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Titolo:
A verotoxin 1 B subunit-lambda CRO chimeric protein specifically binds both DNA and globotriaosylceramide (Gb(3)) to effect nuclear targeting of exogenous DNA in Gb(3) positive cells
Autore:
Facchini, LM; Lingwood, CA;
Indirizzi:
Univ Toronto, Hosp Sick Children, Div Infect Immun Injury & Repair, Res Inst, Toronto, ON M5G 1X8, Canada Univ Toronto Toronto ON Canada M5G 1X8 Inst, Toronto, ON M5G 1X8, Canada Univ Toronto, Dept Lab Med & Pathol, Toronto, ON M5G 1X8, Canada Univ Toronto Toronto ON Canada M5G 1X8 athol, Toronto, ON M5G 1X8, Canada Univ Toronto, Dept Biochem, Toronto, ON M5G 1X8, Canada Univ Toronto Toronto ON Canada M5G 1X8 ochem, Toronto, ON M5G 1X8, Canada
Titolo Testata:
EXPERIMENTAL CELL RESEARCH
fascicolo: 1, volume: 269, anno: 2001,
pagine: 117 - 129
SICI:
0014-4827(20010910)269:1<117:AV1BSC>2.0.ZU;2-7
Fonte:
ISI
Lingua:
ENG
Soggetto:
COMPLEX CLASS-I; RECEPTOR-MEDIATED ENDOCYTOSIS; SHIGA-LIKE TOXIN-1; RETROGRADE TRANSPORT; ESCHERICHIA-COLI; ENDOPLASMIC-RETICULUM; CRYSTAL-STRUCTURE; GOLGI-COMPLEX; BACTERIOPHAGE-LAMBDA; PSEUDOMONAS EXOTOXIN;
Keywords:
retrograde transport; vesicular transport; receptor-mediated endocytosis; glycolipid; membrane translocation; confocal microscopy;
Tipo documento:
Article
Natura:
Periodico
Settore Disciplinare:
Life Sciences
Citazioni:
58
Recensione:
Indirizzi per estratti:
Indirizzo: Lingwood, CA Univ Toronto, Hosp Sick Children, Div Infect Immun Injury & Repair, Res Inst, 555 Univ Ave, Toronto, ON M5G 1X8, Canada Univ Toronto 555Univ Ave Toronto ON Canada M5G 1X8 8, Canada
Citazione:
L.M. Facchini e C.A. Lingwood, "A verotoxin 1 B subunit-lambda CRO chimeric protein specifically binds both DNA and globotriaosylceramide (Gb(3)) to effect nuclear targeting of exogenous DNA in Gb(3) positive cells", EXP CELL RE, 269(1), 2001, pp. 117-129

Abstract

Inefficient nuclear incorporation of foreign DNA remains a critical roadblock in the development of effective nonviral gene delivery systems. DNA delivered by traditional protocols remains within endosomal/lysosomal vesicles, or is rapidly degraded in the cytoplasm. Verotoxin I (VT), an AB(5) subunit toxin produced by enterohaemorrhagic Escherichia coli, binds to the cellsurface glycolipid, globotriaosylceramide (Gb(3)) and is internalized intopreendosomes. VT is then retrograde transported to the Golgi, endoplasmic reticulum. (ER), and nucleus of highly VT-sensitive cells. We have utilizedthis nuclear targeting of VT to design a unique delivery system which transports exogenous DNA via vesicular traffic to the nucleus. The nontoxic VT binding subunit (VTB) was fused to the lambda Cro DNA-binding repressor, generating a 14-kDa VTB-Cro chimera. VTB-Cro binds specifically via the Cro domain to a 25-bp DNA fragment containing the consensus Cro operator. VTB-Cro demonstrates simultaneous specific binding to Gb(3). Treatment of Vero cells with fluorescent-labeled Cro operator DNA in the presence of VTB-Cro, results in DNA internalization to the Golgi, ER, and nucleus, whereas fluorescent DNA alone is incorporated poorly and randomly within the cytoplasm. VTB-Cro mediated nuclear DNA transport is prevented by brefeldin A, consistent with Golgi/ER intracellular routing. Pretreatment with filipin had no effect, indicating that caveoli are not involved. This novel VTB-Cro shuttle protein may find practical applications in the fields of intracellular targeting, gene delivery, and gene therapy. (C) 2001 Academic Press.

ASDD Area Sistemi Dipartimentali e Documentali, Università di Bologna, Catalogo delle riviste ed altri periodici
Documento generato il 20/01/21 alle ore 12:43:33