Cell Growth & Differentiation, Vol 6, Issue 11 1485-1493, Copyright © 1995 by American Association of Cancer Research

ARTICLES

Growth imbalance and altered expression of cyclins B1, A, E, and D3 in MOLT-4 cells synchronized in the cell cycle by inhibitors of DNA replication

J Gong, F Traganos and Z Darzynkiewicz
Cancer Research Institute, New York Medical College, Valhalla 10595, USA.

Expression of cyclins at the translational level is generally studied by immunoblotting lysates of cells synchronized in the cycle. Most methods used to synchronize transformed cells induce growth imbalance. The aim of the present study was to analyze levels of cyclins B1, A, E, and D3 in the respective phases of the cycle in synchronized human leukemic MOLT-4 cells, correlate them with total cellular protein content (reflecting growth imbalance), and compare the synchronized cells with cells from unperturbed, asynchronous cultures. Expression of cyclins detected immunocytochemically in individual permeabilized cells was analyzed by multiparameter flow cytometry, which made it possible to relate position of the cell in the cell cycle with cyclin expression. Cells synchronized at the G1-S boundary by thymidine, mimosine, or aphidicolin had about 40% increased total protein and 4-5 fold higher levels of cyclins E and B1 compared to their G1 counterparts from unperturbed cultures. Expression of cyclin A in synchronized cells was 2-fold higher, while expression of cyclin D3 was essentially unaltered. The synchronized cells traversing S phase after release from the block had elevated but decreasing levels of cyclins E, B1, and A. Although the cyclin expression of cells reentering G1 was similar to that of their counterparts from asynchronous cultures, the total protein content was still elevated by about 30%. The data indicate that due to different degrees of imbalance in total protein and individual cyclin content, levels of cyclins detected by immunoblotting of cell lysates from synchronized cultures may not be representative of their expression in unperturbed cells. The elevated level of cyclin B1 in the cells arrested at the G1-S boundary may reflect the increased half-life of this protein, stabilized as the result of the overexpression of cyclin E.

This article has been cited by other articles:

				A. R. Sedaghat, J. German, T. M. Teslovich, J. Cofrancesco Jr., C. C. Jie, C. C. Talbot Jr., and R. F. Siliciano Chronic CD4+ T-Cell Activation and Depletion in Human Immunodeficiency Virus Type 1 Infection: Type I Interferon-Mediated Disruption of T-Cell Dynamics J. Virol., February 15, 2008; 82(4): 1870 - 1883. [Abstract] [Full Text] [PDF]

				D. C. Vaillant and M. Paulin-Levasseur Evaluation of Mammalian Cell-free Systems of Nuclear Disassembly and Assembly J. Histochem. Cytochem., February 1, 2008; 56(2): 157 - 173. [Abstract] [Full Text] [PDF]

				U. Kim, C.-W. Shu, K. Y. Dane, P. S. Daugherty, J. Y. J. Wang, and H. T. Soh Selection of mammalian cells based on their cell-cycle phase using dielectrophoresis PNAS, December 26, 2007; 104(52): 20708 - 20712. [Abstract] [Full Text] [PDF]

				K. L. Eward, E. C. Obermann, S. Shreeram, M. Loddo, T. Fanshawe, C. Williams, H.-I. Jung, A. T. Prevost, J. J. Blow, K. Stoeber, et al. DNA replication licensing in somatic and germ cells J. Cell Sci., November 15, 2004; 117(24): 5875 - 5886. [Abstract] [Full Text] [PDF]

				M. Shen, Y. Feng, C. Gao, D. Tao, J. Hu, E. Reed, Q. Q. Li, and J. Gong Detection of Cyclin B1 Expression in G1-Phase Cancer Cell Lines and Cancer Tissues by Postsorting Western Blot Analysis Cancer Res., March 1, 2004; 64(5): 1607 - 1610. [Abstract] [Full Text] [PDF]

				J. Litz, P. Carlson, G. S. Warshamana-Greene, S. Grant, and G. W. Krystal Flavopiridol Potently Induces Small Cell Lung Cancer Apoptosis during S Phase in a Manner That Involves Early Mitochondrial Dysfunction Clin. Cancer Res., October 1, 2003; 9(12): 4586 - 4594. [Abstract] [Full Text] [PDF]

				B. A. Narayanan, M. S. Condon, M. C. Bosland, N. K. Narayanan, and B. S. Reddy Suppression of N-Methyl-N-nitrosourea/Testosterone-induced Rat Prostate Cancer Growth by Celecoxib: Effects on Cyclooxygenase-2, Cell Cycle Regulation, and Apoptosis Mechanism(s) Clin. Cancer Res., August 1, 2003; 9(9): 3503 - 3513. [Abstract] [Full Text] [PDF]

				E. A. Fortunato, V. Sanchez, J. Y. Yen, and D. H. Spector Infection of Cells with Human Cytomegalovirus during S Phase Results in a Blockade to Immediate-Early Gene Expression That Can Be Overcome by Inhibition of the Proteasome J. Virol., May 3, 2002; 76(11): 5369 - 5379. [Abstract] [Full Text] [PDF]

				K. Shedden and S. Cooper Analysis of cell-cycle-specific gene expression in human cells as determined by microarrays and double-thymidine block synchronization PNAS, April 2, 2002; 99(7): 4379 - 4384. [Abstract] [Full Text] [PDF]

				M. T. Yip-Schneider, C. J. Sweeney, S.-H. Jung, P. L. Crowell, and M. S. Marshall Cell Cycle Effects of Nonsteroidal Anti-Inflammatory Drugs and Enhanced Growth Inhibition in Combination with Gemcitabine in Pancreatic Carcinoma Cells J. Pharmacol. Exp. Ther., September 1, 2001; 298(3): 976 - 985. [Abstract] [Full Text]

				G. Juan and C. Cordon-Cardo Intranuclear Compartmentalization of Cyclin E during the Cell Cycle: Disruption of the Nucleoplasm-Nucleolar Shuttling of Cyclin E in Bladder Cancer Cancer Res., February 1, 2001; 61(3): 1220 - 1226. [Abstract] [Full Text]

				A. P. Albino, G. Juan, F. Traganos, L. Reinhart, J. Connolly, D. P. Rose, and Z. Darzynkiewicz Cell Cycle Arrest and Apoptosis of Melanoma Cells by Docosahexaenoic Acid: Association with Decreased pRb Phosphorylation Cancer Res., August 1, 2000; 60(15): 4139 - 4145. [Abstract] [Full Text]

				S. Hatse, D. Schols, E. D. Clercq, and J. Balzarini 9-(2-Phosphonylmethoxyethyl)adenine Induces Tumor Cell Differentiation or Cell Death by Blocking Cell Cycle Progression through the S Phase Cell Growth Differ., June 1, 1999; 10(6): 435 - 446. [Abstract] [Full Text]

				G. A. Bjarnason, R. C. K. Jordan, and R. B. Sothern Circadian Variation in the Expression of Cell-Cycle Proteins in Human Oral Epithelium Am. J. Pathol., February 1, 1999; 154(2): 613 - 622. [Abstract] [Full Text] [PDF]

				K. Shedden and S. Cooper Analysis of cell-cycle-specific gene expression in human cells as determined by microarrays and double-thymidine block synchronization PNAS, April 2, 2002; 99(7): 4379 - 4384. [Abstract] [Full Text] [PDF]