Single-molecule imaging of translational output from individual RNA granules in neurons.
Dendritic RNAs are localized and translated in RNA granules. Here we use single-molecule imaging to count the number of RNA molecules in each granule and to record translation output from each granule using Venus fluorescent protein as a reporter. For RNAs encoding activity-regulated cytoskeletal-associated protein (ARC) or fragile X mental retardation protein (FMRP), translation events are spatially clustered near individual granules, and translational output from individual granules is either sporadic or bursty. The probability of bursty translation is greater for Venus-FMRP RNA than for Venus-ARC RNA and is increased in Fmr1-knockout neurons compared to wild-type neurons. Dihydroxyphenylglycine (DHPG) increases the rate of sporadic translation and decreases bursty translation for Venus-FMRP and Venus-ARC RNAs. Single-molecule imaging of translation in individual granules provides new insight into molecular, spatial, and temporal regulation of translation in granules.
University of Connecticut Health Center, Farmington, CT 06030, USA.
Vedakumar
V
Tatavarty
Marius F
MF
Ifrim
Mikhail
M
Levin
George
G
Korza
Elisa
E
Barbarese
Ji
J
Yu
John H
JH
Carson
D000818:Animals; D001426:Bacterial Proteins / Q000096:biosynthesis / Q000235:genetics; D002478:Cells, Cultured; D003598:Cytoskeletal Proteins / Q000096:biosynthesis / Q000235:genetics; D051860:Fragile X Messenger Ribonucleoprotein 1 / Q000096:biosynthesis / Q000235:genetics; D005998:Glycine / Q000031:analogs & derivatives / Q000494:pharmacology; D006624:Hippocampus; D008164:Luminescent Proteins / Q000096:biosynthesis / Q000235:genetics; D057054:Molecular Imaging; D009419:Nerve Tissue Proteins / Q000096:biosynthesis / Q000235:genetics; D009474:Neurons / Q000378:metabolism*; D014176:Protein Biosynthesis; D012313:RNA / Q000235:genetics / Q000378:metabolism*; D051381:Rats; D012118:Resorcinols / Q000494:pharmacology
D001426:Bacterial Proteins; D003598:Cytoskeletal Proteins; D051860:Fragile X Messenger Ribonucleoprotein 1; D005998:Glycine; D008164:Luminescent Proteins; D009419:Nerve Tissue Proteins; D012313:RNA; D012118:Resorcinols; C092069:activity regulated cytoskeletal-associated protein; C054562:yellow fluorescent protein, Bacteria; C079215:3,5-dihydroxyphenylglycine