VISUALIZATION OF INTRACELLULAR BACTERIAL INFECTION IN TUMOR CELL LINES

摘要

Listeria monocytogenes is a Gram~+ intracytoplasmic pathogen. Like other intracellular bacterial pathogens Listeria can adhere to, invade, and release toxins into eukaryotic cells. The basic virulence gene cluster of Listeria ←prfA ←plcA ― hly → mpl → actA → plcB → is separated on the chromosome from another operon also involved in pathogenicity ― inlA → inlB →. Both clusters together form a regulon mostly activated by PrfA (pleiotropic activator protein). Internalins InlA and InlB mediate the entry of Listeria into the host cell by attaching to the E-cadherin receptor. The pore-forming Hly (lysteriolysin O) synergistically operates with PlcA (phosphatidylinositol-specific phospholipase C) to penetrate from the vacuole or phagosome into the cytoplasm. After actA is expressed, the protein (ActA) dimerizes and induces a 'trail' of actin-polymerization which apparently propels the bacterium at ～0.3 μm/s through the cytoplasm into the adjacent host cell within a penetrating projection. This projection pinches off into the adjacent cell to form a double membrane vacuole, which is lysed by PlcB, another phospholipase C, releasing the bacterium into the cytoplasm of the adjacent cell. Listeria infection thus spreads from cell to cell. The function of Mpl, a zinc-dependent metalloprotease is currently unknown. Green fluorescent protein (GFP) from Aequorea victoria serves as an excellent reporter of virulence gene expression and intracellular infectivity of Listeria monocytogenes. The invasive pathogenic ability of Listeria has been exploited to deliver foreign protein products into the cytoplasm of host cells. Listeria infectivity is significant in light of recent findings in our laboratory using GFP and bacterial luciferase that both Gram~+ and Gram bacteria survive and replicate in tumors in murine models. The infectivities of various mammalian cells in culture by wildtype and attenuated strains of Listeria monocytogenes were tested in this study.