Phosphorylated Proteins of Mitogen Stimulated-Rat Peripheral Blood Lymphocytes

분열유발인자에 의한 흰쥐 림프구 단백의 인산화

  • Jou, I-Lo (Department of Pharmacology, Yonsei University College of Medicine) ;
  • Ko, Sung-Soo (Department of Pharmacology, Yonsei University College of Medicine) ;
  • Ahn, Young-Soo (Department of Pharmacology, Yonsei University College of Medicine)
  • 주일로 (연세대학교 의과대학 약리학교실) ;
  • 고성수 (연세대학교 의과대학 약리학교실) ;
  • 안영수 (연세대학교 의과대학 약리학교실)
  • Published : 1993.06.30

Abstract

This study was done to classify the proteins involved in the specific phosphorylation using the rat peripheral blood lymphocytes (rPBL) stimulated with mitogens, phorbol 12-myristate 13-acetate (PMA) and concanavalin A (Con A). The lymphocytes were incubated with $^{32}P-orthophosphate$ before PMA or Con A stimulation. The migration patterns of the phosphorylated proteins of mitogen-treated rPBL in two dimensional electrophoretic fields were analyzed after autoradiography. The stimulation of the lymphocytes with PMA and Con A increased the phosphorylation of thirteen protein fractions. The phosphorylation intensities of the protein spots differ to the treatments of the cells with specific kinase inhibitors, H-7 and W-7. These protein fractions were grouped into 3 classes, namely, PKC-mediated, CaM kinase-mediated, and other kinase mediated proteins. The effect of the duration of the stimulation on the phosphorylated behaviors occurred concurrently, not sequentially, although each individual protein fraction had a different time for the peak phosphorylation during the stimulation period upto 30 minutes. The phosphoproteins found in the cytosolic soluble fraction were phosphorylated prior to those in the pellet, whose phosphorylations were sustained at a high level for over 10 minutes. The above results suggest that the early events in lymphocyte activation involve 3 different sets of proteins which are phosphorylated by CaM kinase, PKC and other kinases, and those kinases do not work sequentially, but rather, independently or cooperatively.