Translocation of spectrin and protein kinase C to a cytoplasmic aggregate upon lymphocyte activation.
We have previously reported that mammalian tissue lymphocytes exhibit significant heterogeneity with respect to the subcellular distribution of spectrin and that this phenomenon may result from a dynamic behavior of spectrin in response to activation signals. Here, we further characterize the involvement of spectrin in lymphocyte activation by examining its relationship with protein kinase C (PKC). PKC isoenzymes are a family of cytosolic kinases that translocate from the soluble to particulate fraction upon cell stimulation. It is reported here that activation of lymph node T cells through the antigen-specific receptor, or direct activation of PKC by phorbol esters, results in a striking increase in cells expressing a cytoplasmic aggregate of spectrin. Additionally, a concurrent increase in cells expressing aggregates of the beta II isozyme of PKC is observed. Immunofluorescence staining revealed that spectrin and PKC beta II are colocalized in untreated lymphocytes and that these two proteins are coincidently translocated to the same focal aggregate within the cytoplasm following stimulation. This redistribution of spectrin and PKC beta is blocked by pretreatment with calphostin C, a specific inhibitor of PKC. Solubility studies showed that there is an increase of both proteins in the detergent-insoluble fraction of lymphocytes upon activation, and immunoprecipitation studies indicated that the soluble form of these molecules may be associated directly or indirectly as part of a complex of proteins. These data indicate that the positioning of the spectrin-based cytoskeleton is sensitive to activation signals and may play a role in the function or positioning of PKC beta II.
Gregorio
C C
CC
Department of Molecular Immunology, Roswell Park Cancer Institute, Buffalo, NY 14263.
Kubo
R T
RT
Bankert
R B
RB
Repasky
E A
EA
D000818:Animals; D002451:Cell Compartmentation / Q000187:drug effects; D003599:Cytoskeleton / Q000378:metabolism*; D005455:Fluorescent Antibody Technique; D008213:Lymphocyte Activation; D008214:Lymphocytes / Q000648:ultrastructure*; D046911:Macromolecular Substances; D051379:Mice; D008807:Mice, Inbred BALB C; D009281:Naphthalenes; D011083:Polycyclic Compounds / Q000494:pharmacology; D011233:Precipitin Tests; D011485:Protein Binding; D011493:Protein Kinase C / Q000737:chemistry / Q000378:metabolism*; D012995:Solubility; D013049:Spectrin / Q000737:chemistry / Q000378:metabolism*
D046911:Macromolecular Substances; D009281:Naphthalenes; D011083:Polycyclic Compounds; D013049:Spectrin; D011493:Protein Kinase C; C058819:calphostin C