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Growth factor dependent regulation of centrosome function and genomic instability by HuR.

Filippova N, Yang X, Nabors LB - Biomolecules (2015)

Bottom Line: This process is regulated by tyrosine phosphorylation of HuR and is abolished by mutating tyrosine residues.HuR is overexpressed in tumor samples from patients with glioblastoma and associated with a reduced survival.These findings suggest HuR plays a significant role in centrosome amplification and genomic instability, which contributes to a worse disease outcome.

View Article: PubMed Central - PubMed

Affiliation: Department of Neurology, University of Alabama at Birmingham, 510 20th Street South, FOT 1020, Birmingham, AL 35209, USA. as1999@uab.edu.

ABSTRACT
The mRNA binding protein HuR is over expressed in cancer cells and contributes to disease progression through post-transcriptional regulation of mRNA. The regulation of HuR and how this relates to glioma is the focus of this report. SRC and c-Abl kinases regulate HuR sub-cellular trafficking and influence accumulation in the pericentriolar matrix (PCM) via a growth factor dependent signaling mechanism. Growth factor stimulation of glioma cell lines results in the associate of HuR with the PCM and amplification of centrosome number. This process is regulated by tyrosine phosphorylation of HuR and is abolished by mutating tyrosine residues. HuR is overexpressed in tumor samples from patients with glioblastoma and associated with a reduced survival. These findings suggest HuR plays a significant role in centrosome amplification and genomic instability, which contributes to a worse disease outcome.

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Stimulation with growth factors induces centrosome amplification in U251 cells. (A) Examples of centrosomes marked with the PACT domain of pericentrin attached to the red fluorescence protein (PACT-mKO1) in clones of U251 cells in control (unstimulated) or stimulated with growth factors (EGF, 40 nM and bFGF 40 nM). There is amplification of centrosomes in cells treated with growth factors compared to the control. Nuclei were stained with DAPI. (B) The graph represents the averaged percentages of cells with amplified centrosome number following stimulation with growth factors (87% ± 8%) versus control (22% ± 6%). The difference is significant with p = 0.0008. The experiments were performed four times. The insert illustrates the total level of γ-tubulin values in cells non-treated (C) and treated with growth factors (GrF). Note that growth factor treatment induced an increase of γ-tubulin/GAPDH ratio (by 44% and 57% in two illustrated experiments in Western blot). The increase is significant (50% ± 7%, p = 0.005, three experiments).
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biomolecules-05-00263-f001: Stimulation with growth factors induces centrosome amplification in U251 cells. (A) Examples of centrosomes marked with the PACT domain of pericentrin attached to the red fluorescence protein (PACT-mKO1) in clones of U251 cells in control (unstimulated) or stimulated with growth factors (EGF, 40 nM and bFGF 40 nM). There is amplification of centrosomes in cells treated with growth factors compared to the control. Nuclei were stained with DAPI. (B) The graph represents the averaged percentages of cells with amplified centrosome number following stimulation with growth factors (87% ± 8%) versus control (22% ± 6%). The difference is significant with p = 0.0008. The experiments were performed four times. The insert illustrates the total level of γ-tubulin values in cells non-treated (C) and treated with growth factors (GrF). Note that growth factor treatment induced an increase of γ-tubulin/GAPDH ratio (by 44% and 57% in two illustrated experiments in Western blot). The increase is significant (50% ± 7%, p = 0.005, three experiments).

Mentions: The amplification of centrosomes is growth factor dependent and can be induced by EGF and bFGF stimulation of cancer cells [10,11,12,13,14]. Figure 1 illustrates the amplification of centrosomes marked with the PACT domain of pericentrin in clones of U251 cells treated with bFGF (40 nM) and EGF (40 nM) for 48 h. We observed that 87% ± 8% of U251 cells had more than two centrosomes per nucleus following stimulation with bFGF and EGF compared to 22% ± 6% of cells with more than two centrosomes per nucleus in the control condition. The difference is significant (p = 0.0008). Similar results have been achieved in four experiments.


Growth factor dependent regulation of centrosome function and genomic instability by HuR.

Filippova N, Yang X, Nabors LB - Biomolecules (2015)

Stimulation with growth factors induces centrosome amplification in U251 cells. (A) Examples of centrosomes marked with the PACT domain of pericentrin attached to the red fluorescence protein (PACT-mKO1) in clones of U251 cells in control (unstimulated) or stimulated with growth factors (EGF, 40 nM and bFGF 40 nM). There is amplification of centrosomes in cells treated with growth factors compared to the control. Nuclei were stained with DAPI. (B) The graph represents the averaged percentages of cells with amplified centrosome number following stimulation with growth factors (87% ± 8%) versus control (22% ± 6%). The difference is significant with p = 0.0008. The experiments were performed four times. The insert illustrates the total level of γ-tubulin values in cells non-treated (C) and treated with growth factors (GrF). Note that growth factor treatment induced an increase of γ-tubulin/GAPDH ratio (by 44% and 57% in two illustrated experiments in Western blot). The increase is significant (50% ± 7%, p = 0.005, three experiments).
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biomolecules-05-00263-f001: Stimulation with growth factors induces centrosome amplification in U251 cells. (A) Examples of centrosomes marked with the PACT domain of pericentrin attached to the red fluorescence protein (PACT-mKO1) in clones of U251 cells in control (unstimulated) or stimulated with growth factors (EGF, 40 nM and bFGF 40 nM). There is amplification of centrosomes in cells treated with growth factors compared to the control. Nuclei were stained with DAPI. (B) The graph represents the averaged percentages of cells with amplified centrosome number following stimulation with growth factors (87% ± 8%) versus control (22% ± 6%). The difference is significant with p = 0.0008. The experiments were performed four times. The insert illustrates the total level of γ-tubulin values in cells non-treated (C) and treated with growth factors (GrF). Note that growth factor treatment induced an increase of γ-tubulin/GAPDH ratio (by 44% and 57% in two illustrated experiments in Western blot). The increase is significant (50% ± 7%, p = 0.005, three experiments).
Mentions: The amplification of centrosomes is growth factor dependent and can be induced by EGF and bFGF stimulation of cancer cells [10,11,12,13,14]. Figure 1 illustrates the amplification of centrosomes marked with the PACT domain of pericentrin in clones of U251 cells treated with bFGF (40 nM) and EGF (40 nM) for 48 h. We observed that 87% ± 8% of U251 cells had more than two centrosomes per nucleus following stimulation with bFGF and EGF compared to 22% ± 6% of cells with more than two centrosomes per nucleus in the control condition. The difference is significant (p = 0.0008). Similar results have been achieved in four experiments.

Bottom Line: This process is regulated by tyrosine phosphorylation of HuR and is abolished by mutating tyrosine residues.HuR is overexpressed in tumor samples from patients with glioblastoma and associated with a reduced survival.These findings suggest HuR plays a significant role in centrosome amplification and genomic instability, which contributes to a worse disease outcome.

View Article: PubMed Central - PubMed

Affiliation: Department of Neurology, University of Alabama at Birmingham, 510 20th Street South, FOT 1020, Birmingham, AL 35209, USA. as1999@uab.edu.

ABSTRACT
The mRNA binding protein HuR is over expressed in cancer cells and contributes to disease progression through post-transcriptional regulation of mRNA. The regulation of HuR and how this relates to glioma is the focus of this report. SRC and c-Abl kinases regulate HuR sub-cellular trafficking and influence accumulation in the pericentriolar matrix (PCM) via a growth factor dependent signaling mechanism. Growth factor stimulation of glioma cell lines results in the associate of HuR with the PCM and amplification of centrosome number. This process is regulated by tyrosine phosphorylation of HuR and is abolished by mutating tyrosine residues. HuR is overexpressed in tumor samples from patients with glioblastoma and associated with a reduced survival. These findings suggest HuR plays a significant role in centrosome amplification and genomic instability, which contributes to a worse disease outcome.

Show MeSH
Related in: MedlinePlus