The Cell Cycle

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  • Created by: rosieevie
  • Created on: 19-05-17 18:31

Parts of the Cell Cycle

Proliferating cells cycle between 3 stages:

  • Cell growth and chromosome replication
  • Chromosome segregation
  • Cell division

M phase divided into mitosis and cytokinesis

Mitosis - when replicated chromosomes separate into 2 new nuclei

Cytokinesis - when the cytoplasm of a single cell divides into 2 daughter cells

Long interphase contains DNA replication

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Studying the Cell Cycle

Xenopus Models - easy to see cell cycle occuring

  • Large diameter, pale spot = nucleus
  • Egg builds up stores of protein in nucleus and grows
  • Cleavage stage - no growth but rapid cell division forms blastomeres
  • Whole embryo size remains constant

Cell-Free Systems

  • Xenopus oocyte extract prepped by centrifugation
  • Easy to manipulate and take samples

Assay of Mammalian Cells in Culture - used to determine how cell density/drugs/growth factors affect cell proliferation

  • Fix cells and stain with Haematoxylin
  • Percentage of cells in M and S phase calculated - M chromosomes visible
  • Mitotic Index = Mitotic cells/Total cell number
  • Used to diagnose diseases/identify tumours
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Studying the Cell Cycle 2

Bromodeoxyuridine Labelling

Newly synthesised DNA labeled w/ bromodeoxyuridine (uracil w/ bromine attatched at 5' carbon)

Maintained levels due to semi-conservative model of DNA replication

Label detected by antibody staining w/ fluorescence or enzymatic activity

Also used to see speed of cycle progression - pulse of BrdU given at S phase - timed how long each phase lasts

Flow cytometry also used to analyse DNA content of population of cells - detect which stage of cycle cells are in

Model Systems

Yeast used - very visible and identified in different stages by morphology

Two types of division - fission and budding

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Studying the Cell Cycle 3

Temperature-Sensitive Cell Division Cycle Mutants

2 possibilities of cell cycle progression control:

  • Progression is free-running, one step follows the next without specific control
  • Progression controlled by specific mechanisms

If 2nd true - specific gene products involved called checkpoints occur

Knocking out checkpoints might be lethal so w/ critial genes temperature-sensitive mutations used

Permissive (low) temperatures - gene product functional as proteins dynamic at low temperatures - not denatured = cells can be in all stages of cycle

Restrictive (high) temperatures - gene product not functional as partially denatured = cells may grow in size but accumulate at specific stage in cycle

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Cell Cycle Control

Minimum of 3 cell cycle checkpoints for cells:

  • Spindle assembly checkpoint - check for chromosome attatchment to spindle
  • G1 checkpoint - check for cell size/nutrients/growth factors/DNA damage
  • G2 checkpint - check for cell size/DNA replication

2 classes of protein required for cell cycle:

  • Cyclin-dependent kinase (Cdk) - phosphorylates target proteins using ATP
  • Cyclin - activates Cdk

Cdk activity oscillates in sync with cell cycle but Cdk protein expression does not chang through cycle = another component must be activating it

Minimum of 3 cyclins required for cell cycle control - concentration of each oscilates w/ different pattern and they only activate cognate Cdks

Cyclin only expressed when cell is ready for corresponding transition and levels of cyclin must reach a threshold point before starting next stage - after threshold reached they are broken down

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