Heterogeneous nuclear ribonucleoprotein E1 binds polycytosine DNA and monitors genome integrity

Heterogeneous nuclear ribonucleoprotein E1 (hnRNP E1) is a tumor suppressor protein that binds site- and structure-specifically to RNA sequences to regulate mRNA stability, facilitate alternative splicing, and suppress protein translation on several metastasis-associated mRNAs. Here, we show that hnRNP E1 binds polycytosine-rich DNA tracts present throughout the genome, including those at promoters of several oncogenes and telomeres and monitors genome integrity. It binds DNA in a site- and structure-specific manner. hnRNP E1-knockdown cells displayed increased DNA damage signals including γ-H2AX at its binding sites and also showed increased mutations. UV and hydroxyurea treatment of hnRNP E1-knockdown cells exacerbated the basal DNA damage signals with increased cell cycle arrest, activation of checkpoint proteins, and monoubiquitination of proliferating cell nuclear antigen despite no changes in deubiquitinating enzymes. DNA damage caused by genotoxin treatment localized to hnRNP E1 binding sites. Our work suggests that hnRNP E1 facilitates functions of DNA integrity proteins at polycytosine tracts and monitors DNA integrity at these sites.

Download Full-text

Distinct Sets of Adjacent Heterogeneous Nuclear Ribonucleoprotein (hnRNP) A1/A2 Binding Sites Control 5′ Splice Site Selection in the hnRNP A1 mRNA Precursor

Journal of Biological Chemistry ◽

10.1074/jbc.m203633200 ◽

2002 ◽

Vol 277 (33) ◽

pp. 29745-29752 ◽

Cited By ~ 54

Author(s):

Stephen Hutchison ◽

Catherine LeBel ◽

Marco Blanchette ◽

Benoit Chabot

Keyword(s):

Splice Site ◽

Site Selection ◽

Binding Sites ◽

Hnrnp A1 ◽

Splice Site Selection ◽

Heterogeneous Nuclear Ribonucleoprotein ◽

Nuclear Ribonucleoprotein ◽

Mrna Precursor

Download Full-text

Heterogeneous Nuclear Ribonucleoprotein (hnRNP) E1 Binds to hnRNP A2 and Inhibits Translation of A2 Response Element mRNAs

Molecular Biology of the Cell ◽

10.1091/mbc.e05-10-0946 ◽

2006 ◽

Vol 17 (8) ◽

pp. 3521-3533 ◽

Cited By ~ 51

Author(s):

Linda D. Kosturko ◽

Michael J. Maggipinto ◽

George Korza ◽

Joo Won Lee ◽

John H. Carson ◽

...

Keyword(s):

Rna Binding ◽

Binding Protein ◽

Rna Binding Protein ◽

In Vitro Translation ◽

Dependent Manner ◽

Response Element ◽

Heterogeneous Nuclear Ribonucleoprotein ◽

Nuclear Ribonucleoprotein ◽

Hnrnp E1

Heterogeneous nuclear ribonucleoprotein (hnRNP) A2 is a trans-acting RNA-binding protein that mediates trafficking of RNAs containing the cis-acting A2 response element (A2RE). Previous work has shown that A2RE RNAs are transported to myelin in oligodendrocytes and to dendrites in neurons. hnRNP E1 is an RNA-binding protein that regulates translation of specific mRNAs. Here, we show by yeast two-hybrid analysis, in vivo and in vitro coimmunoprecipitation, in vitro cross-linking, and fluorescence correlation spectroscopy that hnRNP E1 binds to hnRNP A2 and is recruited to A2RE RNA in an hnRNP A2-dependent manner. hnRNP E1 is colocalized with hnRNP A2 and A2RE mRNA in granules in dendrites of oligodendrocytes. Overexpression of hnRNP E1 or microinjection of exogenous hnRNP E1 in neural cells inhibits translation of A2RE mRNA, but not of non-A2RE RNA. Excess hnRNP E1 added to an in vitro translation system reduces translation efficiency of A2RE mRNA, but not of nonA2RE RNA, in an hnRNP A2-dependent manner. These results are consistent with a model where hnRNP E1 recruited to A2RE RNA granules by binding to hnRNP A2 inhibits translation of A2RE RNA during granule transport.

Download Full-text

A proteomics approach for the identification of nucleophosmin and heterogeneous nuclear ribonucleoprotein C1/C2 as chromatin-binding proteins in response to DNA double-strand breaks

Biochemical Journal ◽

10.1042/bj20042033 ◽

2005 ◽

Vol 388 (1) ◽

pp. 7-15 ◽

Cited By ~ 52

Author(s):

Seung Yun LEE ◽

Ji-Hye PARK ◽

Sungsu KIM ◽

Eun-Jung PARK ◽

Yungdae YUN ◽

...

Keyword(s):

Dna Damage ◽

Dna Repair ◽

Binding Proteins ◽

Double Strand Breaks ◽

Nuclear Proteins ◽

Chromatin Binding ◽

Strand Breaks ◽

Heterogeneous Nuclear Ribonucleoprotein ◽

Proteomics Approach ◽

Nuclear Ribonucleoprotein

Double-strand breaks (DSBs) of chromosomal DNA trigger the cellular response that activates the pathways for DNA repair and cell-cycle checkpoints, and sometimes the pathways leading to cell death if the damage is too severe to be tolerated. Evidence indicates that, upon generation of DNA DSBs, many nuclear proteins that are involved in DNA repair and checkpoints are recruited to chromatin around the DNA lesions. In the present study we used a proteomics approach to identify DNA-damage-induced chromatin-binding proteins in a systematic way. Two-dimensional gel analysis for protein extracts of chromatin from DNA-damage-induced and control HeLa cells identified four proteins as the candidates for DNA-damage-induced chromatin-binding proteins. MALDI–TOF (matrix-assisted laser-desorption ionization–time-of-flight) MS analysis identified these proteins to be NPM (nucleophosmin), hnRNP (heterogeneous nuclear ribonucleoprotein) C1, hnRNP C2 and 37-kDa laminin-receptor precursor, and the identity of these proteins was further confirmed by immunoblot analysis with specific antibodies. We then demonstrated with chromatin-binding assays that NPM and hnRNP C1/C2, the abundant nuclear proteins with pleiotropic functions, indeed bind to chromatin in a DNA-damage-dependent manner, implicating these proteins in DNA repair and/or damage response. Immunofluorescence experiments showed that NPM, normally present in the nucleoli, is mobilized into the nucleoplasm after DNA damage, and that neither NPM nor hnRNP C1/C2 is actively recruited to the sites of DNA breaks. These results suggest that NPM and hnRNP C1/C2 may function at the levels of the global context of chromatin, rather than by specifically targeting the broken DNA.

Download Full-text

Chemical Shift Mapped DNA-Binding Sites and15N Relaxation Analysis of the C-Terminal KH Domain of Heterogeneous Nuclear Ribonucleoprotein K

Biochemistry ◽

10.1021/bi000105e ◽

2000 ◽

Vol 39 (20) ◽

pp. 6022-6032 ◽

Cited By ~ 10

Author(s):

James L. Baber ◽

David Levens ◽

Daniel Libutti ◽

Nico Tjandra

Keyword(s):

Chemical Shift ◽

Dna Binding ◽

Binding Sites ◽

Relaxation Analysis ◽

Heterogeneous Nuclear Ribonucleoprotein ◽

Dna Binding Sites ◽

Kh Domain ◽

Nuclear Ribonucleoprotein

Download Full-text

DNA Damage-induced Heterogeneous Nuclear Ribonucleoprotein K SUMOylation Regulates p53 Transcriptional Activation

Journal of Biological Chemistry ◽

10.1074/jbc.m112.390120 ◽

2012 ◽

Vol 287 (36) ◽

pp. 30789-30799 ◽

Cited By ~ 48

Author(s):

Federico Pelisch ◽

Berta Pozzi ◽

Guillermo Risso ◽

Manuel Javier Muñoz ◽

Anabella Srebrow

Keyword(s):

Dna Damage ◽

Transcriptional Activation ◽

Heterogeneous Nuclear Ribonucleoprotein ◽

Nuclear Ribonucleoprotein

Download Full-text

Arginine methylation of hnRNPK negatively modulates apoptosis upon DNA damage through local regulation of phosphorylation

Nucleic Acids Research ◽

10.1093/nar/gku705 ◽

2014 ◽

Vol 42 (15) ◽

pp. 9908-9924 ◽

Cited By ~ 23

Author(s):

Jen-Hao Yang ◽

Yi-Ying Chiou ◽

Shu-Ling Fu ◽

I-Yun Shih ◽

Tsai-Hsuan Weng ◽

...

Keyword(s):

Dna Damage ◽

Rna Processing ◽

Tumor Development ◽

Arginine Methylation ◽

Wild Type ◽

Post Translational Modifications ◽

Heterogeneous Nuclear Ribonucleoprotein ◽

Damage Response ◽

Nuclear Ribonucleoprotein

Abstract Heterogeneous nuclear ribonucleoprotein K (hnRNPK) is an RNA/DNA-binding protein involved in chromatin remodeling, RNA processing and the DNA damage response. In addition, increased hnRNPK expression has been associated with tumor development and progression. A variety of post-translational modifications of hnRNPK have been identified and shown to regulate hnRNPK function, including phosphorylation, ubiquitination, sumoylation and methylation. However, the functional significance of hnRNPK arginine methylation remains unclear. In the present study, we demonstrated that the methylation of two essential arginines, Arg296 and Arg299, on hnRNPK inhibited a nearby Ser302 phosphorylation that was mediated through the pro-apoptotic kinase PKCδ. Notably, the engineered U2OS cells carrying an Arg296/Arg299 methylation-defective hnRNPK mutant exhibited increased apoptosis upon DNA damage. While such elevated apoptosis can be diminished through addition with wild-type hnRNPK, we further demonstrated that this increased apoptosis occurred through both intrinsic and extrinsic pathways and was p53 independent, at least in part. Here, we provide the first evidence that the arginine methylation of hnRNPK negatively regulates cell apoptosis through PKCδ-mediated signaling during DNA damage, which is essential for the anti-apoptotic role of hnRNPK in apoptosis and the evasion of apoptosis in cancer cells.

Download Full-text