Dynamic interactions between splicing snRNPs, coiled bodies and nucleoli revealed using snRNP protein fusions to the green fluorescent protein.

摘要: The U1, U2, U4/U6, and U5 small nuclear ribonucleoproteins (snRNPs) are subunits of splicing complexes that remove introns from mRNA precursors. snRNPs show a complex, transcription-dependent localization pattern in the nucleoplasm mammalian cells results their association with several distinct subnuclear structures, including interchromatin granule clusters, perichromatin fibrils, coiled bodies. Here we report analysis snRNP interaction body live human using fusions proteins p80 coilin to Green Fluorescent Protein (GFP). Despite large size GFP tag, both core SmE U1 specific U1A assemble into particles give an identical endogenous counterparts. GFP-coilin localizes specifically bodies fashion provides accurate marker for variety cell lines. Treatment selective ser/thr-protein phosphatase inhibitor, okadaic acid, causes GFP-snRNP accumulate within nucleoli, but does not result nucleolar accumulation GFP-fused non-snRNP protein factor ASF/SF2. In all four lines tested, expression mutant single serine aspartate substitution also caused coilin, ASF/SF2, structures resembling when viewed by electron microscopy. This work establishes experimental system analyzing trafficking living evidence reversible phosphorylation mechanism is involved regulating nucleolus.