Baldin notes - 1 Baldin, V., J. Lukas, M.J. Marcote, M....

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Unformatted text preview: 1 Baldin, V., J. Lukas, M.J. Marcote, M. Pagano, G. Draetta. (1993) Cyclin D1 is a nuclear protein required for cell cycle progression in G1. Genes & Development 7:812-821. A. Major Questions •Cyclins and cdks that are acting at mitosis are fairly well deFned at this point in yeast and humans •What is MP¡? •Cdk1(cdc2) is the only yeast cdk and thus is required for G 1 and S phase progression too •But mammalian cdk1 is required only at mitosis •What are the cdks and cyclins required for G 1 and entry into S phase in mammals? •What are the human G 1 cyclins? •Cyclin D1 was identiFed in a screen for human proteins that could complement yeast mutants. •Cyclin D1 is over-expressed in many tumors and contributes to transformation •Difference between transformation of yeast or bacteria and transformation as it applies to mammalian cells! B. Hypothesis •Cyclin D1 is a human G 1 cyclin, important for progression through G 1 2 C. Approach: Biochemistry/ Immunoprecipitations / Cell culture Experimental Strategy: 1. Examine Cyclin D1 levels during the cell cycle •Arrest cells in quiescent state called G ( HOW ?) •G differs from G 1- the cell has degraded essential components needed to complete G 1 . •Some of these components are cdk's (recall vertebrates produce several cdks). •These must be re-synthesized when the cells are provided with serum, and this delays the time it takes the cells to reach S-phase. 3 C. Approach: Biochemistry/ Immunoprecipitations / Cell culture Experimental Strategy: 1. Examine Cyclin D1 levels during the cell cycle •Monitor when S phase begins as cells proceed through cell cycle ( HOW ?) •BrdU is a Thymidine analog •Cells will incorporate BrdU in place of T as they synthesize their DNA •Cells that have incorporated BrdU in their DNA are recognized by indirect...
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This note was uploaded on 10/19/2009 for the course B M B 460 taught by Professor Hanna-rose,w during the Spring '08 term at Penn State.

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Baldin notes - 1 Baldin, V., J. Lukas, M.J. Marcote, M....

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