G1 Cyclin/Cyclin-dependent Kinase-coordinated Phosphorylation of Endogenous Pocket Proteins Differentially Regulates Their Interactions with E2F4 and E2F1 and Gene Expression

Calbó, Joaquim; Parreño, Matilde; Sotillo, Elena; Tan, Yong Zen; Mazo, Adela; Garriga, Judit; Graña, Xavier

doi:10.1074/jbc.m209181200

Cited by 82 publications

(98 citation statements)

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“…26 Pretreatment with TCPOBOP caused a strong enhancement of the nuclear levels of p107 compared with PH alone or unoperated mice (Figure 3b). The results also show that although the lack of liver regeneration observed in aged mice was associated with a strong increase in the hepatic levels of p27, pretreatment with TCPOBOP caused a significant reduction in the expression of this inhibitory protein.…”

Section: Xenobiotic Mitogens and Liver Regeneration A Columbano Et Almentioning

confidence: 91%

Potential utility of xenobiotic mitogens in the context of liver regeneration in the elderly and living-related transplantation

Columbano¹,

Simbula²,

Pibiri³

et al. 2008

Laboratory Investigation

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Although liver regeneration occurring after partial hepatectomy (PH) is greatly reduced in aged mice, liver hyperplasia induced by xenobiotic mitogens was found to be age independent. Here, we investigated the potential utility of mitogens in stimulating liver regeneration in old mice subjected to two-third PH. Although virtually no hepatocytes entered S phase 48 h after PH, pretreatment (2 h prior to surgery) with 1,4-bis(2-(3,5-dichloropyridyloxy)benzene (TCPOBOP), a ligand of constitutive androstane receptor, induced an increase of bromodeoxyuridine incorporation and enhanced the expression of cyclin D1, cyclin A and proliferating cell nuclear antigen . Next, we investigated the potential utility of mitogens in the context of donor conditioning prior to living-related transplantation. Three days after TCPOBOP administration to intact young mice, an almost doubling of the liver mass and DNA content occurred; the regenerative response to two-third resection of the TCPOBOP-induced hyperplastic liver was similar to that of mice subjected to PH alone, suggesting that an increased liver mass at the time of surgery does not inhibit the regenerative capacity. The present results suggest that mitogen-induced hyperplasia is a promising tool in conditions characterized by reduced regenerative capacity, such as in the elderly, or when a rapid increase of liver mass is required, such as in living-related transplantation.

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Section: Xenobiotic Mitogens and Liver Regeneration A Columbano Et Almentioning

confidence: 91%

Potential utility of xenobiotic mitogens in the context of liver regeneration in the elderly and living-related transplantation

Columbano¹,

Simbula²,

Pibiri³

et al. 2008

Laboratory Investigation

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“…All cell lines were maintained in Dulbecco's modified Eagle's medium (Cellgro) supplemented with 10% fetal bovine serum (FBS) (Sigma) at 37°C in a humidified atmosphere with 5% CO 2 . Serum starvation was performed as previously described (11). Density arrest was achieved by growing cells to high density until no dividing cells were visible under the microscope and maintained for an additional 48 h prior to any treatment.…”

Section: Methodsmentioning

confidence: 99%

“…Density arrest was achieved by growing cells to high density until no dividing cells were visible under the microscope and maintained for an additional 48 h prior to any treatment. Progression of cells through the cell cycle was measured by propidium iodide staining followed by flow cytometric analysis (PI/FACS) using a BD FACSCalibur flow cytometer and quantified with Cell Quest software (BD) as described earlier (11).…”

Section: Methodsmentioning

confidence: 99%

“…A mutant derivative of cdc6 (CDC6-S3D) in which all three CDK target residues were converted to aspartic acid was constructed by standard molecular biology methods and packaged identically to normal Cdc6. Adenoviral stocks were amplified using Ad-293 cells and purified by CsCl density gradient centrifugation as described previously (11,17). Viral titers were determined with the Adeno-X TM Rapid Titer Kit (BD Bioscience).…”

Section: Methodsmentioning

confidence: 99%

“…8). Cyclin E deregulation in certain human tumor cell lines and immortalized rat fibroblasts is associated with mitogen-independent cell cycle entry and progression through the cell cycle (11). However, when cyclin E is ectopically expressed in quiescent normal human fibroblasts (NHF), cells remain in G0 (12).…”

mentioning

confidence: 99%

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Coordinated Activation of the Origin Licensing Factor CDC6 and CDK2 in Resting Human Fibroblasts Expressing SV40 Small T Antigen and Cyclin E

Sotillo¹,

Garriga²,

Padgaonkar³

et al. 2009

Journal of Biological Chemistry

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We have previously shown that SV40 small t antigen (st) cooperates with deregulated cyclin E to activate CDK2 and bypass quiescence in normal human fibroblasts (NHF). Here we show that st expression in serum-starved and density-arrested NHF specifically induces up-regulation and loading of CDC6 onto chromatin. Coexpression of cyclin E results in further accumulation of CDC6 onto chromatin concomitantly with phosphorylation of CDK2 on Thr-160 and CDC6 on Ser-54. Investigation of the mechanism leading to CDC6 accumulation and chromatin loading indicates that st is a potent inducer of cdc6 mRNA expression and increases CDC6 protein stability. We also show that CDC6 expression in quiescent NHF efficiently promotes cyclin E loading onto chromatin, but it is not sufficient to activate CDK2. Moreover, we show that CDC6 expression is linked to phosphorylation of the activating T loop of CDK2 in serumstarved NHF stimulated with mitogens or ectopically expressing cyclin E and st. Our data suggest a model where the combination of st and deregulated cyclin E result in cooperative and coordinated activation of both an essential origin licensing factor, CDC6, and an activity required for origin firing, CDK2, resulting in progression from quiescence to S phase.Upon mitogenic stimulation mammalian G1 CDKs 4 trigger passage through the restriction point and the transition into DNA replication. In particular, cyclin E/CDK2 is activated in mid to late G1 and phosphorylates a variety of substrates that play critical roles in these processes. CDK2 cooperates with D-type cyclin/CDKs to inactivate E2F/pocket protein repressor complexes inducing the expression of DNA synthesis factors and other cell cycle regulators (reviewed in Refs. 1 and 2). CDK2 also phosphorylates DNA replication factors facilitating prereplication complex assembly and origin firing and plays additional roles in centrosome duplication and histone synthesis (reviewed in Ref. 1). In particular, it has been proposed that CDK2 phosphorylates the essential origin licensing factor CDC6 promoting its stabilization prior to inactivation of the APC Cdh1 ubiquitin ligase (3). This is thought to ensure that CDC6 accumulation precedes accumulation of other APC substrates that inhibit origin licensing. Moreover, CDK2-independent cyclin E functions have also been reported to be important for prereplication complex assembly in cells in transit from G0 into G1 (4, 5). In keeping with its role as positive regulator of major G1 transitions, deregulation of the cyclin E via gene amplification or defective protein turnover is commonly seen in primary tumors and is associated with poor prognosis (6 -8). In normal fibroblasts, ectopic expression of cyclin E has been associated with shortening of the G1 phase of the cell cycle (9, 10), and with induction of DNA damage (reviewed in Ref. 8). Cyclin E deregulation in certain human tumor cell lines and immortalized rat fibroblasts is associated with mitogen-independent cell cycle entry and progression through the cell cycle (11). However, w...

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Aging does not reduce the hepatocyte proliferative response of mice to the primary mitogen TCPOBOP

et al. 2004

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It has been shown that the magnitude of DNA synthesis and the time at which maximal DNA synthesis occurs after two-thirds partial hepatectomy (PH) is greatly reduced in the liver of aged rodents compared to young animals. This reduction could represent an intrinsic defect in proliferation or a more specialized change in the response to PH. We therefore evaluated the proliferative capacity of hepatocytes in aged animals, following treatment with primary liver mitogens. We show that treatment of 12-month-old CD-1 mice with the hepatomito-gen 1,4-bis[2-(3,5-dichloropyridyloxy)] benzene (TCPOBOP) caused an increase in hepato-cyte proliferation similar to that seen in young (8-week-old) mice. The labeling index was 82% in the livers of aged mice versus 76% in young animals. Histological observation demonstrated that the number of hepatocytes entering mitoses was similar in both groups; the mitotic indices were 2.5 per thousand and 2.7 per thousand, respectively. Additional experiments showed that the timing of DNA synthesis and M phase were nearly identical in both aged and young mice. Stimulation of hepatocyte DNA synthesis was associated with increased expression of several cell cycle-associated proteins (cyclin D1, cyclin A, cyclin B1, E2F, pRb, and p107); all were comparable in aged mice and young mice. TCPOBOP treatment also increased expression of the Forkhead Box transcription factor m l b (Foxmlb) to a similar degree in both groups. In conclusion, hepatocytes retain their proliferative capacity in old age despite impaired liver regeneration. These findings suggest that therapeutic use of mitogens would alleviate the reduction in hepatocyte proliferation observed in the elderly. (HEPATOLOGY 2004;40:981-988.) he liver plays a central role in the metabolic functions of the body, carrying out over 5,000 func-T tions, with a remarkable capacity to adapt to metabolic perturbance. Liver regeneration after partial hepatectomy (PH) typifies one such capacity: the organ Abbreviations: PH, partial hepatectomy; Foxmlb, Forkhead Box mlb; TCPOBOP, 1,4-bis[2-(3,5-dicbloropyridyloxy)]benzene; CAR, constitutive an-drostane receptor; TNF-a tumor necrosis factor (Y IL-6, interleukin 6; mRNA, messenger RNA; B r d u bromodeoyuridine; LI, labeling index; kb, kilobase; cDNA, complementary DNA; P A , R N h e protection assay; SDS, sodium dodecyl suFate. changes instantly from an essentially quiescent state to a rapidly growing one. Experimental models of regenera-tion have been the target of many studies that probed the control mechanisms of liver growth.2-5 This type of re-generative liver growth, termed "compensatory regenera-tion" or "compensatory hyperplasia," also occurs after liver cell necrosis due to chemical, nutritional, vascular, or viral injury. Several studies on the regenerative response of the liver that follows two-thirds PH have shown a striking difference in both the magnitude of the peak response in DNA synthesis and the time at which the maximal DNA synthesis occurs between young (4-8 weeks) and aged (12-15 month...

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G1 Cyclin/Cyclin-dependent Kinase-coordinated Phosphorylation of Endogenous Pocket Proteins Differentially Regulates Their Interactions with E2F4 and E2F1 and Gene Expression

Cited by 82 publications

References 43 publications

Potential utility of xenobiotic mitogens in the context of liver regeneration in the elderly and living-related transplantation

Potential utility of xenobiotic mitogens in the context of liver regeneration in the elderly and living-related transplantation

Coordinated Activation of the Origin Licensing Factor CDC6 and CDK2 in Resting Human Fibroblasts Expressing SV40 Small T Antigen and Cyclin E

Aging does not reduce the hepatocyte proliferative response of mice to the primary mitogen TCPOBOP

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