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Interactions between cells with distinct mutations in c-MYC and Pten in prostate cancer.

Kim J, Eltoum IE, Roh M, Wang J, Abdulkadir SA - PLoS Genet. (2009)

Bottom Line: These lesions were in all cases associated with loss of Pten protein expression from the wild type allele.We conclude that c-MYC overexpression and Pten deficiency cooperate to promote prostate tumorigenesis, but a p53-dependent apoptotic response may present a barrier to further progression.Our results highlight the utility of inducing mutations focally to model the competitive interactions between cell populations with distinct genetic alterations during tumorigenesis.

View Article: PubMed Central - PubMed

Affiliation: Department of Pathology, Vanderbilt University Medical Center, Nashville, TN, USA.

ABSTRACT
In human somatic tumorigenesis, mutations are thought to arise sporadically in individual cells surrounded by unaffected cells. This contrasts with most current transgenic models where mutations are induced synchronously in entire cell populations. Here we have modeled sporadic oncogene activation using a transgenic mouse in which c-MYC is focally activated in prostate luminal epithelial cells. Focal c-MYC expression resulted in mild pathology, but prostate-specific deletion of a single allele of the Pten tumor suppressor gene cooperated with c-MYC to induce high grade prostatic intraepithelial neoplasia (HGPIN)/cancer lesions. These lesions were in all cases associated with loss of Pten protein expression from the wild type allele. In the prostates of mice with concurrent homozygous deletion of Pten and focal c-MYC activation, double mutant (i.e. c-MYC+;Pten-) cells were of higher grade and proliferated faster than single mutant (Pten-) cells within the same glands. Consequently, double mutant cells outcompeted single mutant cells despite the presence of increased rates of apoptosis in the former. The p53 pathway was activated in Pten-deficient prostate cells and tissues, but c-MYC expression shifted the p53 response from senescence to apoptosis by repressing the p53 target gene p21(Cip1). We conclude that c-MYC overexpression and Pten deficiency cooperate to promote prostate tumorigenesis, but a p53-dependent apoptotic response may present a barrier to further progression. Our results highlight the utility of inducing mutations focally to model the competitive interactions between cell populations with distinct genetic alterations during tumorigenesis.

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c-MYC expression increases the proliferation and tumorigenicity of Pten-deficient cells.(A) Proliferation was determined by analysis of phospho-histone H3 staining. (B) Phospho-histone H3 index in c-MYC-positive or c-MYC-negative cells in PbCre4;Z-MYC;Ptenf/f prostates. Inset: double staining shows colocalization of phospho-histone H3 with c-MYC in PbCre4;Z-MYC;Ptenf/f prostate. N = 3–4 mice per group. *p<0.05, **p<0.005, ****p<0.01. (C) c-MYC staining identifies MYC-expressing cells next to MYC-negative cells in the same gland of PbCre4;Z-MYC;Ptenf/f mouse prostate. An adjacent H&E-stained section is also shown. Note distinct, higher grade pathology of c-MYC+ cells. Scale bar: 50 µm. (D) c-MYC+;Pten- cells outcompete Pten- cells. Prostates from 10-week-old and 28-week-old PbCre4;Z-MYC;Ptenf/f mice were stained for c-MYC and smooth muscle actin. At 10 weeks, c-MYC expression is focal; at 28 weeks, it is uniform. Arrows indicate discontinuity of smooth muscle actin (focal micro-invasion). Scale bar: 100 µm.
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pgen-1000542-g005: c-MYC expression increases the proliferation and tumorigenicity of Pten-deficient cells.(A) Proliferation was determined by analysis of phospho-histone H3 staining. (B) Phospho-histone H3 index in c-MYC-positive or c-MYC-negative cells in PbCre4;Z-MYC;Ptenf/f prostates. Inset: double staining shows colocalization of phospho-histone H3 with c-MYC in PbCre4;Z-MYC;Ptenf/f prostate. N = 3–4 mice per group. *p<0.05, **p<0.005, ****p<0.01. (C) c-MYC staining identifies MYC-expressing cells next to MYC-negative cells in the same gland of PbCre4;Z-MYC;Ptenf/f mouse prostate. An adjacent H&E-stained section is also shown. Note distinct, higher grade pathology of c-MYC+ cells. Scale bar: 50 µm. (D) c-MYC+;Pten- cells outcompete Pten- cells. Prostates from 10-week-old and 28-week-old PbCre4;Z-MYC;Ptenf/f mice were stained for c-MYC and smooth muscle actin. At 10 weeks, c-MYC expression is focal; at 28 weeks, it is uniform. Arrows indicate discontinuity of smooth muscle actin (focal micro-invasion). Scale bar: 100 µm.

Mentions: We analyzed proliferation by staining for phospho-histone H3 (pHH3), a mitotic marker. Proliferation was increased significantly in PbCre4;Z-MYC prostates relative to controls, and Pten heterozygosity synergistically increased it further (Figure 5A). The proliferation rates in PbCre4;Ptenf/f and PbCre4;Z-MYC;Ptenf/f were similarly elevated. However, the focal nature of c-MYC expression in our model means that analysis of total proliferation in the PbCre4;Z-MYC;Ptenf/f prostates may not be an accurate measure of the proliferation in foci of c-MYC+;Pten- cells. To overcome this, we performed double staining for c-MYC and phospho-histone H3. As shown in Figure 5B, double mutant (c-MYC+;Pten-) cells were more proliferative than single mutant (Pten-) cells within the same prostate glands. Furthermore, double mutant cells are histologically distinct from adjacent single mutant cells. The double mutant cells are of higher pathological grade with larger nuclei, high nuclear∶cytoplasmic ratios, hyperchromatic nuclei with prominent chromocenters, focal chromatin clearing and prominent single or sometimes multiple nucleoli (Figure 5C and Figure S1). In addition, apoptotic and mitotic figures are prominent. Single mutant (Pten-) cells on the other hand showed low nuclear grade with comparatively small and uniform nuclei, abundant pale cytoplasm and low nuclear∶cytoplasmic ratios. These cells also have inconspicuous nucleoli and the chromatin is comparatively fine (Figure 5C). These observations suggest that c-MYC+;Pten- cells may out-compete Pten- cells within the same prostate gland over time. Indeed, analysis of PbCre4;Z-MYC;Ptenf/f animals showed that at early ages, c-MYC expression was focal within glands, but in older mice, lesions show uniform c-MYC expression, suggesting clonal expansion of c-MYC-positive cells in a time-dependent manner (Figure 5D).


Interactions between cells with distinct mutations in c-MYC and Pten in prostate cancer.

Kim J, Eltoum IE, Roh M, Wang J, Abdulkadir SA - PLoS Genet. (2009)

c-MYC expression increases the proliferation and tumorigenicity of Pten-deficient cells.(A) Proliferation was determined by analysis of phospho-histone H3 staining. (B) Phospho-histone H3 index in c-MYC-positive or c-MYC-negative cells in PbCre4;Z-MYC;Ptenf/f prostates. Inset: double staining shows colocalization of phospho-histone H3 with c-MYC in PbCre4;Z-MYC;Ptenf/f prostate. N = 3–4 mice per group. *p<0.05, **p<0.005, ****p<0.01. (C) c-MYC staining identifies MYC-expressing cells next to MYC-negative cells in the same gland of PbCre4;Z-MYC;Ptenf/f mouse prostate. An adjacent H&E-stained section is also shown. Note distinct, higher grade pathology of c-MYC+ cells. Scale bar: 50 µm. (D) c-MYC+;Pten- cells outcompete Pten- cells. Prostates from 10-week-old and 28-week-old PbCre4;Z-MYC;Ptenf/f mice were stained for c-MYC and smooth muscle actin. At 10 weeks, c-MYC expression is focal; at 28 weeks, it is uniform. Arrows indicate discontinuity of smooth muscle actin (focal micro-invasion). Scale bar: 100 µm.
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Related In: Results  -  Collection

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getmorefigures.php?uid=PMC2697385&req=5

pgen-1000542-g005: c-MYC expression increases the proliferation and tumorigenicity of Pten-deficient cells.(A) Proliferation was determined by analysis of phospho-histone H3 staining. (B) Phospho-histone H3 index in c-MYC-positive or c-MYC-negative cells in PbCre4;Z-MYC;Ptenf/f prostates. Inset: double staining shows colocalization of phospho-histone H3 with c-MYC in PbCre4;Z-MYC;Ptenf/f prostate. N = 3–4 mice per group. *p<0.05, **p<0.005, ****p<0.01. (C) c-MYC staining identifies MYC-expressing cells next to MYC-negative cells in the same gland of PbCre4;Z-MYC;Ptenf/f mouse prostate. An adjacent H&E-stained section is also shown. Note distinct, higher grade pathology of c-MYC+ cells. Scale bar: 50 µm. (D) c-MYC+;Pten- cells outcompete Pten- cells. Prostates from 10-week-old and 28-week-old PbCre4;Z-MYC;Ptenf/f mice were stained for c-MYC and smooth muscle actin. At 10 weeks, c-MYC expression is focal; at 28 weeks, it is uniform. Arrows indicate discontinuity of smooth muscle actin (focal micro-invasion). Scale bar: 100 µm.
Mentions: We analyzed proliferation by staining for phospho-histone H3 (pHH3), a mitotic marker. Proliferation was increased significantly in PbCre4;Z-MYC prostates relative to controls, and Pten heterozygosity synergistically increased it further (Figure 5A). The proliferation rates in PbCre4;Ptenf/f and PbCre4;Z-MYC;Ptenf/f were similarly elevated. However, the focal nature of c-MYC expression in our model means that analysis of total proliferation in the PbCre4;Z-MYC;Ptenf/f prostates may not be an accurate measure of the proliferation in foci of c-MYC+;Pten- cells. To overcome this, we performed double staining for c-MYC and phospho-histone H3. As shown in Figure 5B, double mutant (c-MYC+;Pten-) cells were more proliferative than single mutant (Pten-) cells within the same prostate glands. Furthermore, double mutant cells are histologically distinct from adjacent single mutant cells. The double mutant cells are of higher pathological grade with larger nuclei, high nuclear∶cytoplasmic ratios, hyperchromatic nuclei with prominent chromocenters, focal chromatin clearing and prominent single or sometimes multiple nucleoli (Figure 5C and Figure S1). In addition, apoptotic and mitotic figures are prominent. Single mutant (Pten-) cells on the other hand showed low nuclear grade with comparatively small and uniform nuclei, abundant pale cytoplasm and low nuclear∶cytoplasmic ratios. These cells also have inconspicuous nucleoli and the chromatin is comparatively fine (Figure 5C). These observations suggest that c-MYC+;Pten- cells may out-compete Pten- cells within the same prostate gland over time. Indeed, analysis of PbCre4;Z-MYC;Ptenf/f animals showed that at early ages, c-MYC expression was focal within glands, but in older mice, lesions show uniform c-MYC expression, suggesting clonal expansion of c-MYC-positive cells in a time-dependent manner (Figure 5D).

Bottom Line: These lesions were in all cases associated with loss of Pten protein expression from the wild type allele.We conclude that c-MYC overexpression and Pten deficiency cooperate to promote prostate tumorigenesis, but a p53-dependent apoptotic response may present a barrier to further progression.Our results highlight the utility of inducing mutations focally to model the competitive interactions between cell populations with distinct genetic alterations during tumorigenesis.

View Article: PubMed Central - PubMed

Affiliation: Department of Pathology, Vanderbilt University Medical Center, Nashville, TN, USA.

ABSTRACT
In human somatic tumorigenesis, mutations are thought to arise sporadically in individual cells surrounded by unaffected cells. This contrasts with most current transgenic models where mutations are induced synchronously in entire cell populations. Here we have modeled sporadic oncogene activation using a transgenic mouse in which c-MYC is focally activated in prostate luminal epithelial cells. Focal c-MYC expression resulted in mild pathology, but prostate-specific deletion of a single allele of the Pten tumor suppressor gene cooperated with c-MYC to induce high grade prostatic intraepithelial neoplasia (HGPIN)/cancer lesions. These lesions were in all cases associated with loss of Pten protein expression from the wild type allele. In the prostates of mice with concurrent homozygous deletion of Pten and focal c-MYC activation, double mutant (i.e. c-MYC+;Pten-) cells were of higher grade and proliferated faster than single mutant (Pten-) cells within the same glands. Consequently, double mutant cells outcompeted single mutant cells despite the presence of increased rates of apoptosis in the former. The p53 pathway was activated in Pten-deficient prostate cells and tissues, but c-MYC expression shifted the p53 response from senescence to apoptosis by repressing the p53 target gene p21(Cip1). We conclude that c-MYC overexpression and Pten deficiency cooperate to promote prostate tumorigenesis, but a p53-dependent apoptotic response may present a barrier to further progression. Our results highlight the utility of inducing mutations focally to model the competitive interactions between cell populations with distinct genetic alterations during tumorigenesis.

Show MeSH
Related in: MedlinePlus