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Down-regulation of CYLD expression by Snail promotes tumor progression in malignant melanoma.

Massoumi R, Kuphal S, Hellerbrand C, Haas B, Wild P, Spruss T, Pfeifer A, Fässler R, Bosserhoff AK - J. Exp. Med. (2009)

Bottom Line: Rescue of CYLD expression in melanoma cells reduced proliferation and invasion in vitro and tumor growth and metastasis in vivo.Importantly, tumor thickness and progression-free and overall survival inversely correlated with CYLD expression.Our data suggest that Snail1-mediated suppression of CYLD plays a key role in melanoma malignancy.

View Article: PubMed Central - PubMed

Affiliation: Department of Molecular Medicine, Max Planck Institute of Biochemistry, 82152 Martinsried, Germany.

ABSTRACT
High malignancy and early metastasis are hallmarks of melanoma. Here, we report that the transcription factor Snail1 inhibits expression of the tumor suppressor CYLD in melanoma. As a direct consequence of CYLD repression, the protooncogene BCL-3 translocates into the nucleus and activates Cyclin D1 and N-cadherin promoters, resulting in proliferation and invasion of melanoma cells. Rescue of CYLD expression in melanoma cells reduced proliferation and invasion in vitro and tumor growth and metastasis in vivo. Analysis of a tissue microarray with primary melanomas from patients revealed an inverse correlation of Snail1 induction and loss of CYLD expression. Importantly, tumor thickness and progression-free and overall survival inversely correlated with CYLD expression. Our data suggest that Snail1-mediated suppression of CYLD plays a key role in melanoma malignancy.

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Snail1 regulates tumorigenicity via CYLD repression. (A) Western blot analysis of two Mel Im asSnail clones (clone 1 and clone 2) applying CYLD, Cyclin D1, and N-cadherin antibodies. Stable transduction with viral vectors encoding siRNA against CYLD (siRNA CYLD) results in the complete depletion of CYLD, but induction of Cyclin D1 and N-cadherin in both clones. Transduction with control siRNA (siRNA control) exhibits no effect. Proliferation (B) and migration (C; in monolayer scratch assays) of Mel Im control cells (Control) and Mel Im asSnail clone 1 either stably transduced with viral vectors encoding siRNA against CYLD (siRNA CYLD), control siRNA (siRNA control), or without transduction (asSnail). Data represent proliferation after 72 h, and areas between scratch fronts (after 2 d in monolayer scratch assays) and are given as mean ± SEM. *, P < 0.05. Experiments in A–C were repeated at least three times. (D) Growth of Mel Im asSnail clone 1 (asSnail) transduced with siRNA against CYLD (siRNA CYLD) or control siRNA (siRNA control) and Mel Im control cells (Control) after s.c. implantation into nude mice (106 cells/mouse). Bars represent mean tumor size (± SEM) 3 wk after implantation. *, P < 0.05. (E) Histological analysis of the growths of Mel Im asSnail clone 1 without transduction (I) or stably transduced with siRNA against CYLD (II) 3 wk after s.c. implantation into nude mice. Pictures are representative and arrows indicate diffuse infiltration in II, whereas nodular growth (*) appears in I. Bar, 200 μm. Experiments and analysis in D and E, respectively, have been performed with 10 mice/group.
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fig3: Snail1 regulates tumorigenicity via CYLD repression. (A) Western blot analysis of two Mel Im asSnail clones (clone 1 and clone 2) applying CYLD, Cyclin D1, and N-cadherin antibodies. Stable transduction with viral vectors encoding siRNA against CYLD (siRNA CYLD) results in the complete depletion of CYLD, but induction of Cyclin D1 and N-cadherin in both clones. Transduction with control siRNA (siRNA control) exhibits no effect. Proliferation (B) and migration (C; in monolayer scratch assays) of Mel Im control cells (Control) and Mel Im asSnail clone 1 either stably transduced with viral vectors encoding siRNA against CYLD (siRNA CYLD), control siRNA (siRNA control), or without transduction (asSnail). Data represent proliferation after 72 h, and areas between scratch fronts (after 2 d in monolayer scratch assays) and are given as mean ± SEM. *, P < 0.05. Experiments in A–C were repeated at least three times. (D) Growth of Mel Im asSnail clone 1 (asSnail) transduced with siRNA against CYLD (siRNA CYLD) or control siRNA (siRNA control) and Mel Im control cells (Control) after s.c. implantation into nude mice (106 cells/mouse). Bars represent mean tumor size (± SEM) 3 wk after implantation. *, P < 0.05. (E) Histological analysis of the growths of Mel Im asSnail clone 1 without transduction (I) or stably transduced with siRNA against CYLD (II) 3 wk after s.c. implantation into nude mice. Pictures are representative and arrows indicate diffuse infiltration in II, whereas nodular growth (*) appears in I. Bar, 200 μm. Experiments and analysis in D and E, respectively, have been performed with 10 mice/group.

Mentions: Snail1 has been shown to play an important role in progression of melanoma. To study whether Snail1 facilitates tumorigenicity of melanoma cells via repression of CYLD, the two melanoma asSnail clones (Fig. 2 D) were stably transduced with viral vectors encoding siRNA against CYLD, resulting in complete suppression of CYLD (Fig. 3 A).


Down-regulation of CYLD expression by Snail promotes tumor progression in malignant melanoma.

Massoumi R, Kuphal S, Hellerbrand C, Haas B, Wild P, Spruss T, Pfeifer A, Fässler R, Bosserhoff AK - J. Exp. Med. (2009)

Snail1 regulates tumorigenicity via CYLD repression. (A) Western blot analysis of two Mel Im asSnail clones (clone 1 and clone 2) applying CYLD, Cyclin D1, and N-cadherin antibodies. Stable transduction with viral vectors encoding siRNA against CYLD (siRNA CYLD) results in the complete depletion of CYLD, but induction of Cyclin D1 and N-cadherin in both clones. Transduction with control siRNA (siRNA control) exhibits no effect. Proliferation (B) and migration (C; in monolayer scratch assays) of Mel Im control cells (Control) and Mel Im asSnail clone 1 either stably transduced with viral vectors encoding siRNA against CYLD (siRNA CYLD), control siRNA (siRNA control), or without transduction (asSnail). Data represent proliferation after 72 h, and areas between scratch fronts (after 2 d in monolayer scratch assays) and are given as mean ± SEM. *, P < 0.05. Experiments in A–C were repeated at least three times. (D) Growth of Mel Im asSnail clone 1 (asSnail) transduced with siRNA against CYLD (siRNA CYLD) or control siRNA (siRNA control) and Mel Im control cells (Control) after s.c. implantation into nude mice (106 cells/mouse). Bars represent mean tumor size (± SEM) 3 wk after implantation. *, P < 0.05. (E) Histological analysis of the growths of Mel Im asSnail clone 1 without transduction (I) or stably transduced with siRNA against CYLD (II) 3 wk after s.c. implantation into nude mice. Pictures are representative and arrows indicate diffuse infiltration in II, whereas nodular growth (*) appears in I. Bar, 200 μm. Experiments and analysis in D and E, respectively, have been performed with 10 mice/group.
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fig3: Snail1 regulates tumorigenicity via CYLD repression. (A) Western blot analysis of two Mel Im asSnail clones (clone 1 and clone 2) applying CYLD, Cyclin D1, and N-cadherin antibodies. Stable transduction with viral vectors encoding siRNA against CYLD (siRNA CYLD) results in the complete depletion of CYLD, but induction of Cyclin D1 and N-cadherin in both clones. Transduction with control siRNA (siRNA control) exhibits no effect. Proliferation (B) and migration (C; in monolayer scratch assays) of Mel Im control cells (Control) and Mel Im asSnail clone 1 either stably transduced with viral vectors encoding siRNA against CYLD (siRNA CYLD), control siRNA (siRNA control), or without transduction (asSnail). Data represent proliferation after 72 h, and areas between scratch fronts (after 2 d in monolayer scratch assays) and are given as mean ± SEM. *, P < 0.05. Experiments in A–C were repeated at least three times. (D) Growth of Mel Im asSnail clone 1 (asSnail) transduced with siRNA against CYLD (siRNA CYLD) or control siRNA (siRNA control) and Mel Im control cells (Control) after s.c. implantation into nude mice (106 cells/mouse). Bars represent mean tumor size (± SEM) 3 wk after implantation. *, P < 0.05. (E) Histological analysis of the growths of Mel Im asSnail clone 1 without transduction (I) or stably transduced with siRNA against CYLD (II) 3 wk after s.c. implantation into nude mice. Pictures are representative and arrows indicate diffuse infiltration in II, whereas nodular growth (*) appears in I. Bar, 200 μm. Experiments and analysis in D and E, respectively, have been performed with 10 mice/group.
Mentions: Snail1 has been shown to play an important role in progression of melanoma. To study whether Snail1 facilitates tumorigenicity of melanoma cells via repression of CYLD, the two melanoma asSnail clones (Fig. 2 D) were stably transduced with viral vectors encoding siRNA against CYLD, resulting in complete suppression of CYLD (Fig. 3 A).

Bottom Line: Rescue of CYLD expression in melanoma cells reduced proliferation and invasion in vitro and tumor growth and metastasis in vivo.Importantly, tumor thickness and progression-free and overall survival inversely correlated with CYLD expression.Our data suggest that Snail1-mediated suppression of CYLD plays a key role in melanoma malignancy.

View Article: PubMed Central - PubMed

Affiliation: Department of Molecular Medicine, Max Planck Institute of Biochemistry, 82152 Martinsried, Germany.

ABSTRACT
High malignancy and early metastasis are hallmarks of melanoma. Here, we report that the transcription factor Snail1 inhibits expression of the tumor suppressor CYLD in melanoma. As a direct consequence of CYLD repression, the protooncogene BCL-3 translocates into the nucleus and activates Cyclin D1 and N-cadherin promoters, resulting in proliferation and invasion of melanoma cells. Rescue of CYLD expression in melanoma cells reduced proliferation and invasion in vitro and tumor growth and metastasis in vivo. Analysis of a tissue microarray with primary melanomas from patients revealed an inverse correlation of Snail1 induction and loss of CYLD expression. Importantly, tumor thickness and progression-free and overall survival inversely correlated with CYLD expression. Our data suggest that Snail1-mediated suppression of CYLD plays a key role in melanoma malignancy.

Show MeSH
Related in: MedlinePlus