Effects of Cholera Toxin on Proliferation of Cultured Human Keratinocytes in Relation to Intracellular Cyclic AMP Levels

Okada, Natsuko; Kitano, Yukio; Ichihara, K.

doi:10.1111/1523-1747.ep12510580

Cited by 52 publications

(24 citation statements)

References 19 publications

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“…Fig. 2 a indicates that several keratins of ME-180 cells exhibit isoelectric heterogeneity similar to that described for keratin subunits from other tissues (23)(24)(25)(26) . Immunoblot analysis (Fig.…”

Section: Resultssupporting

confidence: 63%

Dual regulation of intermediate filament phosphorylation.

Gilmartin¹,

Mitchell²,

Vidrich³

et al. 1984

The Journal of Cell Biology

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Intermediate filament proteins have been isolated from ME-180, cells of a human cervical carcinoma. Eight of these proteins have been identified as keratins by immunologic cross-reactivity to antibodies raised against authentic human epidermal keratins. The ME-180 keratin proteins consist of two major subunits designated MEK-1 and MEK-2 with approximate molecular weights of 58,000 and 53,000, respectively, and six minor subunits of 59, 57, 52.5, 50.5, 45, and 40 kilodaltons. When ME-180 cells were incubated for 2-24 h in the presence of [32P]orthophosphate, MEK-1 and MEK-2 as well as the 52.5- and 40-kilodalton keratins were phosphorylated at their serine residues. V8 protease digests revealed that phosphorylation of MEK-2 is restricted to one peptide representing approximately half the molecule. Regulation of MEK-1 and MEK-2 phosphorylation has been studied by prelabeling the cells for 2 h in 32P-labeled medium. This was followed by up to 2 h of continued incubation in the same medium after the addition of a variety of perturbing agents. The phosphorylation of MEK-2 increased in the presence of 10(-4) M dibutyryl cyclic AMP (twofold), 1 mM methylisobutylxanthine (2.5-fold), 10(-5) M isoproterenol (fivefold), and 10(-9) M cholera toxin (sevenfold). In contrast, MEK-1 phosphorylation was unaffected by these agents. Neither cyclic GMP, Ca++, hydrocortisone, nor epidermal growth factor had any effect on the phosphorylation of MEK-1 or MEK-2. The results indicate that the phosphorylation of these two keratins is independently controlled by cyclic AMP-dependent kinase for MEK-2 and by cyclic nucleotide-independent kinase for MEK-1. The observed differences in control suggest distinct functions for MEK-1 and MEK-2 within the cytoskeletal network.

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Section: Resultssupporting

confidence: 63%

Dual regulation of intermediate filament phosphorylation.

Gilmartin¹,

Mitchell²,

Vidrich³

et al. 1984

The Journal of Cell Biology

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“…Incorporation of 3H-thymidine in control cultures is a function of XB cell density, with the maximum occurring at a plating density of 2.3 x lo4 cells/ cm2 and a gradual decrease being seen with higher plating densities. Cholera toxin produced the greatest effect at the lowest XB cell density tested (0.47 x lo4 cells/cm2), consistent with the findings of Okada et al (1982) with cultured human keratinocytes. The maximal effect of EGF occurred in cultures plated at very high cell densities, while TPA produced a moderate increase in thymidine incorporation at both low and high cell densities.…”

Section: Resultssupporting

confidence: 85%

2,3,7,8‐Tetrachlorodibenzo‐p‐dioxin: Examination of biochemical effects involved in the proliferation and differentiation of XB cells

Knutson

Poland

1984

Journal Cellular Physiology

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XB, a cell line derived from a mouse teratoma, differentiates into stratified squamous epithelium when incubated with 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD). To examine the possible biochemical mediators of this response, we compared the effects produced by TCDD to those elicited by other compounds which stimulate epidermal proliferation and/or differentiation in mice. XB/3T3 cultures keratinize when incubated with cholera toxin, epidermal growth factor (EGF), or TCDD, but not 12-0-tetradecanoylphorbol-13-acetate (TPA). Incubation of XB cells with TCDD (10(-9)M) for 48 hours produces a 20% increase in thymidine incorporation, a response which is neither as large nor as rapid as that produced by cholera toxin, TPA, or EGF. Although both cholera toxin and TCDD stimulate differentiation and thymidine incorporation in XB/3T3 cultures, cholera toxin increases cAMP 30-fold in these cells, while TCDD does not affect cAMP accumulation at any of the times studies (15 min to 120 hours). Inhibitors of arachidonic acid metabolism, which block epidermal proliferative responses to TPA in vivo, do not prevent the differentiation of XB cells in response to TCDD. In XB/3T3 cultures, TPA stimulates arachidonic acid release at all times tested (1,6, and 24 hours) and increases the incorporation of 32Pi into total phospholipids and phosphatidylcholine after 3 hours. In contrast, TCDD affects neither arachidonic acid release nor the turnover of phosphatidylinositol or phosphatidylcholine at any of the times tested. Although we examined biochemical effects which have been suggested as part of the mechanism of TCDD and which are produced by other epidermal proliferative compounds in XB cells, no mediator of the TCDD-produced differentiation of XB/3T3 cultures was observed.

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“…The factors that control terminal differentiation in the keratinocytes are not completely understood; however, a role of cyclic adenosine monophosphate has been implicated. Cyclic adenosine monophosphate (cAMP) has been shown to either inhibit keratinocyte cell division [3][4][5] or promote cell division [6][7][8] depending on cell culture conditions. However, the effects of cAMP levels on keratinocyte differentation have been largely overlooked because of conflicting observations on cell division.…”

Section: Introductionmentioning

confidence: 99%

The Induction of Terminal Differentiation Markers by the cAMP Pathway in Human HaCaT Keratinocytes

Mammone

Marenus

Maes

et al. 1998

Skin Pharmacol Physiol

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The terminal differentiation of human epidermal keratinocytes is a complex morphological and biochemical shift from a mitotically active cell to an inert protein cross-linked envelope. This transition is a clearly predetermined cell death mechanism, but it is unlike many other programmed cell deaths in that it is not apoptotic. To explore and contrast the mechanism by which keratinocytes are committed to differentiation rather than apoptosis, we focused on the cyclic adenosine monophosphate (cAMP) signaling pathway using selective modulators of intracelluar cAMP levels. Markers of differentiation were assayed by Western blotting. Raising intracelluar cAMP levels by treating HaCaT cells with forskolin, a diterpene, or with isobutylmethylxanthine, a phosphodiesterase inhibitor, and isoproterenol, a β-adrenergic receptor agonist that selectively activates adenylate cyclase, increased the levels of the differentiation markers keratin K1 and K10, involucrin and transglutaminase. H89 and KT5720, both inhibitors of cAMP-dependent protein kinase, suppressed the expression of keratins K1 and K10. These observations are in line with the defined role for cAMP in the control of keratinocyte differentiation.

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Effects of Cholera Toxin on Proliferation of Cultured Human Keratinocytes in Relation to Intracellular Cyclic AMP Levels

Cited by 52 publications

References 19 publications

Dual regulation of intermediate filament phosphorylation.

Dual regulation of intermediate filament phosphorylation.

2,3,7,8‐Tetrachlorodibenzo‐p‐dioxin: Examination of biochemical effects involved in the proliferation and differentiation of XB cells

The Induction of Terminal Differentiation Markers by the cAMP Pathway in Human HaCaT Keratinocytes

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