Eileen Fong, Jin-Hong Kim,   Alexander Lin    

Mitosis: In Vivo and In Vitro Microscopy

Final Project







   

.: Introduction

Definition of Mitosis
Mitosis is the process of cell division where chromosomes are visibly condensed and the nucleus divides to form daughter chromosomes.  Mitosis is central to the cellular reproduction in which microtubules play a significant role in nearly all stages of the cell cycle.  Although mitosis is generally defined as the five M phases (prophase, prometaphase, metaphase, anaphase, and telophase) for the sake of microtubulin discussion, we also include descriptions of interphase and cytokinesis that complete the entire cell cycle resulting in two daughter cells.

Phases of the Cell Cycle:

Interphase
Interphase

The cell is engaged in metabolic activity and performing its prepare for mitosis (the next four phases that lead up to and include nuclear division). Chromosomes are not clearly discerned in the nucleus, although a dark spot called the nucleolus may be visible. The cell may contain a pair of centrioles  which are organizational sites for microtubules.
Prophase
Prophase

Chromatin in the nucleus begins to condense and becomes visible in the light microscope as chromosomes. The nucleolus disappears. The two centromeres of the cell, each with its pair of centrioles, move to opposite "poles" of the cell.  The mitotic spindle begins to form which is an array of spindles each containing approximately 20 microtubules that are synthesized from the the tubulin polymers in the cytoplasm.
Prometaphase
Prometaphase

The nuclear envelope disintergrates due to the dissolution of the lamins that stabalize the internal membrane.  Chromosomes can now attach to spindle microtubules via kinetochores, and undergo movement.
Metaphase
Metaphase

Spindle fibers align the chromosomes along the metaphase plate in the middle of the cell nucleus.  The proposed mechanism is that microtubules attached to opposite sides shrink or grow until they are at equilibrium.  Microtubule motors move them towards the minus end of shrinking microtubules, or vice versa.
Anaphase
anaphase

The paired chromosomes separate at the kinetochores and move to opposite sides of the cell. This is thought to result from movement along the microtubules powered by minus-end motors while the microtubules themselves shorten at both ends.  The overlapping spindle fibers move past each, pushing the poles farther apart, powered by plus-end kinesin motors.
Telophase
Telophase

Chromatids arrive at opposite poles of cell, and new membranes form around the daughter nuclei. The chromosomes disperse and are no longer visible under the light microscope. The spindle fibers disperse, and cytokinesis or the partitioning of the cell may also begin during this stage which is visible under light microscopy.
Cytokinesis
Cytokinesis

In animal cells, cytokinesis results when a fiber ring composed of a protein called actin around the center of the cell contracts pinching the cell into two daughter cells, each with one nucleus.
                                 Images courtesy of The Biology Project (Dept. of Biochemistry and Molecular Biophysics, University of Arizona)

.: Goals

The goal of our study was to characterize mitosis using light microscopy for in vivo observations of the cell cycle starting from fertilization of sea urchins to complete cell division over several cell cycles.  Given the significant role that microtubules play in mitosis, our study of mitosis in vitro involved microtubulin polymerization which was quantified and characterized using fluorescent microscopy.



















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