A cell-penetrating peptide derived from mammalian cell uptake protein of Mycobacterium tuberculosis.
A Mycobacterium tuberculosis membrane protein called Mycobacterium cell entry protein (Mce1A) was previously shown to mediate the uptake of nonpathogenic Escherichia coli and latex beads by nonphagocytic mammalian cells. Here we characterize further the in vitro invasive activity of Mce1A using colloidal gold nanoparticles and fluorescent latex microspheres. Mce1A-coated colloidal gold particles induced plasma membrane invagination and entered membrane-bound compartments inside HeLa cells. Few of the protein-coated particles were also found in the cytosol compartment. Cytochalasin D and nocodazole inhibited the uptake by HeLa cells, indicating that rearrangement of both microtubules and microfilaments was necessary for the uptake. The functional domain of Mce1A for invasion was narrowed to a highly basic 22-amino acid sequence termed Inv3. A synthetic Inv3 peptide stimulated uptake of colloidal gold particles as well as latex microspheres by HeLa cells. A chimeric protein composed of Inv3 sequence at the N terminus of beta-galactosidase appeared to stain the nuclear membrane, suggesting that it entered the HeLa cell cytoplasm. These observations suggest that the cell uptake activity of Mce1A is confined to a small peptide domain located in the core region of the protein. Inv3 could be used to ferry any protein in fusion with it into mammalian cells and may serve as a potent nonviral delivery system.
Lu
Sangwei
S
Program in Infectious Diseases and Immunity, School of Public Health, University of California, Berkeley, Berkeley, 94720, USA.
Tager
Leah A
LA
Chitale
Sadhana
S
Riley
Lee W
LW
D000595:Amino Acid Sequence / Q000235:genetics; D000942:Antigens, Bacterial / Q000235:genetics; D001426:Bacterial Proteins / Q000378:metabolism / Q000493:pharmacokinetics*; D002462:Cell Membrane / Q000378:metabolism* / Q000648:ultrastructure; D020099:Coated Materials, Biocompatible; D015638:Cytochalasin D / Q000494:pharmacology; D003598:Cytoskeletal Proteins / Q000037:antagonists & inhibitors; D004337:Drug Carriers; D004705:Endocytosis / Q000187:drug effects / Q000502:physiology; D017979:Gold Colloid; D006367:HeLa Cells; D006801:Humans; D008863:Microspheres; D050506:Mutant Chimeric Proteins / Q000378:metabolism / Q000493:pharmacokinetics; D014677:Pharmaceutical Vehicles; D017434:Protein Structure, Tertiary / Q000502:physiology*; D021381:Protein Transport / Q000187:drug effects; D011994:Recombinant Proteins; D013329:Structure-Activity Relationship
D000942:Antigens, Bacterial; D001426:Bacterial Proteins; D020099:Coated Materials, Biocompatible; D003598:Cytoskeletal Proteins; D004337:Drug Carriers; D017979:Gold Colloid; D050506:Mutant Chimeric Proteins; D014677:Pharmaceutical Vehicles; D011994:Recombinant Proteins; C473802:mammalian cell entry protein Mce1A, Mycobacterium tuberculosis; D015638:Cytochalasin D