show daily rhythms in SGs and PBs, respectively, for three variables: number, area, and signal intensity, with periods of approximately 24 h. These findings suggest that the molecular circadian clock controls SGs and PBs. To determine this hypothesis, we analyzed their formation in Bmal1-/- fibroblasts. Surprisingly, the rhythm persisted in these cells. The results presented here reveal new ways in which translation and cytoplasmic mRNA metabolism can be modulated over time.