Howard Goldfine, Ph.D.

Professor of Microbiology

Office Address:
Department of Microbiology
Perelman School of Medicine
University of Pennsylvania
203D Johnson Pavilion
3610 Hamilton Walk
Philadelphia, PA 19104-6076
215-898-6384; FAX 215-573-4856
goldfinh@mail.med.upenn.edu

Dr. Goldfine has retired and is no longer accepting graduate students.

RESEARCH SUMMARY

The anaerobic biosynthetic pathway to ether lipid in bacteria
Recent work with other pathogenic clostridia including C. novyi has shown that the known intermediates in phospholipid biosynthesis in bacteria including phosphatidic acid, CDP-diacylglycerol, and phosphatidylserine in these organisms have no or only trace amounts of plasmalogens, whereas the end-products, phosphatidylethanolamine and phosphatidylglycerol, contain a large proportion of the plasmalogen species. These findings along with earlier work on the kinetics of phospholipid synthesis in clostridia have led to a proposed anaerobic pathway to plasmalogens in which the end-product diacyl phospholipids undergo transformation to plasmalogens. This pathway differs from that found in animal cells, which requires molecular oxygen. We are testing this hypothesis by means of experiments with anaerobic cell-free preparations.

Listeria monocytogenes
Interactions between bacterial pathogens and the mammalian host cell are at the intersection of microbiology and cell biology. Work in Dr.Goldfine's laboratory has been focused on two gram-positive pathogens, Listeria monocytogenes and Bacillus anthracis. L. monocytogenes (Lm) is a facultative intracellular pathogen that is a significant cause of human disease. In its initial interactions with a mammalian host cell, two phospholipases and listeriolysin O, a pore forming cytolysin, act together to initiate calcium and PKC signaling in the host cell. One of the phospholipases is a phosphatidylinositol (PI) specific phospholipase C (PI-PLC), which has been shown to play a role in escape of these bacteria from a primary endocytic vacuole. A second broad range phospholipase C (PC-PLC plays a significant role in cell-to-cell spread. Work in the Goldfine lab has shown that mutants in Lm PI-PLC are not complemented by introduction of the gene for PI-PLC from Bacillus cereus (Bc).or B. anthracis. We have previously shown that LmPI-PLC differs from Bacillus PI-PLC in having weak activity on proteins anchored to eukaryotic cell membranes by a glycosyl-PI modification at the C-terminus. It appears that LmPI-PLC has evolved for intracellular growth and cell-to-cell spread by the absence of a specific amino acid sequence needed for binding of the glycan chain of glycosyl-PI-anchored proteins prior to cleavage of the PI anchor.

Intracellular signaling
Work in this laboratory has shown that prior to internalization by a macrophage, L. monocytogenes induces a series of elevations of intracellular calcium levels in the host cytoplasm. Using isogenic mutants in the virulence factors, we have shown that listeriolysin O and the two phospholipases are required for the full sequence of calcium changes. These calcium elevations alter the kinetics of association of the bacterium with the host cell and the entry of bound bacteria into the host by phagocytosis. The current hypothesis is that these signaling events serve to modify the pathway of maturation of the phagosome so that the bacterium is able to escape from the primary vacuole and grow in the cytoplasm. Our more recent work has shown that PI-PLC and LLO cooperate to produce a protein kinase C cascade which is initiated by the generation of diacylglycerol upon hydrolysis of host phospholipids. This cascade leads to translocation of PKC beta I and II to an early endosome-like compartment. Recent work has implicated ActA, another virulence factor that promotes actin polymerization and intracellular motility, in escape from the primary phagosome in macrophages.