Related Experiment Videos
The Y14 protein communicates to the cytoplasm the position of exon-exon junctions
1Howard Hughes Medical Institute and Department of Biochemistry & Biophysics, University of Pennsylvania School of Medicine, Philadelphia, PA 19104-6148, USA.
Abstract:
We recently described an RNA-binding protein, Y14, that binds preferentially to spliced mRNAs and persists in the cytoplasm. Y14 is part of a multi-protein complex that also contains the mRNA export factor TAP. This suggests that splicing imprints the mRNA with a unique set of proteins that communicate the history of the transcript to the cytoplasm. Here, using microinjection of pre-mRNAs into Xenopus oocyte nuclei followed by immunoprecipitation of RNase-fragmented mRNAs from the cytoplasm, we show that Y14 is stably bound to sequences immediately upstream of exon-exon junctions. This feature appears to be unique to Y14. Using monoclonal antibodies that we produced against Aly/REF, another component recently reported to be an mRNA export factor, we show that Aly/REF is associated with spliced mRNAs in the nucleus but is not detectable on mRNAs in the cytoplasm. Thus, we propose that the splicing- dependent binding of Y14 provides a position-specific molecular memory that communicates to the cytoplasm the location of exon and intron boundaries. This novel mechanism may play an important role in post-splicing events.
Insights
The RNA-binding protein Y14 binds to spliced messenger RNAs (mRNAs) near exon-exon junctions, marking transcript history for the cytoplasm. This splicing-dependent Y14 binding acts as a molecular memory, crucial for post-splicing events.
Area of Science:
- Molecular Biology
- RNA Biology
- Cell Biology
Background:
- Y14 is an RNA-binding protein found in the cytoplasm, associated with the mRNA export factor TAP.
- Splicing may imprint mRNAs with proteins that signal transcript history to the cytoplasm.
Purpose of the Study:
- To investigate the precise binding location of Y14 on spliced mRNAs.
- To determine the cytoplasmic localization of the mRNA export factor Aly/REF.
Main Methods:
- Microinjection of pre-mRNAs into Xenopus oocyte nuclei.
- Immunoprecipitation of RNase-fragmented mRNAs from the cytoplasm.
- Production and use of monoclonal antibodies against Aly/REF.
Main Results:
- Y14 binds stably to sequences upstream of exon-exon junctions on spliced mRNAs.
- Aly/REF is associated with spliced mRNAs in the nucleus but not in the cytoplasm.
Conclusions:
- Splicing-dependent Y14 binding provides position-specific molecular memory of exon-intron boundaries.
- This mechanism likely plays a significant role in post-splicing events and mRNA processing.