Arrest of Listeria movement in host cells by a bacterial ActA analogue: implications for actin-based motility.
Upon entering the host cell's cytoplasm, the pathogen Listeria monocytogenes can subvert the normal contractile system of the host cell; subsequent assembly of polar actin-filament structures is likely to provide the force for rapid intracellular bacterial movement and its cell-to-cell spread. We have now investigated the functional consequences of microinjecting Listeria-infected PtK2 cells with a synthetic peptide, CFEFPPPPTDE. This peptide represents one of four related oligoproline stretches in ActA, a bacterial surface protein necessary for Listeria-induced actin assembly. Over an estimated intracellular concentration range of 80 nM to 0.8 microM, this analogue rapidly blocks the formation of the actin-filament tails and arrests intracellular bacterial motility. Over the same time scale and concentration range, introduction of the ActA analogue also causes host cell membrane retraction. Bodipyphallacidin staining reveals that microinjection of the ActA analogue results in massive retraction of the actin cytoskeleton. Microinjection of 1-20 microM poly(L-proline) (intracellular concentration) fails to block Listeria intracellular movement or polar actin-filament assembly. As observed with ActA, however, poly(L-proline) does cause membrane retraction. Our findings demonstrate the efficacy of low molecular weight peptides in efforts to distinguish mechanistic features in Listeria motility and PtK2 host cell membrane reorganization. These observations also suggest that a cytoskeletal component sensitive to specific oligoproline peptides may participate in protein-protein interactions essential for both of these actin-associated processes.
Southwick
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Department of Medicine, University of Florida College of Medicine, Gainesville 32610-0277.
Purich
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D000199:Actins / Q000378:metabolism*; D000595:Amino Acid Sequence; D001426:Bacterial Proteins / Q000378:metabolism*; D001665:Binding Sites; D002465:Cell Movement; D003285:Contractile Proteins; D008089:Listeria monocytogenes / Q000166:cytology*; D008565:Membrane Proteins / Q000378:metabolism*; D008840:Microfilament Proteins / Q000378:metabolism; D008845:Microinjections; D008969:Molecular Sequence Data; D010446:Peptide Fragments / Q000138:chemical synthesis / Q000378:metabolism; D051304:Profilins; D011392:Proline / Q000378:metabolism
D000199:Actins; D001426:Bacterial Proteins; D003285:Contractile Proteins; D008565:Membrane Proteins; D008840:Microfilament Proteins; D010446:Peptide Fragments; D051304:Profilins; C072980:actA protein, Listeria monocytogenes; D011392:Proline