GARBARINO PICO EDUARDO
Congresos y reuniones científicas
Título:
Stress granules and processing bodies oscillate in a circadian manner.
Autor/es:
MALCOLM M; PENAZZI LG; GARBARINO PICO E
Reunión:
Congreso; LVI SAIB - XV SAMIGE ? Online Joint Meeting 2020; 2020
Resumen:
Stress granules (SG) and processing bodies (PB) are membrane-less organelles, responsible for storage and cytoplasmic
processing of mRNA, which share some components such as mRNA and RNA binding proteins. SGs, which are also
constituted by translation initiation factors, the minor subunit of the ribosome 40S, and translation stalled mRNAs, are formed
in response to different stress stimuli, typically through phosphorylation of the eIF2 factor. PBs, on the other hand, are enriched
in factors involved in mRNA degradation, translational repression, and RNA-mediated silencing, and they are constitutive
membrane-less organelles, although they increase in number with stress. In previous studies we found that SG formation
presents temporary changes in number and size in mouse fibroblast synchronized cultures (NIH/3T3). We wonder whether the
circadian clock controls these temporal changes. The cells were synchronized with dexamethasone and harvested every 4 h
for 68 h. We induced the formation of SGs with sodium arsenite (oxidative stress). On the other hand, we analyzed PBs in not
stressed N2A cells (neuroblastome cell line). We also studied the temporal expression profile of p-eIF2α (factor involved in
SG aggregation) and eIF3, the marker we used for detecting SGs, by Western blot. Cell cultures were arrested in order to
prevent cell cycle progression, and we confirm their quiescence state by flow cytometry. We performed a double
immunolabeling of SGs (eIF3 and G3BP1) and PB (GE-1/HEDLS and DDX6) by immunocytochemistry. 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 the estimated parameters showing periods of approximately 24 h, for two cycles. The subtle differences that we
found between both translation initiation factors eIF3 and eIF2α, would not be responsible for generating the changes observed
in SGs. These findings strongly suggest that the molecular circadian clock rhythmically controls SGs and PBs. The results
presented here reveal new ways in which circadian clocks modulate mRNA translation, stability, and storage in the cytoplasm
processing of mRNA, which share some components such as mRNA and RNA binding proteins. SGs, which are also
constituted by translation initiation factors, the minor subunit of the ribosome 40S, and translation stalled mRNAs, are formed
in response to different stress stimuli, typically through phosphorylation of the eIF2 factor. PBs, on the other hand, are enriched
in factors involved in mRNA degradation, translational repression, and RNA-mediated silencing, and they are constitutive
membrane-less organelles, although they increase in number with stress. In previous studies we found that SG formation
presents temporary changes in number and size in mouse fibroblast synchronized cultures (NIH/3T3). We wonder whether the
circadian clock controls these temporal changes. The cells were synchronized with dexamethasone and harvested every 4 h
for 68 h. We induced the formation of SGs with sodium arsenite (oxidative stress). On the other hand, we analyzed PBs in not
stressed N2A cells (neuroblastome cell line). We also studied the temporal expression profile of p-eIF2α (factor involved in
SG aggregation) and eIF3, the marker we used for detecting SGs, by Western blot. Cell cultures were arrested in order to
prevent cell cycle progression, and we confirm their quiescence state by flow cytometry. We performed a double
immunolabeling of SGs (eIF3 and G3BP1) and PB (GE-1/HEDLS and DDX6) by immunocytochemistry. 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 the estimated parameters showing periods of approximately 24 h, for two cycles. The subtle differences that we
found between both translation initiation factors eIF3 and eIF2α, would not be responsible for generating the changes observed
in SGs. These findings strongly suggest that the molecular circadian clock rhythmically controls SGs and PBs. The results
presented here reveal new ways in which circadian clocks modulate mRNA translation, stability, and storage in the cytoplasm
