A pivotal role of cysteine 3 of Lck tyrosine kinase for localization to glycolipid-enriched microdomains and T cell activation

Citation
A. Kosugi et al., A pivotal role of cysteine 3 of Lck tyrosine kinase for localization to glycolipid-enriched microdomains and T cell activation, IMMUNOL LET, 76(2), 2001, pp. 133-138
Citations number
24
Language
INGLESE
art.tipo
Article
Categorie Soggetti
Immunology
Journal title
IMMUNOLOGY LETTERS
ISSN journal
0165-2478 → ACNP
Volume
76
Issue
2
Year of publication
2001
Pages
133 - 138
Database
ISI
SICI code
0165-2478(20010301)76:2<133:APROC3>2.0.ZU;2-Y
Abstract
Lek, a Src family protein tyrosine kinase (PTKs), is post-translationally m odified by palmitoylation, a process thought to regulate the biological fun ction, membrane affinity and glycolipid-enriched microdomain (GEM) localiza tion of this molecule. To examine the importance o palmitoylation sites Cys 3 and Cys5 in Lck, one or both of these residues was mutated to serine to c reate mutants S3, S5, and S3,5, respectively. Immunofluorescence and confoc al microscopy of COS-7 cells transfected with these constructs showed that while S5 and 53 localized to the plasma membrane, S3,5 was localized to the cytoplasm, suggesting that palmitoylation at at least one site is essentia l for membrane localization. Sucrose gradient based fractionation of these mutants expressed in COS-7 cells slowed that while S5 localized to GEMs in similar fashion to the wild type, GEM localization of 53 was severely inhib ited. Expression of these mutants in Lck-negative JCaM1 cells showed that a lthough Si reconstituted activation of nuclear factor. NFAT as per the wild type, S3 expression failed to do so. These results suggest that Cys3 of Lc k plays a more important role than Cys5 in GEM localization and T cell acti vation. Additionally, it was found that the degree of T cell function recov ery is positively correlated willi the degree of Lck expression in GEMs. (C ) 2001 Elsevier Science B.V. All rights reserved.