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hnRNP H is a component of a splicing enhancer complex that activates a c-src alternative exon in neuronal cells.
Mol Cell Biol. 1999 Jan; 19(1):69-77.MC

Abstract

The regulation of the c-src N1 exon is mediated by an intronic splicing enhancer downstream of the N1 5' splice site. Previous experiments showed that a set of proteins assembles onto the most conserved core of this enhancer sequence specifically in neuronal WERI-1 cell extracts. The most prominent components of this enhancer complex are the proteins hnRNP F, KSRP, and an unidentified protein of 58 kDa (p58). This p58 protein was purified from the WERI-1 cell nuclear extract by ammonium sulfate precipitation, Mono Q chromatography, and immunoprecipitation with anti-Sm antibody Y12. Peptide sequence analysis of purified p58 protein identified it as hnRNP H. Immunoprecipitation of hnRNP H cross-linked to the N1 enhancer RNA, as well as gel mobility shift analysis of the enhancer complex in the presence of hnRNP H-specific antibodies, confirmed that hnRNP H is a protein component of the splicing enhancer complex. Immunoprecipitation of splicing intermediates from in vitro splicing reactions with anti-hnRNP H antibody indicated that hnRNP H remains bound to the src pre-mRNA after the assembly of spliceosome. Partial immunodepletion of hnRNP H from the nuclear extract partially inactivated the splicing of the N1 exon in vitro. This inhibition of splicing can be restored by the addition of recombinant hnRNP H, indicating that hnRNP H is an important factor for N1 splicing. Finally, in vitro binding assays demonstrate that hnRNP H can interact with the related protein hnRNP F, suggesting that hnRNPs H and F may exist as a heterodimer in a single enhancer complex. These two proteins presumably cooperate with each other and with other enhancer complex proteins to direct splicing to the N1 exon upstream.

Authors+Show Affiliations

Howard Hughes Medical Institute, University of California, Los Angeles, Los Angeles, California 90095, USA.No affiliation info availableNo affiliation info availableNo affiliation info available

Pub Type(s)

Journal Article
Research Support, U.S. Gov't, P.H.S.

Language

eng

PubMed ID

9858532

Citation

Chou, M Y., et al. "HnRNP H Is a Component of a Splicing Enhancer Complex That Activates a C-src Alternative Exon in Neuronal Cells." Molecular and Cellular Biology, vol. 19, no. 1, 1999, pp. 69-77.
Chou MY, Rooke N, Turck CW, et al. HnRNP H is a component of a splicing enhancer complex that activates a c-src alternative exon in neuronal cells. Mol Cell Biol. 1999;19(1):69-77.
Chou, M. Y., Rooke, N., Turck, C. W., & Black, D. L. (1999). HnRNP H is a component of a splicing enhancer complex that activates a c-src alternative exon in neuronal cells. Molecular and Cellular Biology, 19(1), 69-77.
Chou MY, et al. HnRNP H Is a Component of a Splicing Enhancer Complex That Activates a C-src Alternative Exon in Neuronal Cells. Mol Cell Biol. 1999;19(1):69-77. PubMed PMID: 9858532.
* Article titles in AMA citation format should be in sentence-case
TY - JOUR T1 - hnRNP H is a component of a splicing enhancer complex that activates a c-src alternative exon in neuronal cells. AU - Chou,M Y, AU - Rooke,N, AU - Turck,C W, AU - Black,D L, PY - 1998/12/22/pubmed PY - 1998/12/22/medline PY - 1998/12/22/entrez SP - 69 EP - 77 JF - Molecular and cellular biology JO - Mol Cell Biol VL - 19 IS - 1 N2 - The regulation of the c-src N1 exon is mediated by an intronic splicing enhancer downstream of the N1 5' splice site. Previous experiments showed that a set of proteins assembles onto the most conserved core of this enhancer sequence specifically in neuronal WERI-1 cell extracts. The most prominent components of this enhancer complex are the proteins hnRNP F, KSRP, and an unidentified protein of 58 kDa (p58). This p58 protein was purified from the WERI-1 cell nuclear extract by ammonium sulfate precipitation, Mono Q chromatography, and immunoprecipitation with anti-Sm antibody Y12. Peptide sequence analysis of purified p58 protein identified it as hnRNP H. Immunoprecipitation of hnRNP H cross-linked to the N1 enhancer RNA, as well as gel mobility shift analysis of the enhancer complex in the presence of hnRNP H-specific antibodies, confirmed that hnRNP H is a protein component of the splicing enhancer complex. Immunoprecipitation of splicing intermediates from in vitro splicing reactions with anti-hnRNP H antibody indicated that hnRNP H remains bound to the src pre-mRNA after the assembly of spliceosome. Partial immunodepletion of hnRNP H from the nuclear extract partially inactivated the splicing of the N1 exon in vitro. This inhibition of splicing can be restored by the addition of recombinant hnRNP H, indicating that hnRNP H is an important factor for N1 splicing. Finally, in vitro binding assays demonstrate that hnRNP H can interact with the related protein hnRNP F, suggesting that hnRNPs H and F may exist as a heterodimer in a single enhancer complex. These two proteins presumably cooperate with each other and with other enhancer complex proteins to direct splicing to the N1 exon upstream. SN - 0270-7306 UR - https://www.unboundmedicine.com/medline/citation/9858532/hnRNP_H_is_a_component_of_a_splicing_enhancer_complex_that_activates_a_c_src_alternative_exon_in_neuronal_cells_ L2 - https://journals.asm.org/doi/10.1128/MCB.19.1.69?url_ver=Z39.88-2003&rfr_id=ori:rid:crossref.org&rfr_dat=cr_pub=pubmed DB - PRIME DP - Unbound Medicine ER -