Wild type and tailless CD8 display similar interaction with microfilaments during capping.
We examined the influence of the intracytoplasmic region of CD8 alpha on capping and interaction with microfilaments. We used cell clones obtained by transfecting a CD4+ T-cell hybridoma with (a) T-cell receptor (TCR) alpha and beta chains from a cytolytic clone and (b) CD8 alpha genes that were either native or modified by extensive deletion of the intracytoplasmic region or replacement of the transmembrane and intracytoplasmic domains with those of a class I major histocompatibility complex gene (Letourneur et al. (1990). Proc. natn. Acad. Sci. U.S.A. 87, 2339-2343). Different cell surface structures were cross-linked with anti-T-cell receptor, anti-CD8 or anti-class I monoclonal antibodies and anti-immunoglobulin (Fab')2. Double labeling and quantitative image analysis were combined to monitor fluorescence anisotropy and correlation between different markers. Microfilaments displayed maximal polarization within two minutes. The correlation between these structures and surface markers was then maximal and started decreasing, whereas the redistribution of surface markers remained stable or continued. Furthermore, wild type and altered CD8 alpha exhibited similar ability to be capped and to induce co-capping of TCR and MHC (major histocompatibility complex) class I: the fraction of cell surface label redistributed into a localized cap ranged between 40% and 80%. Finally, cytochalasin D dramatically decreased CD8 capping in all tested clones. It is concluded that the transmembrane and/or intracellular domains of CD8 molecules are able to drive the extensive redistributions of membrane structures and cytoskeletal elements that are triggered by CD8 cross-linking.
Laboratoire d'Immunologie, Hôpital de Sainte-Marguerite, Marseille, France.
P
P
André
J
J
Gabert
A M
AM
Benoliel
C
C
Capo
C
C
Boyer
A M
AM
Schmitt-Verhulst
B
B
Malissen
P
P
Bongrand
D008841:Actin Cytoskeleton / Q000187:drug effects* / Q000276:immunology; D000199:Actins / Q000032:analysis; D000818:Animals; D015704:CD4 Antigens / Q000032:analysis / Q000276:immunology*; D015496:CD4-Positive T-Lymphocytes / Q000276:immunology*; D016827:CD8 Antigens / Q000032:analysis / Q000276:immunology*; D015638:Cytochalasin D / Q000494:pharmacology*; D005454:Fluorescence Polarization; D006680:HLA Antigens / Q000032:analysis; D006801:Humans; D006825:Hybridomas; D007091:Image Processing, Computer-Assisted; D007152:Immunologic Capping / Q000502:physiology*; D008565:Membrane Proteins / Q000276:immunology*; D014162:Transfection; D014404:Tubulin / Q000032:analysis
D000199:Actins; D015704:CD4 Antigens; D016827:CD8 Antigens; D006680:HLA Antigens; D008565:Membrane Proteins; D014404:Tubulin; D015638:Cytochalasin D