Lecture 19 Study Guide

Cell Cycle I

  1. Name 3 cell cycle checkpoints. What is the purpose of each?

G1/S: regulated by Cdc2/G1 Cyclin; checks for growth factors, nutritional factors, and correct cell size

G2/M: regulated by Cdc2/Mitotic Cyclin; checks for completed replication and DNA integrity

Spindle: controlled by APC; checks if chromosomes are attached at metaphase plate

  1. What are the model systems that are frequently used to study cell cycle?

• Yeast

Two Types:

1. Fission Yeast: S. pombe

2. Budding Yeast: S. cerevisiae

Usefulness:

-reproduce rapidly

-small genome (<1% the size of mammals)

-ease of genetic manipulation

-ability to proliferate in haploid state

-easier to study mutations without second copy to mask mutation

• Animal Embryos

• Cell Culture

  1. What is a temperature sensitive mutant?

Cell-Division-Cycle Genes (cdc):

• First identified in yeast

• Mutations cause arrest of cell cycle

• Includes cyclins and cdk

  • High temps cause the gene to function abnormally

  1. How can BrdU and Flow Cytometry be used to study cell cycle progression?

BrdU:

-round cells indicate mitotic cells

-cytokinesis

-uses a labeled nucleotide that can incorporate into replicating DNA. Therefore, only cells that were replicating their DNA during exposure to the label will light up.

Pink nuclei indicate BRDU labeled cells. (BrdU = Thymidine analog)

Flow Cytometry procedure:

• Stain cells with dye that binds DNA, like Propidium Iodide or Ethidium Bromide

• Place single cell suspension in Flow Cytometer

• Machine sends one cell through at a time and analyzes the fluorescence of each cell

• Brighter cell means more DNA

  1. Draw a cell in metaphase. Label the following components: chromosomes, metaphase plate, nuclear envelope, centrosome, centrioles, spindle, and kinectochore. Label the 3 classes of microtubules.

  1. Why does the mitotic spindle build from the centrosome?

-Each cell has two centrosomes.

-Each centrosome has a pair of centrioles.

-Centrioles contain g-tubulin to initiate microtubule growth

  1. Why do microtubules exhibit dynamic instability instead of treadmilling?

Microtubule Dynamic Instability Increases in Mitosis:

•Microtubules become less stable

• Simultaneously, centrosomes gain increased ability to nucleate microtubules.

• Microtubules are regulated by Microtubule Associated Proteins (MAPs)

  1. Describe the “search and capture” of the spindle to the chromosomes.

1. nuclear envelope breaks down (M-Cdk phosphorylates nuclear lamins)

2. “Search and Capture” –Microtubules extends searching to attach to kinetochore to become stabilized

3. Chromosomes Attach Through Kinetochores – Complex of Proteins at the Centromere

4. Chromosomes are Tugged Until Equidistant from “Metaphase Plate”