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SKP2 cooperates with N-Ras or AKT to induce liver tumor development in mice.

Delogu S, Wang C, Cigliano A, Utpatel K, Sini M, Longerich T, Waldburger N, Breuhahn K, Jiang L, Ribback S, Dombrowski F, Evert M, Chen X, Calvisi DF - Oncotarget (2015)

Bottom Line: We found that forced overexpression of SKP2, N-RasV12 or ΔN90-β-catenin alone as well as co-expression of SKP2 and ΔN90-β-catenin did not induce liver tumor development.In human HCC specimens, nuclear translocation of SKP2 was associated with activation of the AKT/mTOR and Ras/MAPK pathways, but not with β-catenin mutation or activation.Altogether, the present data indicate that SKP2 cooperates with N-Ras and AKT proto-oncogenes to promote hepatocarcinogenesis in vivo.

View Article: PubMed Central - PubMed

Affiliation: Institut für Pathologie, Universitätsmedizin Greifswald, Greifswald, Germany.

ABSTRACT
Mounting evidence indicates that S-Phase Kinase-Associated Protein 2 (SKP2) is overexpressed in human hepatocellular carcinoma (HCC). However, the role of SKP2 in hepatocarcinogenesis remains poorly delineated. To elucidate the function(s) of SKP2 in HCC, we stably overexpressed the SKP2 gene in the mouse liver, either alone or in combination with activated forms of N-Ras (N-RasV12), AKT1 (myr-AKT1), or β-catenin (ΔN90-β-catenin) protooncogenes, via hydrodynamic gene delivery. We found that forced overexpression of SKP2, N-RasV12 or ΔN90-β-catenin alone as well as co-expression of SKP2 and ΔN90-β-catenin did not induce liver tumor development. Overexpression of myr-AKT1 alone led to liver tumor development after long latency. In contrast, co-expression of SKP2 with N-RasV12 or myr-AKT1 resulted in early development of multiple hepatocellular tumors in all SKP2/N-RasV12 and SKP2/myr-AKT1 mice. At the molecular level, preneoplastic and neoplastic liver lesions from SKP2/N-RasV12 and SKP2/myr-AKT1 mice exhibited a strong induction of AKT/mTOR and Ras/MAPK pathways. Noticeably, the tumor suppressor proteins whose levels have been shown to be downregulated by SKP2-dependent degradation in various tumor types, including p27, p57, Dusp1, and Rassf1A were not decreased in liver lesions from SKP2/N-RasV12 and SKP2/myr-AKT1 mice. In human HCC specimens, nuclear translocation of SKP2 was associated with activation of the AKT/mTOR and Ras/MAPK pathways, but not with β-catenin mutation or activation. Altogether, the present data indicate that SKP2 cooperates with N-Ras and AKT proto-oncogenes to promote hepatocarcinogenesis in vivo.

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Immunohistochemical patterns of SKP2, activated/phosphorylated AKT (p-AKT), activated/phosphorylated ERK1/2 (p-ERK1/2), and β-catenin proteins in human hepatocellular carcinoma (HCC)Upper panel: HCC specimen showing strong immunolabeling for nuclear SKP2, p-AKT, and p-ERK1/2 and membranous β-catenin (better appreciable in the inset). Lower panel: HCC specimen exhibiting strong immunoreactivity for nuclear SKP2, p-AKT, p-ERK1/2, and nuclear β-catenin. Abbreviation: HE, hematoxylin and eosin staining. Original magnification: 400x in inset; 200x in all the other pictures.
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Figure 6: Immunohistochemical patterns of SKP2, activated/phosphorylated AKT (p-AKT), activated/phosphorylated ERK1/2 (p-ERK1/2), and β-catenin proteins in human hepatocellular carcinoma (HCC)Upper panel: HCC specimen showing strong immunolabeling for nuclear SKP2, p-AKT, and p-ERK1/2 and membranous β-catenin (better appreciable in the inset). Lower panel: HCC specimen exhibiting strong immunoreactivity for nuclear SKP2, p-AKT, p-ERK1/2, and nuclear β-catenin. Abbreviation: HE, hematoxylin and eosin staining. Original magnification: 400x in inset; 200x in all the other pictures.

Mentions: To further investigate the possible relationship between SKP2 and the AKT/mTOR, Ras/MAPK, and Wnt/β-catenin pathways, we analyzed a collection of human HCC specimens (n = 64; Supplementary Table 3) by immunohistochemistry for SKP2, p-AKT, p-ERK, and β-catenin staining (Fig. 6). SKP2 nuclear accumulation was detected in 18 of 64 HCC (28.1%), whereas positive immunolabeling for p-AKT, p-ERK, and nuclear β-catenin was found in 54.7%, 32.8%, and 29.7% of HCC, respectively. β-catenin mutations were detected in 12 of 64 HCCs (18.8%). Of note, 83.3% (15/18) and 66.6% (12/18) of HCCs displaying nuclear accumulation of SKP2 concomitantly showed up-regulation of p-AKT and p-ERK, respectively. In contrast, only 16.7% (3/18) of HCC specimens with nuclear SKP2 concomitantly exhibited nuclear accumulation of β-catenin (P < 0.001 and P < 0.01 vs. p-AKT and p-ERK, respectively). Of the samples showing simultaneous nuclear SKP2 and β-catenin immunoreactivity, two harbored β-catenin mutations. Importantly, 11 of 13 HCCs showing concomitantly nuclear SKP2 accumulation and p-ERK and p-AKT activation belonged to the HCC subset with poorer outcome, suggesting that simultaneous activation of the latter cascades is associated with a dismal prognosis in HCC. No association between the staining patterns of SKP2, p-AKT, and p-ERK and other clinicopathologic features of the patients, including etiology, presence of cirrhosis, α-fetoprotein levels, and tumor grading was found. Subsequent analysis of an additional collection of human HCC (n = 44) whose survival data were missing, showed nuclear accumulation of SKP2 in 11 of 44 (25%) samples, and activation of p-AKT, p-ERK, and β-catenin in 59.1%, 31.9%, and 36,4%, respectively. β-catenin mutations were detected in 11 of 44 HCCs (25%). Once again, nuclear accumulation of SKP2 was frequently paralleled by p-AKT and p-ERK activation, as eight of 11 HCC exhibited concomitantly nuclear SKP2 and p-AKT and p-ERK immunoreactivity, whereas only two HCC displayed concomitant nuclear SKP2 and β-catenin accumulation (P < 0.05). Altogether, the present data indicate that HCC with nuclear SKP2 translocation are often characterized by activation of the AKT/mTOR and Ras/MAPK pathways, with β-catenin mutations and/or activation rarely occurring in this HCC subset.


SKP2 cooperates with N-Ras or AKT to induce liver tumor development in mice.

Delogu S, Wang C, Cigliano A, Utpatel K, Sini M, Longerich T, Waldburger N, Breuhahn K, Jiang L, Ribback S, Dombrowski F, Evert M, Chen X, Calvisi DF - Oncotarget (2015)

Immunohistochemical patterns of SKP2, activated/phosphorylated AKT (p-AKT), activated/phosphorylated ERK1/2 (p-ERK1/2), and β-catenin proteins in human hepatocellular carcinoma (HCC)Upper panel: HCC specimen showing strong immunolabeling for nuclear SKP2, p-AKT, and p-ERK1/2 and membranous β-catenin (better appreciable in the inset). Lower panel: HCC specimen exhibiting strong immunoreactivity for nuclear SKP2, p-AKT, p-ERK1/2, and nuclear β-catenin. Abbreviation: HE, hematoxylin and eosin staining. Original magnification: 400x in inset; 200x in all the other pictures.
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Related In: Results  -  Collection

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getmorefigures.php?uid=PMC4385847&req=5

Figure 6: Immunohistochemical patterns of SKP2, activated/phosphorylated AKT (p-AKT), activated/phosphorylated ERK1/2 (p-ERK1/2), and β-catenin proteins in human hepatocellular carcinoma (HCC)Upper panel: HCC specimen showing strong immunolabeling for nuclear SKP2, p-AKT, and p-ERK1/2 and membranous β-catenin (better appreciable in the inset). Lower panel: HCC specimen exhibiting strong immunoreactivity for nuclear SKP2, p-AKT, p-ERK1/2, and nuclear β-catenin. Abbreviation: HE, hematoxylin and eosin staining. Original magnification: 400x in inset; 200x in all the other pictures.
Mentions: To further investigate the possible relationship between SKP2 and the AKT/mTOR, Ras/MAPK, and Wnt/β-catenin pathways, we analyzed a collection of human HCC specimens (n = 64; Supplementary Table 3) by immunohistochemistry for SKP2, p-AKT, p-ERK, and β-catenin staining (Fig. 6). SKP2 nuclear accumulation was detected in 18 of 64 HCC (28.1%), whereas positive immunolabeling for p-AKT, p-ERK, and nuclear β-catenin was found in 54.7%, 32.8%, and 29.7% of HCC, respectively. β-catenin mutations were detected in 12 of 64 HCCs (18.8%). Of note, 83.3% (15/18) and 66.6% (12/18) of HCCs displaying nuclear accumulation of SKP2 concomitantly showed up-regulation of p-AKT and p-ERK, respectively. In contrast, only 16.7% (3/18) of HCC specimens with nuclear SKP2 concomitantly exhibited nuclear accumulation of β-catenin (P < 0.001 and P < 0.01 vs. p-AKT and p-ERK, respectively). Of the samples showing simultaneous nuclear SKP2 and β-catenin immunoreactivity, two harbored β-catenin mutations. Importantly, 11 of 13 HCCs showing concomitantly nuclear SKP2 accumulation and p-ERK and p-AKT activation belonged to the HCC subset with poorer outcome, suggesting that simultaneous activation of the latter cascades is associated with a dismal prognosis in HCC. No association between the staining patterns of SKP2, p-AKT, and p-ERK and other clinicopathologic features of the patients, including etiology, presence of cirrhosis, α-fetoprotein levels, and tumor grading was found. Subsequent analysis of an additional collection of human HCC (n = 44) whose survival data were missing, showed nuclear accumulation of SKP2 in 11 of 44 (25%) samples, and activation of p-AKT, p-ERK, and β-catenin in 59.1%, 31.9%, and 36,4%, respectively. β-catenin mutations were detected in 11 of 44 HCCs (25%). Once again, nuclear accumulation of SKP2 was frequently paralleled by p-AKT and p-ERK activation, as eight of 11 HCC exhibited concomitantly nuclear SKP2 and p-AKT and p-ERK immunoreactivity, whereas only two HCC displayed concomitant nuclear SKP2 and β-catenin accumulation (P < 0.05). Altogether, the present data indicate that HCC with nuclear SKP2 translocation are often characterized by activation of the AKT/mTOR and Ras/MAPK pathways, with β-catenin mutations and/or activation rarely occurring in this HCC subset.

Bottom Line: We found that forced overexpression of SKP2, N-RasV12 or ΔN90-β-catenin alone as well as co-expression of SKP2 and ΔN90-β-catenin did not induce liver tumor development.In human HCC specimens, nuclear translocation of SKP2 was associated with activation of the AKT/mTOR and Ras/MAPK pathways, but not with β-catenin mutation or activation.Altogether, the present data indicate that SKP2 cooperates with N-Ras and AKT proto-oncogenes to promote hepatocarcinogenesis in vivo.

View Article: PubMed Central - PubMed

Affiliation: Institut für Pathologie, Universitätsmedizin Greifswald, Greifswald, Germany.

ABSTRACT
Mounting evidence indicates that S-Phase Kinase-Associated Protein 2 (SKP2) is overexpressed in human hepatocellular carcinoma (HCC). However, the role of SKP2 in hepatocarcinogenesis remains poorly delineated. To elucidate the function(s) of SKP2 in HCC, we stably overexpressed the SKP2 gene in the mouse liver, either alone or in combination with activated forms of N-Ras (N-RasV12), AKT1 (myr-AKT1), or β-catenin (ΔN90-β-catenin) protooncogenes, via hydrodynamic gene delivery. We found that forced overexpression of SKP2, N-RasV12 or ΔN90-β-catenin alone as well as co-expression of SKP2 and ΔN90-β-catenin did not induce liver tumor development. Overexpression of myr-AKT1 alone led to liver tumor development after long latency. In contrast, co-expression of SKP2 with N-RasV12 or myr-AKT1 resulted in early development of multiple hepatocellular tumors in all SKP2/N-RasV12 and SKP2/myr-AKT1 mice. At the molecular level, preneoplastic and neoplastic liver lesions from SKP2/N-RasV12 and SKP2/myr-AKT1 mice exhibited a strong induction of AKT/mTOR and Ras/MAPK pathways. Noticeably, the tumor suppressor proteins whose levels have been shown to be downregulated by SKP2-dependent degradation in various tumor types, including p27, p57, Dusp1, and Rassf1A were not decreased in liver lesions from SKP2/N-RasV12 and SKP2/myr-AKT1 mice. In human HCC specimens, nuclear translocation of SKP2 was associated with activation of the AKT/mTOR and Ras/MAPK pathways, but not with β-catenin mutation or activation. Altogether, the present data indicate that SKP2 cooperates with N-Ras and AKT proto-oncogenes to promote hepatocarcinogenesis in vivo.

Show MeSH
Related in: MedlinePlus