Specialty:
Basic Reproductive Sciences
Location:
University of Colorado Anschutz Medical Campus, Aurora
Positions Available: 1
Objective:
Estrogen (E2) binds to and
activates the estrogen receptor to regulate both normal and cancerous growth of
uterine, breast and ovarian tissues. Estrogen receptors (ER) function as
ligand-dependent (estrogen is the ligand) transcription factors which control
target gene expression.
Upon estrogen binding,
receptors localize to the nucleus, bind estrogen response elements in gene
promoters, and regulate transcription. Recently, estrogen receptors have been
shown to localize to the plasma membrane, associate with membrane proteins, and
function as signal transduction molecules—this activity is termed extranuclear,
nongenomic or nonclassical action.
The
goal of this project is to elucidate the mechanisms by which E2 and ER initiate
rapid cytoplasmic signaling. We will test the hypothesis that E2-activated ER interact
with c-Src and PI3 kinases to initiate the signaling cascades that control
uterine cancer cell growth.
Description:
Endometrial (uterine) cancer
cells expressing a mutant version of the ER that is solely cytosolic will be
treated +/- estrogen and harvested under non-denaturing (native protein
folding) conditions. Subsequent co-immunoprecipitation (co-IP) will be used to
determine if ER can interact with the signaling kinases, c-Src and PI3 kinase. Co-IP
uses antibodies to ER to precipitate ER and associated proteins out of the cell
lysate. The immunoprecipitated proteins will be resolved by polyacrylamide gel
electrophoresis and Western blotting will be used to determine if c-Src and PI3
kinase interact with ER.
The
contribution of these kinases to uterine cancer cell growth will be determined
using kinase inhibitors in conjuction with proliferation assays.
The student will
learn:
basic tissue culture techniques
protein extraction, immunoprecipitation,
and immunoblotting
densitometric quantitation and basic statistical
analyses, and
how to keep a lab notebook, interpret data and write an
abstract. My goal is for the student to produce publishable data and leave the
lab with potential co-authorship on a peer-reviewed journal article.