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

SUNDAY
the SNARE protein Vamp-7. Lysosome secretion allows the extracellular release of proteases,
whose activities promote the extraction of the immobilized Ag. We further show that local reorganization
of cortical actin by type I Myosins, which link the cortex to the plasma membrane,
is required for lysosome exocytosis and Ag extraction. Remarkably, while the short-tail Myosin
IC is recruited at the synapse and facilitates vesicle secretion and Ag uptake, the long-tail
Myosin IE negatively regulates both processes. These results suggest that the two class I
Myosins play antagonistic roles in the local reorganization of cortical actin for vesicle secretion
and Ag uptake. The B cell synapse therefore emerges as a highly specialized site where tightly
regulated exocytic and endocytic events take place thanks to the local shaping of the
membrane-cytoskeleton interface by class I Myosins
1976
Determination of the Subcellular, Surface, and Extracellular Localization of Hsp70s in
Mammalian Cells.
R. Medina* 1 , M. Rashedan* 1 , N. Nikolaidis 1 ; 1 Biological Science, California State University,
Fullerton, Fullerton, CA
70-kD Heat shock proteins (Hsp70s) are a family of molecular chaperones that play essential
roles in stress response by promoting protein homeostasis. Members of this family are primarily
localized in different subcellular compartments, including the cytosol, the endoplasmic reticulum,
and the mitochondria. Apart from their primary location in specific parts of the cell, different
Hsp70s have also been found in other places within the cell, at the cellular membrane, and at
the extracellular milieu. Although a few stresses and pathophysiological conditions, like cancer,
have been related with the re-localization of Hsp70s within the cell, their translocation to the
membrane, and their secretion from viable cells, the majority of these conditions remain
unknown. Additionally, the signaling mechanisms involved in the trafficking of intracellular
Hsp70s remain elusive. To this end we studied the intra-, membrane- and extra-cellular
localization of four members of the Hsp70 family. Specifically, we determined the re-localization
of HSPA1A and HSPA8, primarily localized in the cytosol, HSPA5, primarily localized in the
endoplasmic reticulum, and HSPA9, primarily localized in the mitochondria. Human embryonic
cells treated with different stressors, including heat and ethanol, or untreated were subjected to
sub-cellular fractionation and the presence of Hsp70s was determined by Western. The amount
of Hsp70s in the different fractions was quantified and normalized for equal loads using fractionspecific
antibodies, e.g., beta-actin for the cytosolic fraction. These experiments revealed that
under normal growth conditions all four proteins were present in the nuclear, cytosolic,
mitochondrial, and membrane fractions, as well as in the extracellular medium. In heat-shocked
cells, although all proteins were still present in all fractions their amounts in each fraction were
significantly different from the untreated cells. These results strongly suggest that after stress
the different Hsp70s are being re-localized within the cell, are anchored at the cellular
membrane, and are secreted from the cell. Additional experiments using pharmacological
manipulation of intracellular trafficking pathways are currently being performed to identify the
molecular mechanism used by Hsp70s to achieve their translocation and subsequent secretion.
This study is the first step towards elucidating the biological importance of the relocalization and
membrane occurrence of these chaperones.
*equal contribution