title: Induction of protein expression within Escherichia coli vector for entry into mammalian cells.
abstract: Direct protein delivery into the cytosol of mammalian cells by invasive Escherichia coli (E. coli) bacterial vector will bypass the need to achieve nuclear entry and transcription of DNA, a major hurdle that is known to seriously limit gene transfer. The bacterial vector is induced to express the protein during its growth phase, before presentation for entry into mammalian cells and release of its content into the cellular environment. For this class of vector, crossing the plasma membrane becomes the primary step that determines the success of protein delivery. Yet, how the mechanics of protein expression within the vector affect its entry into the host is poorly understood. We found the vector's effectiveness to enter HeLa cells diminished together with its viability when phage N15 protelomerase (TelN) expression was induced continuously in the invasive E. coli despite producing an abundant amount of functional protein. By comparison, shorter induction, even as little as 3 hr, produced sufficient amounts of functional TelN and showed more effective invasion of HeLa cells, comparable to that of uninduced invasive E. coli. These results demonstrate that brief induction of protein expression during vector growth is essential for optimal entry into mammalian cells, an important step for achieving bacteria-mediated protein delivery.
authors.affiliation: 1 School of Science, Monash University , Bandar Sunway 46150, Malaysia .
authors.forename: Qingwen
authors.initials: Q
authors.lastname: Chen
authors.forename: Choon-Weng
authors.initials: CW
authors.lastname: Lee
authors.forename: Edmund Ui-Hang
authors.initials: EU
authors.lastname: Sim
authors.forename: Kumaran
authors.initials: K
authors.lastname: Narayanan
mesh_terms: D004792:Enzyme Precursors / Q000235:genetics* / Q000378:metabolism; D004926:Escherichia coli / Q000235:genetics* / Q000378:metabolism; D015870:Gene Expression / Q000187:drug effects; D018014:Gene Transfer Techniques; D006367:HeLa Cells; D006801:Humans; D010957:Plasmids / Q000235:genetics / Q000378:metabolism*; D019098:Telomerase / Q000235:genetics* / Q000378:metabolism; D013752:Tetracycline / Q000494:pharmacology; D013997:Time Factors; D014764:Viral Proteins / Q000235:genetics* / Q000378:metabolism
chemical_list: D004792:Enzyme Precursors; D014764:Viral Proteins; C411662:protelomerase protein, Bacteriophage N15; D019098:Telomerase; D013752:Tetracycline