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T-cell intracellular antigens function as tumor suppressor genes.

Sánchez-Jiménez C, Ludeña MD, Izquierdo JM - Cell Death Dis (2015)

Bottom Line: Genome-wide profiling illustrates a selective upregulation of p53 signaling pathway-related genes.Remarkably, low expressions of TIA1 and TIAR correlate with poor prognosis in patients with lung squamous cell carcinoma.These findings strongly support the concept that TIA proteins act as tumor suppressor genes.

View Article: PubMed Central - PubMed

Affiliation: Centro de Biología Molecular Severo Ochoa, Consejo Superior de Investigaciones Científicas, Universidad Autónoma de Madrid (CSIC/UAM), C/ Nicolás Cabrera 1, Madrid, Spain.

ABSTRACT
Knockdown of T-cell intracellular antigens TIA1 and TIAR in transformed cells triggers cell proliferation and tumor growth. Using a tetracycline-inducible system, we report here that an increased expression of TIA1 or TIAR in 293 cells results in reduced rates of cell proliferation. Ectopic expression of these proteins abolish endogenous TIA1 and TIAR levels via the regulation of splicing of their pre-mRNAs, and partially represses global translation in a phospho-eukaryotic initiation factor 2 alpha-dependent manner. This is accompanied by cell cycle arrest at G1/S and cell death through caspase-dependent apoptosis and autophagy. Genome-wide profiling illustrates a selective upregulation of p53 signaling pathway-related genes. Nude mice injected with doxycycline-inducible cells expressing TIA1 or TIAR retard, or even inhibit, growth of xenotumors. Remarkably, low expressions of TIA1 and TIAR correlate with poor prognosis in patients with lung squamous cell carcinoma. These findings strongly support the concept that TIA proteins act as tumor suppressor genes.

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Expression patterns of GFP-tagged proteins in tetracycline-induced FT293 cells. (a) Fluorescence images from FT293 cells expressing indicated proteins by confocal microcopy. The middle panels are the sum of GFP and To-Pro-3 images (Merge). Phase contrast photographs are also shown. Scale bars represent 20 μm. The fluorescence levels were analyzed by flow cytometry (b–d) Time-course and expression profiles of ectopic and endogenous TIA1, TIAR and HuR proteins in indicated FT293 cells by immunoblotting
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fig1: Expression patterns of GFP-tagged proteins in tetracycline-induced FT293 cells. (a) Fluorescence images from FT293 cells expressing indicated proteins by confocal microcopy. The middle panels are the sum of GFP and To-Pro-3 images (Merge). Phase contrast photographs are also shown. Scale bars represent 20 μm. The fluorescence levels were analyzed by flow cytometry (b–d) Time-course and expression profiles of ectopic and endogenous TIA1, TIAR and HuR proteins in indicated FT293 cells by immunoblotting

Mentions: The Flp-In T-REx System was used to generate inducible FT293 cell lines expressing GFP, TIA1, TIAR or HuR proteins, as well as truncated versions of TIA1 and TIAR lacking the C-terminal Q-rich (ΔQ) domains. The subcellular localization and protein expression patterns from different GFP-tagged constructs, expressed in FT293 cells for 3 days, were examined by confocal microscopy and western blotting. Results showed that the expression of tetracycline-inducible TIA1, TIA1ΔQ, TIAR, TIARΔQ and HuR proteins was consistent with their well-established patterns of nucleocytoplasmic and nuclear (with nucleolar exclusion) localizations for endogenous TIA1, TIAR and HuR, respectively (Figure 1a and refs 27, 28). Compared with the oxidative stress agent arsenite, analysis demonstrated that cells overexpressing TIA proteins did not trigger the massive formation of stress granules (Figure 1a and Supplementary Figure S1). However, ectopic TIA1 or TIAR expression promoted the depletion of endogenous TIA proteins (Figures 1b and Supplementary Figure S2A). This depletion occurred in a ΔQ domain-dependent manner (Figures 1b and Supplementary Figures S2B and D) and was associated solely with expression of TIA proteins as the ectopic expression of TIA1 or TIAR did not alter endogenous HuR expression (Figures 1b and d and Supplementary Figure S2). Similarly, the endogenous expression of TIA1 and TIAR was unaffected by ectopic HuR expression, whereas the endogenous HuR levels were decreased (Supplementary Figure S2A and ref. 29). Importantly, α-tubulin levels were not significantly changed (Figures 1b and d and Supplementary Figure S2A). These results are consistent with the notion of regulatory crosstalk between TIA proteins.


T-cell intracellular antigens function as tumor suppressor genes.

Sánchez-Jiménez C, Ludeña MD, Izquierdo JM - Cell Death Dis (2015)

Expression patterns of GFP-tagged proteins in tetracycline-induced FT293 cells. (a) Fluorescence images from FT293 cells expressing indicated proteins by confocal microcopy. The middle panels are the sum of GFP and To-Pro-3 images (Merge). Phase contrast photographs are also shown. Scale bars represent 20 μm. The fluorescence levels were analyzed by flow cytometry (b–d) Time-course and expression profiles of ectopic and endogenous TIA1, TIAR and HuR proteins in indicated FT293 cells by immunoblotting
© Copyright Policy - open-access
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC4385921&req=5

fig1: Expression patterns of GFP-tagged proteins in tetracycline-induced FT293 cells. (a) Fluorescence images from FT293 cells expressing indicated proteins by confocal microcopy. The middle panels are the sum of GFP and To-Pro-3 images (Merge). Phase contrast photographs are also shown. Scale bars represent 20 μm. The fluorescence levels were analyzed by flow cytometry (b–d) Time-course and expression profiles of ectopic and endogenous TIA1, TIAR and HuR proteins in indicated FT293 cells by immunoblotting
Mentions: The Flp-In T-REx System was used to generate inducible FT293 cell lines expressing GFP, TIA1, TIAR or HuR proteins, as well as truncated versions of TIA1 and TIAR lacking the C-terminal Q-rich (ΔQ) domains. The subcellular localization and protein expression patterns from different GFP-tagged constructs, expressed in FT293 cells for 3 days, were examined by confocal microscopy and western blotting. Results showed that the expression of tetracycline-inducible TIA1, TIA1ΔQ, TIAR, TIARΔQ and HuR proteins was consistent with their well-established patterns of nucleocytoplasmic and nuclear (with nucleolar exclusion) localizations for endogenous TIA1, TIAR and HuR, respectively (Figure 1a and refs 27, 28). Compared with the oxidative stress agent arsenite, analysis demonstrated that cells overexpressing TIA proteins did not trigger the massive formation of stress granules (Figure 1a and Supplementary Figure S1). However, ectopic TIA1 or TIAR expression promoted the depletion of endogenous TIA proteins (Figures 1b and Supplementary Figure S2A). This depletion occurred in a ΔQ domain-dependent manner (Figures 1b and Supplementary Figures S2B and D) and was associated solely with expression of TIA proteins as the ectopic expression of TIA1 or TIAR did not alter endogenous HuR expression (Figures 1b and d and Supplementary Figure S2). Similarly, the endogenous expression of TIA1 and TIAR was unaffected by ectopic HuR expression, whereas the endogenous HuR levels were decreased (Supplementary Figure S2A and ref. 29). Importantly, α-tubulin levels were not significantly changed (Figures 1b and d and Supplementary Figure S2A). These results are consistent with the notion of regulatory crosstalk between TIA proteins.

Bottom Line: Genome-wide profiling illustrates a selective upregulation of p53 signaling pathway-related genes.Remarkably, low expressions of TIA1 and TIAR correlate with poor prognosis in patients with lung squamous cell carcinoma.These findings strongly support the concept that TIA proteins act as tumor suppressor genes.

View Article: PubMed Central - PubMed

Affiliation: Centro de Biología Molecular Severo Ochoa, Consejo Superior de Investigaciones Científicas, Universidad Autónoma de Madrid (CSIC/UAM), C/ Nicolás Cabrera 1, Madrid, Spain.

ABSTRACT
Knockdown of T-cell intracellular antigens TIA1 and TIAR in transformed cells triggers cell proliferation and tumor growth. Using a tetracycline-inducible system, we report here that an increased expression of TIA1 or TIAR in 293 cells results in reduced rates of cell proliferation. Ectopic expression of these proteins abolish endogenous TIA1 and TIAR levels via the regulation of splicing of their pre-mRNAs, and partially represses global translation in a phospho-eukaryotic initiation factor 2 alpha-dependent manner. This is accompanied by cell cycle arrest at G1/S and cell death through caspase-dependent apoptosis and autophagy. Genome-wide profiling illustrates a selective upregulation of p53 signaling pathway-related genes. Nude mice injected with doxycycline-inducible cells expressing TIA1 or TIAR retard, or even inhibit, growth of xenotumors. Remarkably, low expressions of TIA1 and TIAR correlate with poor prognosis in patients with lung squamous cell carcinoma. These findings strongly support the concept that TIA proteins act as tumor suppressor genes.

Show MeSH
Related in: MedlinePlus