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Heterogeneous nuclear RNA-ribonucleoprotein F binds to DNA via an oligo(dG)-motif and is associated with RNA
Background:
The heterogeneous nuclear ribonucleoprotein F (hnRNP-F) is one of the constituents of the splicing-related hnRNP complex. Recent studies suggest that pre-mRNA modification and splicing factors are associated with transcriptional initiation factors and RNA polymerase II (RNA pol II) at a promoter, implying that pre-mRNA-engaged factors might be associated with a promoter.
Results:
We isolated a cDNA of rat hnRNP-F and expressed the recombinant protein. HnRNP-F selectively bound to oligo(dG) in constructions with other oligohomonucleotides. The ssDNA of an SV40 promoter sequence having the GC-boxes was bound to hnRNP-F, while hnRNP-F bound to the (G/C)-stretch as dsDNA. Consequently, hnRNP-F was designated as an oligo(dG)-binding protein. None of the RNA-binding domains (RBDs) in hnRNP-F were critical for the oligo(dG)-binding. In contrast, the GY-rich region that exists between RBD-II and -III exhibited an oligo(dG)-binding ability. We suggest that the GY-rich region is a novel DNA-binding motif. HnRNP-F was shown to contain two types of binding motifs for RNA and DNA. Additionally, we found that hnRNP-F was co-precipitated with RNA pol II. Interestingly, the RNA pol II holoenzyme also contained hnRNP-F. The tissue distribution profile of hnRNP-F was similar to that of both RNA pol II and TBP.
Conclusion:
The above results suggest that hnRNP-F is associated with a transcriptional initiation apparatus that includes RNA pol II. The DNA-binding ability of hnRNP-F might facilitate the entry of pre-mRNA modification/splicing factors at a promoter.
Insights
Heterogeneous nuclear ribonucleoprotein F (hnRNP-F) binds DNA and RNA, associating with RNA polymerase II. This suggests hnRNP-F plays a role in transcriptional initiation and pre-mRNA processing at gene promoters.
Area of Science:
- Molecular Biology
- Gene Regulation
- Protein-Nucleic Acid Interactions
Background:
- Heterogeneous nuclear ribonucleoprotein F (hnRNP-F) is part of the splicing-related hnRNP complex.
- Emerging evidence links pre-mRNA processing factors to transcriptional initiation at promoters.
Purpose of the Study:
- To characterize the DNA-binding properties of hnRNP-F.
- To investigate the association of hnRNP-F with transcriptional machinery.
Main Methods:
- Isolation and expression of rat hnRNP-F cDNA.
- Oligonucleotide binding assays (DNA and RNA).
- Co-immunoprecipitation with RNA polymerase II.
Main Results:
- hnRNP-F selectively binds oligo(dG) and GC-rich promoter DNA sequences.
- A novel GY-rich region, distinct from RNA-binding domains, mediates DNA binding.
- hnRNP-F co-precipitates with RNA polymerase II (RNA pol II) and is found in the RNA pol II holoenzyme.
- Tissue distribution of hnRNP-F mirrors that of RNA pol II and TATA-binding protein (TBP).
Conclusions:
- hnRNP-F functions as a DNA- and RNA-binding protein.
- The GY-rich region represents a novel DNA-binding motif.
- hnRNP-F is associated with the RNA pol II transcriptional initiation complex.
- Its DNA-binding capacity may facilitate the recruitment of pre-mRNA processing factors to promoters.