TGFbeta regulates the expression and activities of G2 checkpoint kinases in human myeloid leukemia cells.
Transforming Growth Factor-beta (TGFbeta) is known to be a negative regulator of G1 cyclin/cdk activity. It is not clear whether TGFbeta has any effect on G2 checkpoint kinases. We have found that TGFbeta downregulated the expression of several G2 checkpoint kinases including cdc2, cyclin B1, and cdc25c without causing cell accumulation in G2/M phases in two human leukemia cell lines. The inhibition was time-dependent with a maximal inhibition being observed by 24h for cyclin B1 and cdc2 and by 48h for cdc25c. The inhibition was not a result of G1 arrest but a direct effect of TGFbeta which downregulates their expression at mRNA level. In proliferating cells, there was a significant formation of cdc2-pRb complexes, which was decreased to 30% of control levels by 48h after initiating TGFbeta treatment. Cdc2 showed a marked kinase activity on GST-Rb protein in proliferating cells detected by in vitro kinase assay, which was downregulated in response to TGFbeta. In addition, TGFbeta caused a rapid and transient dephosphorylation of cdc2 (Tyr15) and cdc25c (Ser216) for about 2-3h before a dramatic decrease of both molecules by 48h. Taken together, our data suggest that TGFbeta has a direct inhibitory effect on G2 checkpoint kinases, which is regulated at mRNA level. The transient activation of cdc2 and cdc25c and subsequent inhibition of cdc2, cyclin B1, and cdc25c could amplify TGFbeta-induced G1 arrest and growth inhibition.
Hu
Xiaotang
X
Interdisciplinary Oncology Program, University of South Florida, H Lee Moffitt Cancer Center and Research Institute, Tampa, FL 33161, USA. xthu@mail.barry.edu
Cui
Dongming
D
Moscinski
Lynn C
LC
Zhang
Xiaohong
X
Maccachero
Vivian
V
Zuckerman
Kenneth S
KS
D016203:CDC2 Protein Kinase / Q000235:genetics / Q000378:metabolism; D018797:Cell Cycle Proteins / Q000235:genetics / Q000378:metabolism; D045744:Cell Line, Tumor; D019926:Cyclin B / Q000235:genetics / Q000378:metabolism; D056744:Cyclin B1; D016195:G2 Phase / Q000502:physiology*; D006801:Humans; D007951:Leukemia, Myeloid / Q000276:immunology*; D011993:Recombinant Fusion Proteins / Q000235:genetics / Q000378:metabolism; D016160:Retinoblastoma Protein / Q000235:genetics / Q000378:metabolism; D016212:Transforming Growth Factor beta / Q000378:metabolism*; D020687:cdc25 Phosphatases / Q000235:genetics / Q000378:metabolism
C534699:CCNB1 protein, human; D018797:Cell Cycle Proteins; D019926:Cyclin B; D056744:Cyclin B1; D011993:Recombinant Fusion Proteins; D016160:Retinoblastoma Protein; D016212:Transforming Growth Factor beta; D016203:CDC2 Protein Kinase; C517009:CDC25C protein, human; D020687:cdc25 Phosphatases