SUNDAY, DECEMBER 4- Late Abstracts 1 - Molecular Biology of the ...

SUNDAY
xenografts in mice. Comparative studies in spontaneous vs experimental metastasis models
suggest that loss of Pfn1 expression likely promotes metastasis-promoting events at the primary
tumor site. ECM invasion (an important step for tumor metastasis) of breast cancer cells is
increased upon loss of Pfn1 expression. Experiments involving selective perturbation of
signaling pathway downstream of PI3-Kinase (PI3K) further revealed that hyper-invasiveness of
Pfn1-depleted cells is mediated by a PI3K-PI(3,4)P2-lamellipodin signaling axis. Finally, as a
clinical correlate of these in vitro and in vivo findings, analyses of Pfn1 expression in primary
tumors revealed that Pfn1 downregulation occurs in all molecular subtypes of breast cancer,
somewhat more prominently in estrogen receptor-negative tumors. Furthermore, tumors
involving significant lymph node infiltration and/or distant metastasis present the most dramatic
downregulation in Pfn1 expression. These findings imply that Pfn1 could serve as a new
molecular marker for breast cancer malignancy.
2051
Reestablishing the polarity of Dictyostelium cells to understand the organization of the
cytoskeleton and signaling complexes during chemotaxis.
D. Jowhar 1 , G. Wright 1 , C. Janetopoulos 1,2 ;
1 Biological Sciences, Vanderbilt University, Nashville, TN, 2 Cell and Developmental Biology,
Vanderbilt University, Nashville, TN
Cells migrating in a chemical gradient typically have a polarized morphology, where they display
a distinct front and back. Cells can also have different localized sensitivities to chemoattractants
depending on their degree and type of polarity. To better understand these phenomena, we
used the social amoeba Dictyostelium discoideum that expressed GFP-tagged proteins known
to localize to distinct regions of the cell during cell migration. Dictyostelium cells were lured into
a specialized microfluidic device developed in our laboratory which confined the cells in narrow
3-dimensional Polydimethylsiloxane (PDMS) channels. The chemoattractant gradient in this
experimental platform could be reversed so
that the rear of the cell was exposed to a higher concentration of chemoattractant than the front
of the cell. Cells in the channels would freeze, become unpolarized, and were then capable of
forming a new front at the former rear of the cell. We have observed the temporal and spatial
loss and gain of localization of various signaling and cytoskeletal molecules at both the rear and
front of the cell as polarity is broken down and re-established. We monitored the localization of
PHcrac (a biosensor for PI(3,4)P2 and PI(3,4,5)P3), RBD (a biosensor for Ras activity), PTEN-
GFP (a phosphatase which elevates levels of PI(4,5)P2), LimE (a marker for F-actin), and
TACC-GFP (a microtubule centrosome and plus-end
marker.) The signaling markers showed a re-distribution to opposite sides of the cell during the
reversals, as did LimE. We also monitored the microtubules and centrosomes and tracked their
redistribution during polarity re-establishment using TACC. Further experiments were performed
in the absence of an actin cytoskeleton and in a variety of signaling mutants. These experiments
have helped elucidate the molecular mechanisms that contribute to the establishment and
maintenance of cell polarity.
2052
Alteration of α6 integrin expression and chemotactic motility of microglia upon ADP
stimulation.
S-H. Lee 1 , C. Y. Chung 2 ; 1 Pharmacology, Vanderbilt University Medical Center, Nashville, TN,
2 Vanderbilt University Medical Center, Nashville, TN
Microglia are the immune effector cells that are rapidly activated in response to even minor
pathological changes in the central nervous system. It has been demonstrated that microglia