Research in
our laboratory is focused primarily on three areas. Our most longstanding
project concerns the roles for microtubules (MTs) and MT motor proteins
in membrane trafficking. In addition, we are studying how actin filament
organization is regulated by a protein known as IQGAP1, and how hyperphosphorylated
forms of the MT-associated protein, tau, self assemble into paired
helical filaments in Alzheimer's disease. To address these topics,
we rely on the tools of biochemistry, cell and molecular biology,
and biophysics, with the ultimate aim being to understand in vivo
functions at the level of individual proteins.
Much of our work on membrane trafficking has concentrated on the MT
motor protein, kinesin. Over the course of several years, we demonstrated
that kinesin is a MT stimulated ATPase which moves organelles along
MTs, and is a motor for anterograde fast axonal transport in neurons,
and for movement of membranes from the Golgi apparatus to the endoplasmic
reticulum (ER). Now we are concentrating on how organelle motility
along MTs is regulated. We have obtained evidence that kinesin-mediated
membrane motility away from the Golgi is regulated by multiple GTPases,
including Cdc42 and Rac1. We are now attempting to determine how those
two proteins, and whatever additional GTPases may prove to be relevant,
control membrane movement along MTs.
In a related project, we are using live cell fluorescence microscopy
to study the dynamic behavior of membrane proteins fused to the naturally
fluorescent protein, GFP. Our efforts in this area are focused on
proteins that normally reside in specialized plasma membrane domains
that are known as caveolae, and represent sites at which many signalling
moclecules are concentrated. Our work on membrane trafficking has
profited from collaborations with several laboratories, including
those of Scott Brady and Dick Anderson (University of Texas Southwestern
Medical Center), Nobutaka Hirokawa (Univesity of Tokyo), Nancy Ratner
(University of Cincinnati) and Jennifer Lippincott-Schwartz (NIH).
IQGAP1 is a protein which was shown by us to bind directly to actin
filaments, and by other labs to bind directly to calmodulin, and to
activated (GTP-bound) forms of Rac1 and Cdc42. Because activated Rac1
and Cdc42 induce cortical actin filaments to rearrange dramatically,
we suspect that IQGAP1 serves as a direct molecular link between these
GTPases and cortical actin. We have purified native IQGAP1, and demonstrated
that it is a dimeric protein that cross-links actin filaments into
bundles and gels. We are now studying how Cdc42, Rac1, calmodulin,
GTP and calcium act coordinately to regulate the actin filament binding
and cross linking activities of IQGAP1.
Our work on tau has concentrated primarily on its interactions with
protein phosphatase 2A (PP2A). We have obtained evidence that PP2A
is a major tau phosphatase, and have mapped the binding sites on PP2A
for tau and microtubules, and on tau and microtubules for PP2A. Our
ultimate goal is to understand the role of tau phosphorylation in
the assembly of paired helical filamnets in vivo. Labs that have collaborated
with us to study tau have included those of Estelle Sontag and Marc
Mumby (University of Texas Southwestern Medical Center), Gloria Lee
(University of Iowa), Jeff Kuret (Ohio State University) and Roland
Brandt (University of Heidelberg).
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