Sequences enriched in Alu repeats drive nuclear localization of long RNAs in human cells

AbstractHelicobacter pylori (Hp) CagA protein has a key role in the development of gastric cancer by the intruding in many intracellular processes of host human cell. Endoplasmic reticulum (ER) stress is an essential process for cellular homeostasis that modulates survival and death and is linked to several complex diseases including cancer. CagA protein is found in the serum of Hp-positive individuals and also in the supernatant of Hp culture. Limited studies report that recombinant CagA can alter gene expression and signaling pathways and induce the death of human cells. In this study, we investigated the effect of exogenous recombinant CagA protein treatment on ER stress and autophagy of human cell. AGS, MKN45, and HEK293 cells were treated with 1 µg/ml of recombinant CagA protein and then ER stress was studied by quantitative-PCR of spliced XBP-1 mRNA, immunofluorescence staining of ATF6 protein nuclear localization and real-time quantitative-PCR and/or western blot expression of GRP78, GRP94, ATF4 and CHOP genes. Autophagy was studied by western blot assessment of the conversion of LC3-I to LC3-II and LC3 aggregation. Cell proliferation and death were investigated by MTT assay and trypan blue staining respectively. As result, treatment with recombinant CagA enhanced XBP-1mRNA splicing, nuclear localization of ATF6, and the expression of ER stress signaling target genes in the cells. Recombinant CagA also induced LC3 protein conversion and aggregation in the cells. Reduced cell proliferation and increased cell death were determined in the cells treated with recombinant CagA. These results show that exogenous recombinant CagA protein causes cell death by inducing ER stress and autophagy in human cells. We conclude that CagA protein exogenously localizes in/on human cells and induces ER stress via disturbing protein machinery leading the human cell to death, however, the mechanism of CagA-host cell interaction is to be investigated.

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Sequences enriched in Alu repeats drive nuclear localization of long RNAs in human cells

10.1101/189746 ◽

2017 ◽

Cited By ~ 2

Author(s):

Yoav Lubelsky ◽

Igor Ulitsky

Keyword(s):

Nuclear Localization ◽

Nuclear Protein ◽

Noncoding Rnas ◽

Nuclear Accumulation ◽

Untranslated Regions ◽

Alu Elements ◽

Alu Repeats ◽

Key Players ◽

Cellular Pathways ◽

Rna Accumulation

AbstractLong noncoding RNAs (lncRNAs) are emerging as key players in multiple cellular pathways, but their modes of action, and how those are dictated by sequence remain elusive. While lncRNAs share most molecular properties with mRNAs, they are more likely to be enriched in the nucleus, a feature that is likely to be crucial for function of many lncRNAs, but whose molecular underpinnings remain largely unclear. In order identify elements that can force nuclear localization we screened libraries of short fragments tiled across nuclear RNAs, which were cloned into the untranslated regions of an efficiently exported mRNA. The screen identified a short sequence derived from Alu elements and found in many mRNAs and lncRNAs that increases nuclear accumulation and reduces overall expression levels. Measurements of the contribution of individual bases and short motifs to the element functionality identified a combination of RCCTCCC motifs that are bound by the abundant nuclear protein HNRNPK. Increased HNRNPK binding and C-rich motifs are predictive of substantial nuclear enrichment in both lncRNAs and mRNAs, and this mechanism is conserved across species. Our results thus detail a novel pathway for regulation of RNA accumulation and subcellular localization that has been co-opted to regulate the fate of transcripts that integrated Alu elements.

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Nuclear-localized Asunder regulates cytoplasmic dynein localization via its role in the Integrator complex

Molecular Biology of the Cell ◽

10.1091/mbc.e13-05-0254 ◽

2013 ◽

Vol 24 (18) ◽

pp. 2954-2965 ◽

Cited By ~ 10

Author(s):

Jeanne N. Jodoin ◽

Poojitha Sitaram ◽

Todd R. Albrecht ◽

Sarah B. May ◽

Mohammad Shboul ◽

...

Keyword(s):

Nuclear Localization ◽

Rna Processing ◽

Small Interfering Rna ◽

Cytoplasmic Dynein ◽

Human Cells ◽

Nuclear Localization Sequence ◽

Small Nuclear Rnas ◽

Localization Sequence ◽

Cultured Human Cells ◽

Interfering Rna

We previously reported that Asunder (ASUN) is essential for recruitment of dynein motors to the nuclear envelope (NE) and nucleus–centrosome coupling at the onset of cell division in cultured human cells and Drosophila spermatocytes, although the mechanisms underlying this regulation remain unknown. We also identified ASUN as a functional component of Integrator (INT), a multisubunit complex required for 3′-end processing of small nuclear RNAs. We now provide evidence that ASUN acts in the nucleus in concert with other INT components to mediate recruitment of dynein to the NE. Knockdown of other individual INT subunits in HeLa cells recapitulates the loss of perinuclear dynein in ASUN–small interfering RNA cells. Forced localization of ASUN to the cytoplasm via mutation of its nuclear localization sequence blocks its capacity to restore perinuclear dynein in both cultured human cells lacking ASUN and Drosophila asun spermatocytes. In addition, the levels of several INT subunits are reduced at G2/M when dynein is recruited to the NE, suggesting that INT does not directly mediate this step. Taken together, our data support a model in which a nuclear INT complex promotes recruitment of cytoplasmic dynein to the NE, possibly via a mechanism involving RNA processing.

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Mutational Analysis of Vpr-Induced G2Arrest, Nuclear Localization, and Cell Death in Fission Yeast

Journal of Virology ◽

10.1128/jvi.73.4.3236-3245.1999 ◽

1999 ◽

Vol 73 (4) ◽

pp. 3236-3245 ◽

Cited By ~ 51

Author(s):

Mingzhong Chen ◽

Robert T. Elder ◽

Min Yu ◽

Maurice G. O’Gorman ◽

Luc Selig ◽

...

Keyword(s):

Cell Cycle ◽

Cell Death ◽

Fission Yeast ◽

Nuclear Localization ◽

Mutational Analysis ◽

Single Point ◽

Point Mutations ◽

Cell Killing ◽

Human Cells ◽

Α Helix

ABSTRACT Cell cycle G2 arrest, nuclear localization, and cell death induced by human immunodeficiency virus type 1 Vpr were examined in fission yeast by using a panel of Vpr mutations that have been studied previously in human cells. The effects of the mutations on Vpr functions were highly similar between fission yeast and human cells. Consistent with mammalian cell studies, induction of cell cycle G2 arrest by Vpr was found to be independent of nuclear localization. In addition, G2 arrest was also shown to be independent of cell killing, which only occurred when the mutant Vpr localized to the nucleus. The C-terminal end of Vpr is crucial for G2 arrest, the N-terminal α-helix is important for nuclear localization, and a large part of the Vpr protein is responsible for cell killing. It is evident that the overall structure of Vpr is essential for these cellular effects, as N- and C-terminal deletions affected all three cellular functions. Furthermore, two single point mutations (H33R and H71R), both of which reside at the end of each α-helix, disrupted all three Vpr functions, indicating that these two mutations may have strong effects on the overall Vpr structure. The similarity of the mutant effects on Vpr function in fission yeast and human cells suggests that fission yeast can be used as a model system to evaluate these Vpr functions in naturally occurring viral isolates.

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025 Three-dimensional analysis of nuclear localization of repair sites of UV-induced DNA damage in human cells

Journal of Dermatological Science ◽

10.1016/s0923-1811(97)81730-6 ◽

1997 ◽

Vol 15 (2) ◽

pp. 107

Author(s):

A. Nakagawa ◽

N. Kobayashj ◽

Y. Yamashina ◽

T. Muramatsu ◽

T. Shirai ◽

...

Keyword(s):

Dna Damage ◽

Dimensional Analysis ◽

Nuclear Localization ◽

Three Dimensional ◽

Human Cells

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386: Nuclear Localization of ERBB3: A Potential Diagnostic/Prognostic Marker of Prostate Cancer

The Journal of Urology ◽

10.1016/s0022-5347(18)34639-1 ◽

2005 ◽

Vol 173 (4S) ◽

pp. 105-106

Author(s):

Ismaël H. Koumakpayi ◽

Jean-Simon Diallo ◽

Cecile Le Page ◽

Laurent Lessard ◽

Martin E. Gleave ◽

...

Keyword(s):

Prostate Cancer ◽

Prognostic Marker ◽

Nuclear Localization

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Alpha-Tocopherol Protects Cultured Human Cells from the Acute Lethal Cytotoxicity of Dioxin

International Journal for Vitamin and Nutrition Research ◽

10.1024/0300-9831.72.3.147 ◽

2002 ◽

Vol 72 (3) ◽

pp. 147-153 ◽

Cited By ~ 8

Author(s):

Kei-Ichi Hirai ◽

Jie-Hong Pan ◽

Ying-Bo Shui ◽

Eriko Simamura ◽

Hiroki Shimada ◽

...

Keyword(s):

Cell Death ◽

Cell Damage ◽

Smooth Endoplasmic Reticulum ◽

Dehydroascorbic Acid ◽

Human Cells ◽

Alpha Tocopherol ◽

Cervical Cancer Cells ◽

Cultured Human Cells ◽

Nuclear Damage ◽

Conjunctival Epithelial Cells

The possible protection of cultured human cells from acute dioxin injury by antioxidants was investigated. The most potent dioxin, 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD), caused vacuolization of the smooth endoplasmic reticulum and Golgi apparatus in cultured human conjunctival epithelial cells and cervical cancer cells. Subsequent nuclear damage included a deep irregular indentation resulting in cell death. A dosage of 30–40 ng/mL TCDD induced maximal intracellular production of H2O2 at 30 minutes and led to severe cell death (0–31% survival) at two hours. A dose of 1.7 mM alpha-tocopherol or 1 mM L-dehydroascorbic acid significantly protected human cells against acute TCDD injuries (78–97% survivals), but vitamin C did not provide this protection. These results indicate that accidental exposure to fatal doses of TCDD causes cytoplasmic free radical production within the smooth endoplasmic reticular systems, resulting in severe cytotoxicity, and that vitamin E and dehydroascorbic acid can protect against TCDD-induced cell damage.

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