Characterisation of protein kinase C isoforms and enzymic activity from the αT3‐1 gonadotroph‐derived cell line

Johnson, Mark; MacEwan, David J.; Simpson, J. A.; Mitchell, Rory

doi:10.1016/0014-5793(93)80376-6

Cited by 33 publications

(11 citation statements)

References 40 publications

(19 reference statements)

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“…We also found that the expression of keratinocyte differentiation markers (e.g., involucrin and filaggrin) is greatly decreased in the epidermis from aged wild-type (CD44+/+) mouse skin treated with a PKNγ inhibitor, Ro31-8220 [45–47] in the presence of HA L treatment (Fig. 4C-c vs. Fig.…”

Section: Resultsmentioning

confidence: 75%

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Selective matrix (hyaluronan) interaction with CD44 and RhoGTPase signaling promotes keratinocyte functions and overcomes age-related epidermal dysfunction

Bourguignon

Wong

Xia

et al. 2013

Journal of Dermatological Science

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Background Mouse epidermal chronologic aging is closely associated with aberrant matrix (hyaluronan, HA) -size distribution/production and impaired keratinocyte proliferation/differentiation, leading to a marked thinning of the epidermis with functional consequence that causes a slower recovery of permeability barrier function. Objective The goal of this study is to demonstrate mechanism-based, corrective therapeutic strategies using topical applications of small HA (HAS) and/or large HA (HAL) [or a sequential small HA (HAS) and large HA(HAL) (HAs-»HAL) treatment] as well as RhoGTPase signaling perturbation agents to regulate HA/CD44-mediated signaling, thereby restoring normal epidermal function, and permeability barrier homeostasis in aged mouse skin. Methods A number of biochemical, cell biological/molecular, pharmacological and physiological approaches were used to investigate matrix HA-CD44-mediated RhoGTPase signaling in regulating epidermal functions and skin aging. Results In this study we demonstrated that topical application of small HA (HAS) promotes keratinocyte proliferation and increases skin thickness, while it fails to upregulate keratinocyte differentiation or permeability barrier repair in aged mouse skin. In contrast, large HA (HAL) induces only minimal changes in keratinocyte proliferation and skin thickness, but restores keratinocyte differentiation and improves permeability barrier function in aged epidermis. Since neither HAS nor HAL corrects these epidermal defects in aged CD44 knock-out mice, CD44 likely mediates HA-associated epidermal functions in aged mouse skin. Finally, blockade of Rho-kinase activity with Y27632 or protein kinase-Nγ activity with Ro31-8220 significantly decreased the HA (HAS or HAL)-mediated changes in epidermal function in aged mouse skin. Conclusion The results of our study show first that HA application of different sizes regulates epidermal proliferation, differentiation and barrier function in aged mouse skin. Second, manipulation of matrix (HA) interaction with CD44 and RhoGTPase signaling could provide further novel therapeutic approaches that could be targeted for the treatment of various aging-related skin disorders.

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Section: Resultsmentioning

confidence: 75%

“…4B-b and Table 4). In contrast, aged wild-type (CD44+/+) mice similarly treated with a PKNγ inhibitor, Ro31-8220 [45–47] do not display noticeable changes in HA S -induced PCNA-positive cell numbers and subsequent changes in epidermal thickness (Fig. 4B-c and Table 4).…”

Section: Resultsmentioning

confidence: 99%

Selective matrix (hyaluronan) interaction with CD44 and RhoGTPase signaling promotes keratinocyte functions and overcomes age-related epidermal dysfunction

Bourguignon

Wong

Xia

et al. 2013

Journal of Dermatological Science

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“…In vivo, the selectivity of Ro-31-8220 and GF109203X was addressed in ␣T3-1 gonadotrophs expressing PKC␣, -⑀, and -. In that study, Ro-31-8220 was shown to be a selective inhibitor of calcium-, diterpine-, and phorbol ester-activable PKC isozymes ␣ and ⑀, while GF109203X was an equally effective inhibitor of basal PKC activity composed of ␣, ⑀, and activity as of activated PKC activity (34). Other studies have addressed the selectivity of staurosporine and its analogs to inhibit soluble PKC and membrane-associated PKC.…”

Section: Discussionmentioning

confidence: 99%

The Selective Protein Kinase C Inhibitor, Ro-31-8220, Inhibits Mitogen-activated Protein Kinase Phosphatase-1 (MKP-1) Expression, Induces c-Jun Expression, and Activates Jun N-terminal Kinase

Beltman¹,

McCormick²,

Cook³

1996

Journal of Biological Chemistry

179

145

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The role of protein kinase C (PKC) in inflammation, mitogenesis, and differentiation has been deduced in part through the use of a variety of PKC inhibitors. Two widely used inhibitors are the structurally related compounds GF109203X and Ro-31-8220, both of which potently inhibit PKC activity and are believed to be highly selective. While using GF109203X and Ro-31-8220 to address the role of PKC in immediate early gene expression, we observed striking differential effects by each of these two compounds. Growth factors induce the expression of the immediate early gene products MAP kinase phosphatase-1 (MKP-1), c-Fos and c-Jun. Ro-31-8220 inhibits growth factor-stimulated expression of MKP-1 and c-Fos but strongly stimulated c-Jun expression, even in the absence of growth factors. GF109203X displays none of these properties.These data suggest that Ro-31-8220 may have other pharmacological actions in addition to PKC inhibition. Indeed, Ro-31-8220 strongly stimulates the stress-activated protein kinase, JNK1. Furthermore, Ro-31-8220 apparently activates JNK in a PKC-independent manner. Neither the down-regulation of PKC by phorbol esters nor the inhibition of PKC by GF109203X affected the ability of Ro-31-8220 to activate JNK1. These data suggest that, in addition to potently inhibiting PKC, Ro-31-8220 exhibits novel pharmacological properties which are independent of its ability to inhibit PKC. Protein kinase C (PKC)1 is a large and diverse family of protein kinases, whose members are differentially regulated by calcium, phorbol esters, diglycerides, and phosphatidic acid (for a review, see Ref. 1). The various PKC isozymes play important signaling roles in cellular growth, differentiation, and homeostasis. The reported cellular roles of PKC have been identified largely through the use of PKC activators and inhibitors. The bisindolemaleimide class of compounds are staurosporine analogs which by all reports are specific for PKC (2, 3). Two widely used bisindolemaleimides, GF109203X and Ro-31-8220, differ by a single functional group, and although their isozyme specificity varies slightly, they are both potent and selective PKC inhibitors.In addition to cellular responses that may be attributed directly to PKC, PKC signaling pathways likely also contribute to cellular signaling through cross-talk mechanisms with other signaling cascades. Multiple lines of evidence indicate a role for PKC in regulating the ERK and JNK cascades which regulate transcription factors in response to external stimuli. The ERK cascade, a growth factor-stimulated signaling cascade is regulated by the Ras and Raf protooncogenes which activate MEK which in turn activates ERK1 and ERK2 (4, 5). ERK1 and -2 accumulate in the nucleus and phosphorylate and transactivate the transcription factor TCF/Elk-1 (6). A parallel signaling pathway activated by cellular stresses such as uv light and translational inhibitors leads to the activation of MEKK1, SEK, and in turn JNK1 and JNK2 (7, 8). The major role of the JNK signaling pathway is believed to be the...

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“…To test whether PKC is involved in this effect of EGF, we used two PKC inhibitors, Ro318220 and Gö6983, which have been shown to inhibit differentially novel and atypical PKCs, namely ⑀ and isoforms, respectively. The reported IC 50 of Gö6983 for PKC is 60 nM (17) and that of Ro318220 for PKC⑀ is 24 -48 nM (18,19), depending on the cell type studied. As shown in Fig.…”

Section: Effect Of Ro318220 and Gö6983 On The Ability Of Egf To Inhibmentioning

confidence: 99%

A Role for Protein Kinase Cε in the Inhibitory Effect of Epidermal Growth Factor on Calcium-stimulated Chloride Secretion in Human Colonic Epithelial Cells

Chow

Uribe²,

Barrett

2000

Journal of Biological Chemistry

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Epidermal growth factor (EGF) inhibits carbachol-induced chloride secretion in T 84 colonic epithelial cells and has been shown to activate phosphatidylinositol (PI) 3-kinase, leading to inhibition of a basolateral potassium conductance. We asked whether the inhibitory effect of EGF on secretion is due to activation of specific isoforms of protein kinase C (PKC) by PI 3-kinase. Western analysis revealed that PKC␣, ␥, ⑀, , , /, and were expressed in T 84 cells. Ro318220 (an inhibitor active against PKC⑀, 10 M) but not Gö 6983 (an inhibitor active against PKC, 10 M) reversed the inhibitory effect of EGF (100 ng/ml) on carbachol-stimulated chloride secretion. EGF induced the rapid translocation of PKC⑀ from the cytoplasm to the membrane. Wortmannin (50 M) and LY294002 (20 nM), which are PI 3-kinase inhibitors that by themselves had no effect on PKC⑀ activity, significantly suppressed PKC⑀ translocation activated by EGF. LY294002 also reversed the inhibitory action of EGF on chloride secretion. PI (3,4)P 2 increased membrane-associated PKC⑀ and reduced carbachol-induced 86 Rb؉ efflux. Antisense oligonucleotides against PKC⑀ decreased PKC⑀ mass and prevented the inhibitory effect of EGF on carbachol-induced 86 Rb ؉ efflux. Thus, the inhibitory effect of EGF on carbachol-induced chloride secretion involves the activation of PKC⑀ mediated by PI 3-kinase. Our findings contribute to the understanding of the cellular mechanisms that control chloride secretion.

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Characterisation of protein kinase C isoforms and enzymic activity from the αT3‐1 gonadotroph‐derived cell line

Cited by 33 publications

References 40 publications

Selective matrix (hyaluronan) interaction with CD44 and RhoGTPase signaling promotes keratinocyte functions and overcomes age-related epidermal dysfunction

Selective matrix (hyaluronan) interaction with CD44 and RhoGTPase signaling promotes keratinocyte functions and overcomes age-related epidermal dysfunction

The Selective Protein Kinase C Inhibitor, Ro-31-8220, Inhibits Mitogen-activated Protein Kinase Phosphatase-1 (MKP-1) Expression, Induces c-Jun Expression, and Activates Jun N-terminal Kinase

A Role for Protein Kinase Cε in the Inhibitory Effect of Epidermal Growth Factor on Calcium-stimulated Chloride Secretion in Human Colonic Epithelial Cells

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