Targeting of PKCζ and PKB to caveolin-enriched microdomains represents a crucial step underpinning the disruption in PKB-directed signalling by ceramide

Elevated ceramide concentrations in adipocytes and skeletal muscle impair PKB (protein kinase B; also known as Akt)-directed insulin signalling to key hormonal end points. An important feature of this inhibition involves the ceramide-induced activation of atypical PKCζ (protein kinase C-ζ), which associates with and negatively regulates PKB. In the present study, we demonstrate that this inhibition is critically dependent on the targeting and subsequent retention of PKCζ–PKB within CEM (caveolin-enriched microdomains), which is facilitated by kinase interactions with caveolin. Ceramide also recruits PTEN (phosphatase and tensin homologue detected on chromosome 10), a 3′-phosphoinositide phosphatase, thereby creating a repressive membrane microenvironment from which PKB cannot signal. Disrupting the structural integrity of caveolae by cholesterol depletion prevented caveolar targeting of PKCζ and PKB and suppressed kinase–caveolin association, but, importantly, also ameliorated ceramide-induced inhibition of PKB. Consistent with this, adipocytes from caveolin-1−/− mice, which lack functional caveolae, exhibit greater resistance to ceramide compared with caveolin-1+/+ adipocytes. We conclude that the recruitment and retention of PKB within CEM contribute significantly to ceramide-induced inhibition of PKB-directed signalling.

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Combined thiazolidinedione–metformin treatment synergistically improves insulin signalling to insulin receptor substrate-1-dependent phosphatidylinositol 3-kinase, atypical protein kinase C and protein kinase B/Akt in human diabetic muscle

Diabetologia ◽

10.1007/s00125-008-1180-z ◽

2008 ◽

Vol 52 (1) ◽

pp. 60-64 ◽

Cited By ~ 14

Author(s):

N. Temofonte ◽

M. P. Sajan ◽

S. Nimal ◽

T. Pastoor ◽

C. Fumero ◽

...

Keyword(s):

Protein Kinase C ◽

Protein Kinase ◽

Insulin Receptor ◽

Insulin Signalling ◽

Protein Kinase B ◽

Insulin Receptor Substrate ◽

Metformin Treatment ◽

Phosphatidylinositol 3 Kinase ◽

Atypical Protein Kinase ◽

Kinase C

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Faculty Opinions recommendation of Exercise improves phosphatidylinositol-3,4,5-trisphosphate responsiveness of atypical protein kinase C and interacts with insulin signalling to peptide elongation in human skeletal muscle.

Faculty Opinions – Post-Publication Peer Review of the Biomedical Literature ◽

10.3410/f.1089955.543122 ◽

2008 ◽

Author(s):

Mark Hargreaves

Keyword(s):

Skeletal Muscle ◽

Protein Kinase C ◽

Protein Kinase ◽

Human Skeletal Muscle ◽

Insulin Signalling ◽

Atypical Protein Kinase C ◽

Atypical Protein Kinase ◽

Kinase C ◽

Peptide Elongation

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Inhibition of growth-factor-induced phosphorylation and activation of protein kinase B/Akt by atypical protein kinase C in breast cancer cells

Biochemical Journal ◽

10.1042/bj3520475 ◽

2000 ◽

Vol 352 (2) ◽

pp. 475-482 ◽

Cited By ~ 34

Author(s):

Muling MAO ◽

Xianjun FANG ◽

Yiling LU ◽

Ruth LAPUSHIN ◽

Robert C. BAST ◽

...

Keyword(s):

Breast Cancer ◽

Protein Kinase C ◽

Growth Factor ◽

Protein Kinase ◽

Cancer Cells ◽

Breast Cancer Cells ◽

Protein Kinase B ◽

Akt Phosphorylation ◽

Kinase C ◽

Pkc Inhibitors

The protein kinase B/Akt serine/threonine kinase, located downstream of phosphoinositide 3-kinase (PI-3K), is a major regulator of cellular survival and proliferation. Atypical protein kinase C (aPKC) family members are activated by PI-3K and also contribute to cell proliferation, suggesting that Akt and aPKC might interact to activate signalling through the PI-3K cascade. Here we demonstrate that blocking PKC activity in MDA-MB-468 breast cancer cells increased the phosphorylation and activity of Akt. Functional PI-3K was required for the PKC inhibitors to increase Akt phosphorylation and activation, potentially owing to the activation of specific PKC isoforms by PI-3K. The concentration dependence of the action of the PKC inhibitors implicates aPKC in the inhibition of Akt phosphorylation and activity. In support of a role for aPKC in the regulation of Akt, Akt and PKCζ or PKCλ/ℓ were readily co-precipitated from the BT-549 breast cancer cell line. Furthermore, the overexpression of PKCζ inhibited growth-factor-induced increases in Akt phosphorylation and activity. Thus PKCζ associates physically with Akt and decreases Akt phosphorylation and enzyme activity. The effects of PKC on Akt were transmitted through the PI-3K cascade as indicated by changes in p70 s6 kinase (p70s6k) phosphorylation. Thus PKCζ, and potentially other PKC isoenzymes, regulate growth-factor-mediated Akt phosphorylation and activation, which is consistent with a generalized role for PKCζ in limiting growth factor signalling through the PI-3K/Akt pathway.

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Prevention of tumour cell apoptosis associated with sustained protein kinase B phosphorylation is more sensitive to regulation by insulin signalling than stimulation of proliferation and extracellular signal-regulated kinase

Molecular and Cellular Biochemistry ◽

10.1007/s11010-017-2996-y ◽

2017 ◽

Vol 432 (1-2) ◽

pp. 41-54 ◽

Cited By ~ 4

Author(s):

Christoph Schmid ◽

Claudia Ghirlanda ◽

Markus Niessen

Keyword(s):

Protein Kinase ◽

Tumour Cell ◽

Cell Apoptosis ◽

Insulin Signalling ◽

Protein Kinase B ◽

Extracellular Signal Regulated Kinase ◽

Extracellular Signal ◽

Stimulation Of

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The role of Akt (protein kinase B) and protein kinase C in ischemia–reperfusion injury

Neurological Research ◽

10.1080/01616412.2015.1133024 ◽

2016 ◽

Vol 38 (4) ◽

pp. 301-308 ◽

Cited By ~ 19

Author(s):

Ethan Y. Zhao ◽

Aslan Efendizade ◽

Lipeng Cai ◽

Yuchuan Ding

Keyword(s):

Protein Kinase C ◽

Protein Kinase ◽

Reperfusion Injury ◽

Ischemia Reperfusion Injury ◽

Protein Kinase B ◽

Ischemia Reperfusion ◽

Kinase C

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Protein Kinase C-ε Induces Caveolin-Dependent Internalization of Vascular Adenosine 5′-Triphosphate–Sensitive K + Channels

Hypertension ◽

10.1161/hypertensionaha.108.110817 ◽

2008 ◽

Vol 52 (3) ◽

pp. 499-506 ◽

Cited By ~ 32

Author(s):

Jundong Jiao ◽

Vivek Garg ◽

Baofeng Yang ◽

Terry S. Elton ◽

Keli Hu

Keyword(s):

Smooth Muscle ◽

Protein Kinase C ◽

Angiotensin Ii ◽

Protein Kinase ◽

Cell Surface ◽

Small Interfering Rna ◽

Caveolin 1 ◽

Kinase C ◽

Interfering Rna ◽

Pkc Activation

Vascular ATP-sensitive K + (K ATP ) channels are critical regulators of arterial tone and, thus, blood flow in response to local metabolic needs. They are important targets for clinically used drugs to treat hypertensive emergency and angina. It is known that protein kinase C (PKC) activation inhibits K ATP channels in vascular smooth muscles. However, the mechanism by which PKC inhibits the channel remains unknown. Here we report that caveolin-dependent internalization is involved in PKC-ε–mediated inhibition of vascular K ATP channels (Kir6.1 and SUR2B) by phorbol 12-myristate 13-acetate or angiotensin II in human embryonic kidney 293 cells and immortalized human saphenous vein vascular smooth muscle cells. We showed that Kir6.1 substantially overlapped with caveolin-1 at the cell surface. Cholesterol depletion with methyl-β-cyclodextrin significantly reduced, whereas overexpression of caveolin-1 largely enhanced, PKC-induced inhibition of Kir6.1/SUR2B currents. Importantly, we demonstrated that activation of PKC-ε caused internalization of K ATP channels, the effect that was blocked by depletion of cholesterol with methyl-β-cyclodextrin, expression of dominant-negative dynamin mutant K44E, or knockdown of caveolin-1 with small interfering RNA. Moreover, patch-clamp studies revealed that PKC-ε–mediated inhibition of the K ATP current induced by PMA or angiotensin II was reduced by a dynamin mutant, as well as small interfering RNA targeting caveolin-1. The reduction in the number of plasma membrane K ATP channels by PKC activation was further confirmed by cell surface biotinylation. These studies identify a novel mechanism by which the levels of vascular K ATP channels could be rapidly downregulated by internalization. This finding provides a novel mechanistic insight into how K ATP channels are regulated in vascular smooth muscle cells.

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