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Titolo:
Precursor cystatin C in cultured retinal pigment epithelium cells: evidence for processing through the secretory pathway
Autore:
Paraoan, L; White, MRH; Spiller, DG; Grierson, I; Maden, BEH;
Indirizzi:
Univ Liverpool, Sch Biol Sci, Liverpool L69 7ZB, Merseyside, England Univ Liverpool Liverpool Merseyside England L69 7ZB , Merseyside, England Univ Liverpool, Dept Med, Unit Ophthalmol, Liverpool L69 3GA, Merseyside, England Univ Liverpool Liverpool Merseyside England L69 3GA , Merseyside, England
Titolo Testata:
MOLECULAR MEMBRANE BIOLOGY
fascicolo: 3, volume: 18, anno: 2001,
pagine: 229 - 236
SICI:
0968-7688(200107)18:3<229:PCCICR>2.0.ZU;2-5
Fonte:
ISI
Lingua:
ENG
Soggetto:
RAY CRYSTAL-STRUCTURE; POST-GAMMA-GLOBULIN; PHYSICOCHEMICAL CHARACTERIZATION; CYSTEINE PROTEINASES; BIOLOGICAL-FLUIDS; CATHEPSIN-B; IN-VITRO; TRACE; INHIBITORS; SEQUENCE;
Keywords:
cystatin C; EGFP-fusion protein; retinal pigment epithelium; Golgi apparatus; secretion;
Tipo documento:
Article
Natura:
Periodico
Settore Disciplinare:
Life Sciences
Citazioni:
34
Recensione:
Indirizzi per estratti:
Indirizzo: Paraoan, L Univ Liverpool, Sch Biol Sci, Life Sci Bldg,Crown St, LiverpoolL69 7ZB, Merseyside, England Univ Liverpool Life Sci Bldg,Crown St Liverpool Merseyside England L69 7ZB
Citazione:
L. Paraoan et al., "Precursor cystatin C in cultured retinal pigment epithelium cells: evidence for processing through the secretory pathway", MOL MEMBR B, 18(3), 2001, pp. 229-236

Abstract

Evidence was recently reported that the cysteine proteinase inhibitor, cystatin C, is highly expressed by cultured human retinal pigment epithelial (RPE) cells. As a step towards understanding possible functions of this protein associated with the RPE, the localization, targetting and trafficking of cystatin C were investigated. Constructs encoding an enhanced variant of green fluorescent protein (EGFP) fused to precursor cystatin C and to mature cystatin C were made and transfected into cultured human RPE cells. Expression of fusion proteins was monitored in vivo by fluorescence confocal microscopy. In cells transfected with precursor cystatin C-EGFP, fluorescence was initially targetted to the perinuclear zone, co-localizing with the Golgi apparatus. Transfected cells were observed at intervals over a period ofup to 3 weeks, during which time fluorescent vesicles developed peripherally and basally while fluorescence continued to be detected in the Golgi region. Immunochemical analysis of cell lysates confirmed the expression of a fusion protein recognized by antibodies to both cystatin C and EGFP. Cells transfected with the construct lacking the leader peptide of precursor cystatin C presented a diffuse and weak fluorescence. Together, these results imply a leader sequence-dependent processing of cystatin C through the secretory pathway of RPE cells. This was confirmed by the detection, by Western blotting, of the chimaeric protein alongside endogenous cystatin C in the medium of transfected RPE cells.

ASDD Area Sistemi Dipartimentali e Documentali, Università di Bologna, Catalogo delle riviste ed altri periodici
Documento generato il 04/04/20 alle ore 14:31:31