Calcium/Calmodulin-dependent Phosphorylation and Activation of Human Cdc25-C at the G2/M Phase Transition in HeLa Cells

Patel, Rajnikant; Holt, Mark R.; Philipova, Rada; Moss, Stephen J.; Schulman, Howard; Hidaka, Hiroyoshi; Whitaker, Michael

doi:10.1074/jbc.274.12.7958

Cited by 127 publications

(99 citation statements)

References 82 publications

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“…CaM is also known to interact with and regulate the activity and͞or nuclear localization of several cell-cycle regulatory proteins, including p21 Cip1 , cyclin D1-Cdk4, and CaM kinase II (46)(47)(48). Furthermore, CaM is involved in the expression of cell-cycle regulatory proteins Cdk2, Cdk4, cdc2, p21 Cip1 , cyclin B, and cyclin A and the enzymes of DNA replication, DNA polymerase ␣ and proliferating cell nuclear antigen (49,50).…”

Section: Discussionmentioning

confidence: 99%

Physical and functional interaction of androgen receptor with calmodulin in prostate cancer cells

Cifuentes

Mataraza

Yoshida

et al. 2003

Proc. Natl. Acad. Sci. U.S.A.

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Ca 2؉ and calmodulin (CaM) play a critical role in proliferation and viability of a wide variety of cells, including prostate cancer cells. We examined two prostate cancer cell lines, androgen-sensitive LNCaP and androgen-independent PC-3. Proliferation of LNCaP cells was six to eight times more sensitive to the inhibitory effect of the CaM antagonist N-(6-aminohexyl)-5-chloro-1-naphthalenesulfonamide hydrochloride (W-7) than were PC-3 cells. Because LNCaP cell proliferation is sensitive to stimulation by androgen, we assessed the physical and functional interaction between androgen receptor (AR) and CaM. We observed tight binding of AR to CaM when LNCaP cell extracts were subjected to CaM-affinity column chromatography. AR binding to CaM was Ca 2؉ -dependent and was inhibited by pretreatment of the cell extracts with W-7. Using immunofluorescence staining and confocal microscopy, we demonstrated colocalization of AR and CaM in the nucleus of LNCaP cells. Furthermore, the functional relevance of AR-CaM interactions in intact cells was revealed by the observation that W-7 was as effective as Casodex, an antiandrogen, in blocking AR-regulated expression of prostate-specific antigen in LNCaP cells. AR seems to interact with CaM directly because purified human AR could bind to CaM-agarose, and CaM could be detected in AR-immunoprecipitate prepared from purified soluble proteins. These studies provide direct evidence for physical and functional interaction between AR and CaM and suggest the potential usefulness of CaM antagonists in blocking AR activity in prostate cancer.

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Section: Discussionmentioning

confidence: 99%

Physical and functional interaction of androgen receptor with calmodulin in prostate cancer cells

Cifuentes

Mataraza

Yoshida

et al. 2003

Proc. Natl. Acad. Sci. U.S.A.

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“…Cell Culture and Preparation of Cell Extracts-Human cervical carcinoma cells (HeLa) were cultured as described previously (33). To obtain mitotically arrested cells, an asynchronous population of HeLa cells was treated with either nocodazole (3 M), taxol (1 M), vincristine (1 M), or vinblastine (1 M) for various times (0 -30 h).…”

Section: Methodsmentioning

confidence: 99%

“…To obtain mitotically arrested cells, an asynchronous population of HeLa cells was treated with either nocodazole (3 M), taxol (1 M), vincristine (1 M), or vinblastine (1 M) for various times (0 -30 h). Mitotic cells were collected by mechanical shake-off, washed in Dulbecco's phosphate-buffered saline, and lysed in the appropriate buffer depending on the protein kinase being assayed as described previously (33). Radioimmunoprecipitation buffer containing 1% (v/v) Triton X-100 was used to prepare cell lysates for immunoblotting with the I B␣ antibodies.…”

Section: Methodsmentioning

confidence: 99%

NF-κB Promotes Survival during Mitotic Cell Cycle Arrest

Mistry

Deacon

Mistry

et al. 2004

Journal of Biological Chemistry

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By activating the mitotic checkpoint, anti-microtubule drugs such as nocodazole cause mammalian cells to arrest in mitosis and then undergo apoptosis. Microtubule depolymerization is rapid and results in the activation of the transcription factor NF-B and induction of NF-B-dependent gene expression. However, the functional consequence of NF-B activation has remained unclear. Evidence has accumulated to suggest that NF-B transcriptional activity is required to suppress apoptosis. In the present study, we confirm and extend previous findings that microtubule depolymerization leads to the rapid activation of NF-B and test the hypothesis that the induction of NF-B regulates cell survival during mitotic cell cycle arrest in order to define its role. Using a range of functional assays, we have shown that microtubule depolymerization correlates with the activation of IKK␣ and IKK␤; the phosphorylation, ubiquitination, and degradation of I B␣; the translocation of native p65 (RelA) into the nucleus; and increased NF-B transcriptional activity. By inhibiting either the activation of the IKKs or the degradation of I B␣, we find that the level of apoptosis is significantly increased in the mitotically arrested cells. Inhibition of NF-B signaling in the nonmitotic cells did not affect their survival. We establish that although NF-B is activated rapidly in response to microtubule depolymerization, its cell survival function is not required until mitotic cell cycle arrest, when the mitotic checkpoint is activated and apoptosis is triggered. We conclude that NF-B may regulate the transcription of one or more antiapoptotic proteins that may regulate cell survival during mitotic cell cycle arrest.

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“…Thus the regulation of 14-3-3 binding to the S323 site is likely to be at the level of phosphorylation. The S216 site on cdc25C, which is identical to the S323 motif in cdc25B, is a substrate for a wide range of kinase in vitro, including c-TAK1 (Peng et al, 1998), CaM kinase II (Patel et al, 1999), and the checkpoint kinases chk1 and chk2 (Peng et al, 1997;Blasina et al, 1999). It is possible that these kinases may also be involved in regulating the conserved S323 site on cdc25B.…”

Section: Discussionmentioning

confidence: 99%

Cdc25B activity is regulated by 14-3-3

Forrest

Gabrielli

2001

Oncogene

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Calcium/Calmodulin-dependent Phosphorylation and Activation of Human Cdc25-C at the G2/M Phase Transition in HeLa Cells

Abstract: cdc25-c is not phosphorylated, p34 cdc2 remains tyrosine phosphorylated, and there is no increase in histone H1 kinase activity. Our data suggest that a calcium-calmodulin-dependent step may be involved in the initial activation of p54 cdc25-c .

Cited by 127 publications

References 82 publications

Physical and functional interaction of androgen receptor with calmodulin in prostate cancer cells

Physical and functional interaction of androgen receptor with calmodulin in prostate cancer cells

NF-κB Promotes Survival during Mitotic Cell Cycle Arrest

Cdc25B activity is regulated by 14-3-3

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