Live-cell spectroscopy and single-particle tracking revealed that eIF4E binding to 4E-BP1 triggers its release from the mRNA 5' cap upon mTOR inhibition, while eIF4G resides longer on the mRNA.
Live-cell spectroscopy and single-particle tracking can resolve the binding dynamics of endogenous translation initiation factors at single-cell and single-mRNA resolution.
mRNA translation is tightly regulated to preserve cellular homeostasis. Despite extensive biochemical, genetic, and structural studies, a detailed understanding of mRNA translation regulation is lacking. Imaging methodologies able to resolve the binding dynamics of translation factors at single-cell and single-mRNA resolution were necessary to fully elucidate regulation of this paramount process. Here live-cell spectroscopy and single-particle tracking were combined to interrogate the binding dynamics of endogenous initiation factors to the 5'cap. The diffusion of initiation factors (IFs) changed markedly upon their association with mRNA. Quantifying their diffusion characteristics revealed the sequence of IFs assembly and disassembly in cell lines and the clustering of translation in neurons. This approach revealed translation regulation at high spatial and temporal resolution that can be applied to the formation of any endogenous complex that results in a measurable shift in diffusion.
Gandin et al. (Wed,) conducted a other in Translation initiation dynamics. Torin-1 (mTOR inhibitor) vs. Vehicle (DMSO) was evaluated on Diffusion characteristics and binding dynamics of translation initiation factors. Live-cell spectroscopy and single-particle tracking revealed that eIF4E binding to 4E-BP1 triggers its release from the mRNA 5' cap upon mTOR inhibition, while eIF4G resides longer on the mRNA.
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