Inhibition of rat hepatocyte proliferation by transforming growth factor ? and glucagon is associated with inhibition of ERK2 and p70 S6 kinase

Dixon, Mark C.; Agius, Loranne; Yeaman, Stephen J.; Day, Christopher P.

doi:10.1002/hep.510290516

Cited by 43 publications

(29 citation statements)

References 59 publications

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“…A lower phosphorylation of p70S6K could contribute to the decline in the proliferative response of hepatocytes. 44,45 We demonstrated that downregulation of p70S6K expression with siRNA significantly inhibits DNA synthesis. Furthermore, inhibition of ERK2 by siRNA decreased p70S6K phosphorylation, showing that p70S6K was located downstream of ERK2 in hepatocytes.…”

Section: Discussionmentioning

confidence: 92%

An MLCK-dependent window in late G1 controls S phase entry of proliferating rodent hepatocytes via ERK-p70S6K pathway

et al. 2006

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We show that MLCK (myosin light chain kinase) plays a key role in cell cycle progression of hepatocytes: either chemical inhibitor ML7 or RNA interference led to blockade of cyclin D1 expression and DNA replication, providing evidence that MLCK regulated S phase entry. Conversely, inhibition of RhoK by specific inhibitor Y27632 or RhoK dominant-negative vector did not influence progression in late G1 and S phase entry. Inhibition of either MLCK or RhoK did not block ERK1/2 phosphorylation, whereas MLCK regulated ERK2-dependent p70S6K activation. In addition, DNA synthesis was reduced in hepatocytes treated with p70S6K siRNA, demonstrating the key role played by the kinase in S phase entry. Interestingly, after the G1/S transition, DNA replication in S phase was no longer dependent on MLCK activity. We strengthened this result by ex vivo experiments and evidenced an MLCK-dependent window in late G1 phase of regenerating liver after two-thirds partial hepatectomy. In conclusion, our results underline an MLCK-dependent restriction point in G1/S transition, occurring downstream of ERK2 through the regulation of p70S6K activation, and highlighting a new signaling pathway critical for hepatocyte proliferation. T he G1 phase is a preparative step where cells temporally integrate complex signals from the microenvironment. At this time, the cells are able to switch between proliferation, differentiation or apoptosis, depending on extracellular matrix composition, cytokines, and cell-cell contacts. In the hepatocyte, a highly differentiated cell, cell cycle progression in vivo during liver regeneration is regulated by a sequential acquisition of complex signals depending on transduction pathway activation. 1-3 Primary cultures of hepatocytes are a powerful model in studying the precise sequence of events that are necessary for cell progression from a G0-like state to S phase. The model mimics the physiological process of hepatic regeneration after liver injury or partial hepatectomy. In this context, we have already demonstrated that epidermal growth factor (EGF) possesses two distinct and complementary effects on G1 phase hepatocytes. 4 First, the growth factor is a morphogen in early G1 phase by inducing controlled spreading of hepatocytes via integrin ␤1 regulation. Second, a mitogenic effect occurs in midlate G1 phase and allows hepatocytes to progress through a restriction point located two thirds of the way through G1 phase. [5][6][7] In addition, EGF promotes cell progression up to late G1. 7 In hepatocytes, EGF activates specific transduction pathways. MEK and PI3K signaling cascades are essential for progression past the G1/S checkpoint and hepatocyte progression to S phase. 8 They both control expression of cyclin D1, a key cell cycle protein that is upregulated in the pre-replicative phase of liver regeneration and in primary culture hepatocytes. 4,9,10 MLCK and RhoK, two kinases linked to the contractile apparatus, are known to be involved in the regulation of a wide range of cellular functions, including prol...

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Section: Discussionmentioning

confidence: 92%

An MLCK-dependent window in late G1 controls S phase entry of proliferating rodent hepatocytes via ERK-p70S6K pathway

et al. 2006

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“…ERK MAP kinase and PKB(Akt) are activated by EGF in a number of cell types. 15,18,37,49 Time course experiments showed that addition of EGF to primary rat hepatocyte cultures caused sustained stimulation of ERK MAP kinase (p44 MAPK and p42 MAPK) 5 minutes to 2 hours after EGF addition ( Fig. 1A and 1B).…”

Section: Egf Stimulates Erk Map Kinase Activation and Pkb(akt) Activamentioning

confidence: 98%

“…TGF-␤ 1 is a potent inducer of apoptosis in hepatocytes and several hepatoma cell lines, as well as in regressing liver in vivo. 3,36 In vitro, TGF-␤ 1 acts as a physiological inhibitor of cellular proliferation, 1,37 and the addition of TGF-␤ 1 to primary hepatocyte cultures results in high levels of apoptosis. [38][39][40][41] In this study, we have investigated the roles of different signaling pathways engaged by EGF and TNF-␣ in the suppression of TGF-␤ 1 -induced apoptosis.…”

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confidence: 99%

The role of protein kinase B and mitogen-activated protein kinase in epidermal growth factor and tumor necrosis factor ?-mediated rat hepatocyte survival and apoptosis

2000

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Perturbation of hepatocyte growth regulation is associated with a number of liver diseases such as fibrosis and cancer. These diseases are mediated by a network of growth factors and cytokines that regulate the induction of hepatocyte proliferation and apoptosis. In this study, we have investigated the role of signaling pathways activated by tumor necrosis factor ␣ (TNF-␣) and epidermal growth factor (EGF) in the regulation of apoptosis induced by transforming growth factor ␤ 1 (TGF-␤ 1 ), because this physiological factor is believed to regulate spontaneous apoptosis in the liver. We show that pretreatment with (10 ng/mL) EGF or (25 ng/mL) TNF-␣ can suppress TGF-␤ 1 -induced apoptosis by 73% and 50%, respectively, in isolated rat hepatocytes. However, suppression of TGF-␤ 1 -induced apoptosis by EGF and TNF-␣ occurs via different protein kinase signaling pathways. Using specific inhibitors, we show that suppression of apoptosis by EGF is dependent on activation of phosphoinositide 3-kinase (PI 3-kinase) and the extracellular signal regulated kinase (ERK) mitogenactivated protein (MAP) kinase pathways, but not p38 MAP kinase. In contrast, suppression of TGF-␤ 1 -induced apoptosis by TNF-␣ does not require PI 3-kinase and protein kinase B (PKB or Akt)-mediated pathways, but is dependent on ERK and p38 MAP kinase activity. These data contribute to our understanding of the intracellular survival signals that play a role in normal liver homeostasis and in diverse pathological conditions. (HEPATOLOGY 2000;31:420-427.)

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“…Studies using a perfused rat liver preparation show that glucagon stimulates protein degradation, inhibits protein synthesis, and prevents completely the stimulation of protein synthesis induced by elevated levels of perfusate amino acid concentration (6). Studies using rat hepatocytes show that glucagon attenuates the stimulatory effect of epidermal growth factor on phosphorylation of the ribosomal protein (rp) 1 S6 kinase S6K1 (7).…”

mentioning

confidence: 99%

Glucagon Represses Signaling through the Mammalian Target of Rapamycin in Rat Liver by Activating AMP-activated Protein Kinase

Kimball

Siegfried

Jefferson

2004

Journal of Biological Chemistry

120

104

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The opposing actions of glucagon and insulin on glucose metabolism within the liver are essential mechanisms for maintaining plasma glucose concentrations within narrow limits. Less well studied are the counterregulatory actions of glucagon on protein metabolism. In the present study, the effect of glucagon on amino acid-induced signaling through the mammalian target of rapamycin (mTOR), an important controller of the mRNA binding step in translation initiation, was examined using the perfused rat liver as an experimental model. The results show that amino acids enhance signaling through mTOR resulting in phosphorylation of eukaryotic initiation factor 4E-binding protein (4E-BP)1, the 70-kDa ribosomal protein (rp)S6 kinase, S6K1, and rpS6. In contrast, glucagon repressed both basal and amino acid-induced signaling through mTOR, as assessed by changes in the phosphorylation of 4E-BP1 and S6K1. The repression was associated with the activation of protein kinase A and enhanced phosphorylation of LKB1 and the AMP-activated protein kinase (AMPK). Surprisingly, the phosphorylation of two S6K1 substrates, rpS6 and eukaryotic initiation factor 4B, was not repressed but instead was increased by glucagon treatment, regardless of the amino acid concentration. The latter finding could be explained by the glucagoninduced phosphorylation of the ERK1 and the 90-kDa rpS6 kinase p90 rsk . Thus, glucagon represses phosphorylation of 4E-BP1 and S6K1 through the activation of a protein kinase A-LKB-AMPK-mTOR signaling pathway, while simultaneously enhancing phosphorylation of other downstream effectors of mTOR through the activation of the extracellular signal-regulated protein kinase 1-p90 rsk signaling pathway. Amino acids also enhance AMPK phosphorylation, although to a lesser extent than glucagon and amino acids combined.The counter-regulatory action of glucagon on insulin-stimulated glucose metabolism is well documented in the literature (1). In contrast, few studies have examined the effects of the hormone on protein metabolism. Reports suggest that in vivo glucagon has an overall catabolic effect on protein metabolism, although the mechanisms through which it acts are unclear (2-5). Studies using a perfused rat liver preparation show that glucagon stimulates protein degradation, inhibits protein synthesis, and prevents completely the stimulation of protein synthesis induced by elevated levels of perfusate amino acid concentration (6). Studies using rat hepatocytes show that glucagon attenuates the stimulatory effect of epidermal growth factor on phosphorylation of the ribosomal protein (rp) 1 S6 kinase S6K1 (7).In rat liver, the stimulation of S6K1 phosphorylation by hormones and nutrients is regulated through a signal transduction pathway involving a protein kinase referred to as the mammalian target of rapamycin (mTOR). Phosphorylation activates S6K1, which then phosphorylates rpS6 (8) and eukaryotic initiation factor (eIF)4B (9), resulting in an enhanced translation of subsets of mRNA such as those containing an oligopyri...

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Inhibition of rat hepatocyte proliferation by transforming growth factor ? and glucagon is associated with inhibition of ERK2 and p70 S6 kinase

Cited by 43 publications

References 59 publications

An MLCK-dependent window in late G1 controls S phase entry of proliferating rodent hepatocytes via ERK-p70S6K pathway

An MLCK-dependent window in late G1 controls S phase entry of proliferating rodent hepatocytes via ERK-p70S6K pathway

The role of protein kinase B and mitogen-activated protein kinase in epidermal growth factor and tumor necrosis factor ?-mediated rat hepatocyte survival and apoptosis

Glucagon Represses Signaling through the Mammalian Target of Rapamycin in Rat Liver by Activating AMP-activated Protein Kinase

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