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Retinoblastoma loss modulates DNA damage response favoring tumor progression.

Seoane M, Iglesias P, Gonzalez T, Dominguez F, Fraga M, Aliste C, Forteza J, Costoya JA - PLoS ONE (2008)

Bottom Line: It has been described in premalignant lesions that OIS requires DNA damage response (DDR) activation, safeguard of the integrity of the genome.In human gliomas most of the genetic alterations that have been previously identified result in abnormal activation of cell growth signaling pathways and deregulation of cell cycle, features recapitulated in our model by oncogenic Ras expression and retinoblastoma (Rb) inactivation respectively.Moreover, Rb loss inactivates the stress kinase DDR-associated p38MAPK by specific Wip1-dependent dephosphorylation.

View Article: PubMed Central - PubMed

Affiliation: Molecular Oncology Lab, Departamento de Fisioloxia, Facultade de Medicina, Universidade de Santiago de Compostela, Santiago de Compostela, Spain.

ABSTRACT
Senescence is one of the main barriers against tumor progression. Oncogenic signals in primary cells result in oncogene-induced senescence (OIS), crucial for protection against cancer development. It has been described in premalignant lesions that OIS requires DNA damage response (DDR) activation, safeguard of the integrity of the genome. Here we demonstrate how the cellular mechanisms involved in oncogenic transformation in a model of glioma uncouple OIS and DDR. We use this tumor type as a paradigm of oncogenic transformation. In human gliomas most of the genetic alterations that have been previously identified result in abnormal activation of cell growth signaling pathways and deregulation of cell cycle, features recapitulated in our model by oncogenic Ras expression and retinoblastoma (Rb) inactivation respectively. In this scenario, the absence of pRb confers a proliferative advantage and activates DDR to a greater extent in a DNA lesion-independent fashion than cells that express only HRas(V12). Moreover, Rb loss inactivates the stress kinase DDR-associated p38MAPK by specific Wip1-dependent dephosphorylation. Thus, Rb loss acts as a switch mediating the transition between premalignant lesions and cancer through DDR modulation. These findings may have important implications for the understanding the biology of gliomas and anticipate a new target, Wip1 phosphatase, for novel therapeutic strategies.

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HRasV12 in the absence of Rb induces malignant transformation both in vitro and in vivo.a, Ability to form foci, representative pictures of wells stained with crystal violet (day 7). b, Representative pictures of wells stained on the indicated days of culture showing the morphological change observed in each group are also shown. c, Tumor-free survival curve (Kaplan-Meier plot) for tumor formation in SCID mice. Tumor was found in cRb−/−/RasV12 cohort (P<0.0001) d, Photographs of SCID mice after subcutaneous injection of cRbloxP/loxP/RasV12 and cRb−/−/RasV12 astrocytes at eleven and two weeks respectively. Actual sizes of tumors after biopsy. e. Low-grade gliomas (cRbloxP/loxP/RasV12; H&E, upper left) showed a rather monomorphic appearance, with polygonal or rounded cell shape and smaller rounded nuclei without prominent nucleoli. Note the absence of mitosis and necrosis. In contrast, high-grade gliomas (cRb−/−/RasV12; H&E, lower left) displayed cellular pleomorphism, with large, fusiform and irregular nuclei that frequently exhibit mitotic figures (upper left corner) and necrotic foci (lower right corner). Co-expression of EGFP permits the visualization of the previously modified astrocytes.
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pone-0003632-g002: HRasV12 in the absence of Rb induces malignant transformation both in vitro and in vivo.a, Ability to form foci, representative pictures of wells stained with crystal violet (day 7). b, Representative pictures of wells stained on the indicated days of culture showing the morphological change observed in each group are also shown. c, Tumor-free survival curve (Kaplan-Meier plot) for tumor formation in SCID mice. Tumor was found in cRb−/−/RasV12 cohort (P<0.0001) d, Photographs of SCID mice after subcutaneous injection of cRbloxP/loxP/RasV12 and cRb−/−/RasV12 astrocytes at eleven and two weeks respectively. Actual sizes of tumors after biopsy. e. Low-grade gliomas (cRbloxP/loxP/RasV12; H&E, upper left) showed a rather monomorphic appearance, with polygonal or rounded cell shape and smaller rounded nuclei without prominent nucleoli. Note the absence of mitosis and necrosis. In contrast, high-grade gliomas (cRb−/−/RasV12; H&E, lower left) displayed cellular pleomorphism, with large, fusiform and irregular nuclei that frequently exhibit mitotic figures (upper left corner) and necrotic foci (lower right corner). Co-expression of EGFP permits the visualization of the previously modified astrocytes.

Mentions: Unlike normal primary astrocytes, flattened and non-refractile cells, Ras-expressing astrocytes showed noticeable morphological changes and loss of contact inhibition, and although flat cells were also present, the population was heterogeneous and large displaying at the same time a refractile cytoplasm with thin and long projections (Figure 2A). It is also revealing the fact that although Rb-deficient astrocytes displayed a marked proliferative increase its morphology did not appear significantly changed. On the contrary, Ras-expressing astrocytes seemed to undergo dramatic morphological alterations suggesting a transformed phenotype. We then wondered if these astrocytes would also exhibit foci growth, which is a characteristic feature of transformed cells. As Figure 2B shows, HRasV12 seems to induce foci growth as seen in cRbloxP/loxP/RasV12 and cRb−/−/RasV12 cells, whereas the remaining groups do not show significant foci formation.


Retinoblastoma loss modulates DNA damage response favoring tumor progression.

Seoane M, Iglesias P, Gonzalez T, Dominguez F, Fraga M, Aliste C, Forteza J, Costoya JA - PLoS ONE (2008)

HRasV12 in the absence of Rb induces malignant transformation both in vitro and in vivo.a, Ability to form foci, representative pictures of wells stained with crystal violet (day 7). b, Representative pictures of wells stained on the indicated days of culture showing the morphological change observed in each group are also shown. c, Tumor-free survival curve (Kaplan-Meier plot) for tumor formation in SCID mice. Tumor was found in cRb−/−/RasV12 cohort (P<0.0001) d, Photographs of SCID mice after subcutaneous injection of cRbloxP/loxP/RasV12 and cRb−/−/RasV12 astrocytes at eleven and two weeks respectively. Actual sizes of tumors after biopsy. e. Low-grade gliomas (cRbloxP/loxP/RasV12; H&E, upper left) showed a rather monomorphic appearance, with polygonal or rounded cell shape and smaller rounded nuclei without prominent nucleoli. Note the absence of mitosis and necrosis. In contrast, high-grade gliomas (cRb−/−/RasV12; H&E, lower left) displayed cellular pleomorphism, with large, fusiform and irregular nuclei that frequently exhibit mitotic figures (upper left corner) and necrotic foci (lower right corner). Co-expression of EGFP permits the visualization of the previously modified astrocytes.
© Copyright Policy
Related In: Results  -  Collection

Show All Figures
getmorefigures.php?uid=PMC2573954&req=5

pone-0003632-g002: HRasV12 in the absence of Rb induces malignant transformation both in vitro and in vivo.a, Ability to form foci, representative pictures of wells stained with crystal violet (day 7). b, Representative pictures of wells stained on the indicated days of culture showing the morphological change observed in each group are also shown. c, Tumor-free survival curve (Kaplan-Meier plot) for tumor formation in SCID mice. Tumor was found in cRb−/−/RasV12 cohort (P<0.0001) d, Photographs of SCID mice after subcutaneous injection of cRbloxP/loxP/RasV12 and cRb−/−/RasV12 astrocytes at eleven and two weeks respectively. Actual sizes of tumors after biopsy. e. Low-grade gliomas (cRbloxP/loxP/RasV12; H&E, upper left) showed a rather monomorphic appearance, with polygonal or rounded cell shape and smaller rounded nuclei without prominent nucleoli. Note the absence of mitosis and necrosis. In contrast, high-grade gliomas (cRb−/−/RasV12; H&E, lower left) displayed cellular pleomorphism, with large, fusiform and irregular nuclei that frequently exhibit mitotic figures (upper left corner) and necrotic foci (lower right corner). Co-expression of EGFP permits the visualization of the previously modified astrocytes.
Mentions: Unlike normal primary astrocytes, flattened and non-refractile cells, Ras-expressing astrocytes showed noticeable morphological changes and loss of contact inhibition, and although flat cells were also present, the population was heterogeneous and large displaying at the same time a refractile cytoplasm with thin and long projections (Figure 2A). It is also revealing the fact that although Rb-deficient astrocytes displayed a marked proliferative increase its morphology did not appear significantly changed. On the contrary, Ras-expressing astrocytes seemed to undergo dramatic morphological alterations suggesting a transformed phenotype. We then wondered if these astrocytes would also exhibit foci growth, which is a characteristic feature of transformed cells. As Figure 2B shows, HRasV12 seems to induce foci growth as seen in cRbloxP/loxP/RasV12 and cRb−/−/RasV12 cells, whereas the remaining groups do not show significant foci formation.

Bottom Line: It has been described in premalignant lesions that OIS requires DNA damage response (DDR) activation, safeguard of the integrity of the genome.In human gliomas most of the genetic alterations that have been previously identified result in abnormal activation of cell growth signaling pathways and deregulation of cell cycle, features recapitulated in our model by oncogenic Ras expression and retinoblastoma (Rb) inactivation respectively.Moreover, Rb loss inactivates the stress kinase DDR-associated p38MAPK by specific Wip1-dependent dephosphorylation.

View Article: PubMed Central - PubMed

Affiliation: Molecular Oncology Lab, Departamento de Fisioloxia, Facultade de Medicina, Universidade de Santiago de Compostela, Santiago de Compostela, Spain.

ABSTRACT
Senescence is one of the main barriers against tumor progression. Oncogenic signals in primary cells result in oncogene-induced senescence (OIS), crucial for protection against cancer development. It has been described in premalignant lesions that OIS requires DNA damage response (DDR) activation, safeguard of the integrity of the genome. Here we demonstrate how the cellular mechanisms involved in oncogenic transformation in a model of glioma uncouple OIS and DDR. We use this tumor type as a paradigm of oncogenic transformation. In human gliomas most of the genetic alterations that have been previously identified result in abnormal activation of cell growth signaling pathways and deregulation of cell cycle, features recapitulated in our model by oncogenic Ras expression and retinoblastoma (Rb) inactivation respectively. In this scenario, the absence of pRb confers a proliferative advantage and activates DDR to a greater extent in a DNA lesion-independent fashion than cells that express only HRas(V12). Moreover, Rb loss inactivates the stress kinase DDR-associated p38MAPK by specific Wip1-dependent dephosphorylation. Thus, Rb loss acts as a switch mediating the transition between premalignant lesions and cancer through DDR modulation. These findings may have important implications for the understanding the biology of gliomas and anticipate a new target, Wip1 phosphatase, for novel therapeutic strategies.

Show MeSH
Related in: MedlinePlus