MUTATIONAL ANALYSIS OF AMOEBOID CHEMOTAXIS

Project: Research project

Project Details

Description

Amoeboid chemotaxis is an important phenomenon in which signal
transduction and cell motility interact. It may be critical to movement
of cancer cells away from the original in metastasis as well as the
ability of white blood cells such as to respond rapidly to infection.
Dictyostelium discoideum amoebae provide a powerful model system for
analyzing amoeboid motility and chemotaxis using a genetic approach.
They have a habloid genome which allbws expression of be analyzed
genetically, and are easily cultivited. The ultimate goal of this study
is a dissection of amoeboid motility and chemotaxis by analyzing the
biochemical and behavioral properties of mutants.

Mutants will be generated by chemical mutagenesis. Dictyostelium
mutants will be selected using an established selection chamber and
screening assay. The mutants that are selected will be analyzed in
terms of their biochemistry and behavior. Chemoattractant binding,
production of intracellular messengers (such as IP3, calcium, and cGMP),
and actin/myosin association with the cytoskeleton will assayed.
Computer-controlled image analysis will be used to determine single cell
speed, turning, orientation and shape in response to both stable
chemoattractant gradients and rapid changes in concentration. Genetic
studies will determine complementation groups and insure that the
biochemical defect is associated with the behavioral alteration.

The wild-type copies of the mutant genes will be cloned by complementing
selected mutants with a library of genomic DNA in a Dictyostelium
transformation vector. Chemotactic transformants will be selected on
the basis of morphology or using the selection chamber. Recovery of the
transformation vector from chemotactic transformants will lead to the
identification of wild-type sequences that can complement the mutation.
Tbe long term objective is to identify the biochemical processes
responsible for particular behavioral responses.
StatusFinished
Effective start/end date7/1/926/30/93

Funding

  • National Institute of General Medical Sciences

ASJC

  • Genetics
  • Molecular Biology
  • Biophysics
  • Biotechnology
  • Microbiology

Fingerprint Explore the research topics touched on by this project. These labels are generated based on the underlying awards/grants. Together they form a unique fingerprint.