A genetic screen using forced protein-protein interactions to cause
dominant, gain-of-function phenotypes.
Michael DeVit,
Stan Fields
Dept of Genome Sciences, University of Washington, 1959
NE Pacific Ave, Seattle, WA 98195, USA
We are developing a novel
genetic screen to identify proteins that function in a common pathway or
process. The screen takes advantage of the observation that cellular
processes, such as signal transduction pathways, are often initiated by
the formation of multiprotein complexes, or by colocalization of
proteins. By artificially forcing components of a pathway, e.g., a
protein kinase and its substrate, to interact we might be able to cause
a gain-of-function phenotype, such as signal-independent activation of
the pathway. Thus, it should be possible to identify novel proteins in a
particular process by forcing a known component of a pathway to interact
with every other yeast protein, and testing each combination for a
phenotype associated with activation of the process. To force proteins
to interact, we are fusing them to the heterodimerizing leucine zippers
from the mammalian Fos and Jun proteins. We have identified the minimal
portions of Fos and Jun that will interact in yeast, and have shown that
a Fos-GFP fusion will colocalize with Jun-protein fusions. We are making
a library of yeast proteins fused to Fos, which will be used to carry
out test screens with this approach.
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