Stimulation of Melanogenesis in a Human Melanoma Cell Line by Bistratene A

Watters, Dianne Josephine; Garrone, Bernadette; Coomer, Joanne; Johnson, William E.; Brown, Geoffrey; Parsons, Peter G.

doi:10.1016/s0006-2952(97)00680-1

Cited by 26 publications

(23 citation statements)

References 46 publications

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“…As shown in Figure 1, PKC-d overexpression caused a significant 30% decrease in cell proliferation relative to empty vector (EV) transfected cells (Po0.05, n ¼ 6). We then treated cells with 12-Otetradecanoylphorbol 13-acetate (TPA), a phorbol ester, known to activate classical and novel PKC isoforms (Castagna et al, 1982;Ryves et al, 1991), or with Bistratene A (BisA), a PKC-d specific activator (Watters et al, 1998). Treatment of EV transfected Caco-2 cells with TPA or with BisA for 48 h, significantly inhibited cellular proliferation by 33 and 29%, respectively, compared with DMSO (vehicle treated) controls (Po0.05, n ¼ 6).…”

Section: Resultsmentioning

confidence: 99%

Protein kinase C delta inhibits Caco-2 cell proliferation by selective changes in cell cycle and cell death regulators

et al. 2006

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PKC-d is a serine/threonine kinase that mediates diverse signal transduction pathways. We previously demonstrated that overexpression of PKC-d slowed the G1 progression of Caco-2 colon cancer cells, accelerated apoptosis, and induced cellular differentiation. In this study, we further characterized the PKC-d dependent signaling pathways involved in these tumor suppressor actions in Caco-2 cells overexpressing PKC-d using a Zn 2 þ inducible expression vector. Consistent with a G1 arrest, increased expression of PKC-d caused rapid and significant downregulation of cyclin D1 and cyclin E proteins (50% decreases, Po0.05), while mRNA levels remained unchanged. The PKC agonist, phorbol 12-myristate 13-acetate (TPA, 100 nM, 4 h), induced two-fold higher protein and mRNA levels of p21 Waf1, a cyclin-dependent kinase (cdk) inhibitor in PKC-d transfectants compared with empty vector (EV) transfected cells, whereas the PKC-d specific inhibitor rottlerin (3 lM) or knockdown of this isoenzyme with specific siRNA oligonucleotides blocked p21Waf1 expression. Concomitantly, compared to EV control cells, PKC-d upregulation decreased cyclin D1 and cyclin E proteins co-immunoprecipitating with cdk6 and cdk2, respectively. In addition, overexpression of PKC-d increased binding of cdk inhibitor p27Kip1 to cdk4. These alterations in cyclin-cdks and their inhibitors are predicted to decrease G1 cyclin kinase activity. As an independent confirmation of the direct role PKC-d plays in cell growth and cell cycle regulation, we knocked down PKC-d using specific siRNA oligonucleotides. PKC-d specific siRNA oligonucleotides, but not irrelevant control oligonucleotides, inhibited PKCd protein by more than 80% in Caco-2 cells. Moreover, PKC-d knockdown enhanced cell proliferation (B1.4-2-fold, Po0.05) and concomitantly increased cyclin D1 and cyclin E expression (B1.7-fold, Po0.05). This was a specific effect, as nontargeted PKC-f was not changed by PKC-d siRNA oligonucleotides. Consistent with accelerated apoptosis in PKC-d transfectants, compared to EV cells, PKC-d upregulation increased proapoptotic regulator Bax two-fold at mRNA and protein levels, while antiapoptotic Bcl-2 protein was decreased by 50% at a post-transcriptional level. PKC-d specific siRNA oligonucleotides inhibited Bax protein expression by more than 50%, indicating that PKC-d regulates apoptosis through Bax. Taken together, these results elucidate two critical mechanisms regulated by PKC-d that inhibit cell cycle progression and enhance apoptosis in colon cancer cells. We postulate these antiproliferative pathways mediate an important tumor suppressor function for PKC-d in colonic carcinogenesis.

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Section: Resultsmentioning

confidence: 99%

Protein kinase C delta inhibits Caco-2 cell proliferation by selective changes in cell cycle and cell death regulators

et al. 2006

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“…Another skin response to UVR is an increase in melanin content to provide a further level of protection against UV-induced DNA damage and possibly oxidative damage (Thody and Graham, 1998). aMSH expression increases after UVR, and the binding of aMSH and tyrosinase activity are increased in normal G2 phase and G2 phase arrested melanoma cells, these e ects co-operate to increase the sensitivity of irradiated cells to aMSH and stimulate melanin synthesis (McLane and Pawelek, 1988;Watters et al, 1998;Chakraborty et al, 1999). The G2 arrest in the surrounding basal layer keratinocytes may also be a response to increased aMSH binding to these cells (Chakraborty and Pawelek, 1993) and contribute to the increased melanization of the basal layer.…”

Section: Discussionmentioning

confidence: 99%

G2 phase cell cycle arrest in human skin following UV irradiation

2001

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The contribution of the short wavelength ultraviolet (UV) component of sunlight to the aetiology of skin cancer has been widely acknowledged, although its direct contribution to tumour initiation or progression is still poorly understood. The loss of normal cell cycle controls, particularly checkpoint controls, are a common feature of cancer. UV radiation causes both G1 and G2 phase checkpoint arrest in vitro cultured cells. In this study we have investigated the cell cycle responses to suberythemal doses of UV on skin. We have utilized short-term whole organ skin cultures, and multi parameter immunohistochemical and biochemical analysis to demonstrate that basal and suprabasal layer melanocytes and keratinocytes undergo a G2 phase cell cycle arrest for up to 48 h following irradiation. The arrest is associated with increased p16 expression but no apparent p53 involvement. This type of organ culture provides a very useful model system, combining the ease of in vitro manipulation with the ability to perform detailed molecular analysis in a normal tissue environment. Oncogene (2001) 20, 6103 ± 6110.

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“…The growth arrest and alterations in collagen expression of bis A-treated chondrocytes may therefore represent part of a "default" response of these cells that is associated with inductions of cell death, rather than being part of any physiological, maturation pathway. Conversely, as PKCδ activation has also been associated with increased cell maturation in a number of systems (Denning et al, 2000;Watters and Parsons, 1999;Watters et al, 1998), the decreased viability of chondrocytes following bis A-treatment may occur independently or even as a consequence of this maturation. To some extent, the findings are thereby limited by the use of pharmacological agents, where pleiotropic effects may operate.…”

Section: Discussionmentioning

confidence: 99%

“…In fibroblasts, bis A treatment induces reorganisation of the actin cytoskeleton, a rounded morphology and detachment from tissue culture substrates , whilst in cultured melanoma cells treatment with bis A similarly alters cell morphology and induces onset of melanocyte differentiation (Watters et al, 1998). Here, we describe the effects of bis A on the shape, growth and phenotype of cultured human articular chondrocytes.…”

Section: Introductionmentioning

confidence: 95%

“…Bistratene A (bis A) is a macrolide polyether toxin that specifically activates PKCδ in a variety of cell types Griffiths et al, 1996;Watters et al, 1998). In fibroblasts, bis A treatment induces reorganisation of the actin cytoskeleton, a rounded morphology and detachment from tissue culture substrates , whilst in cultured melanoma cells treatment with bis A similarly alters cell morphology and induces onset of melanocyte differentiation (Watters et al, 1998).…”

Section: Introductionmentioning

confidence: 99%

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Phenotypic modulation of human articular chondrocytes by bistratene A

Cragg²,

Richardson

et al. 2002

eCM

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Chondrocytes undergo phenotypic alterations following extended periods in monolayer culture, i.e., they become bipolar and flattened, proliferate, and synthesise type I as opposed to type II collagen. This process has been termed chondrocyte dedifferentiation. Bistratene A is a macrolide polyether that specifically activates the delta isoform of protein kinase C (PKCδ) in some cell types. Here, we show that dedifferentiated human articular chondrocytes became rounded and underwent cell growth arrest after treatment with bistratene A. In addition, bistratene Atreated chondrocytes became more immunopositive for type II collagen, but less immunopositive for type I collagen. These phenotypic changes were associated with a prior and extensive disruption of actin microfilaments and translocation of PKCδ to the nuclear membrane. Concurrent treatments of chondrocytes with a specific inhibitor of PKCδ, rottlerin, partially blocked the morphological effects of bistratene A.

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Stimulation of Melanogenesis in a Human Melanoma Cell Line by Bistratene A

Cited by 26 publications

References 46 publications

Protein kinase C delta inhibits Caco-2 cell proliferation by selective changes in cell cycle and cell death regulators

Protein kinase C delta inhibits Caco-2 cell proliferation by selective changes in cell cycle and cell death regulators

G2 phase cell cycle arrest in human skin following UV irradiation

Phenotypic modulation of human articular chondrocytes by bistratene A

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