The objectives of this research are threefold. First, to develop the analysis of phospholipid vesicle fusion at two levels: (i) Kinetic - to ascertain which step of the overall process is rate limiting and (ii) Structural - identifying the intermembrane intermediates involved either during or subsequent to fusion. The effects of lipid composition,cation binding temperature and liposome size on the destabilization and fusion of these liposomes is being developed into a rigorous biophysical theory of membrane interactions. This theory is centered upon relating equilibrium thermodynamic properties of the lipids, e.g., phase transitions and morphological changes, to the initial interaction and destabilization of apposed bilayers. Our second objective is to enlarge our study of cytoplasmic fusion agents using liposomes as target membranes. The protein synexin and the polyamine spermine have both been implicated as mediators of intracellular fusion events involved in exocytosis. Our preliminary data show that both these agents act on liposomal membranes solely by enhancing aggregation rates, i.e., the rate of close approach. The destabilization of the apposed bilayers requires another factor, e.g., bound Ca2+. We will establish the generality of this result with other membranes. Of particular interest here is elucidating the effect of triphosphoinositides on membrane fusion in the presence of the polyamines. Our third objective is to begin studies on biological fusion proteins, specifically hemagglutinin from the influenza virus and the (putative) myoblast fusion protein. For hemagglutinin, we will focus on the role of cell surface receptors on the fusion mechanism using intact virus and reconstituted systems, virosomes, which we will develop into a reliable method for delivery of macromolecules into cells. For the myoblasts, we will focus on the ability of muscle cell vesicles, secreted during active myoblast fusion, to induce the fusion of a cloned line of nonfusing myoblasts.

Agency
National Institute of Health (NIH)
Institute
National Institute of General Medical Sciences (NIGMS)
Type
Research Project (R01)
Project #
5R01GM031506-09
Application #
3279554
Study Section
Biophysical Chemistry Study Section (BBCB)
Project Start
1982-04-01
Project End
1993-06-30
Budget Start
1991-07-01
Budget End
1992-06-30
Support Year
9
Fiscal Year
1991
Total Cost
Indirect Cost
Name
Drexel University
Department
Type
Schools of Arts and Sciences
DUNS #
061197161
City
Philadelphia
State
PA
Country
United States
Zip Code
19104
Shangguan, T; Siegel, D P; Lear, J D et al. (1998) Morphological changes and fusogenic activity of influenza virus hemagglutinin. Biophys J 74:54-62
Shangguan, T; Alford, D; Bentz, J (1996) Influenza-virus-liposome lipid mixing is leaky and largely insensitive to the material properties of the target membrane. Biochemistry 35:4956-65
Goldfine, H; Knob, C; Alford, D et al. (1995) Membrane permeabilization by Listeria monocytogenes phosphatidylinositol-specific phospholipase C is independent of phospholipid hydrolysis and cooperative with listeriolysin O. Proc Natl Acad Sci U S A 92:2979-83
Nieva, J L; Nir, S; Muga, A et al. (1994) Interaction of the HIV-1 fusion peptide with phospholipid vesicles: different structural requirements for fusion and leakage. Biochemistry 33:3201-9
Alford, D; Ellens, H; Bentz, J (1994) Fusion of influenza virus with sialic acid-bearing target membranes. Biochemistry 33:1977-87
Nir, S (1993) Kinetics and extent of fusion of viruses with target membranes. Methods Enzymol 220:379-91
Duzgunes, N; Pedroso de Lima, M C; Stamatatos, L et al. (1992) Fusion activity and inactivation of influenza virus: kinetics of low pH-induced fusion with cultured cells. J Gen Virol 73 ( Pt 1):27-37
Bentz, J (1992) Intermediates and kinetics of membrane fusion. Biophys J 63:448-59
Pedroso de Lima, M C; Nir, S; Flasher, D et al. (1991) Fusion of Sendai virus with human HL-60 and CEM cells: different kinetics of fusion for two isolates. Biochim Biophys Acta 1070:446-54
Ellens, H; Bentz, J; Mason, D et al. (1990) Fusion of influenza hemagglutinin-expressing fibroblasts with glycophorin-bearing liposomes: role of hemagglutinin surface density. Biochemistry 29:9697-707

Showing the most recent 10 out of 31 publications