TY - JOUR
T1 - Characterization of the Mr 65,000 lymphokine-activated killer proteins phosphorylated after tumor target binding
T2 - Evidence that pp65a and pp65b are phosphorylated forms of L-plastin
AU - Frederick, Mitchell J.
AU - Rodriguez, Lewis V.
AU - Johnston, Dennis A.
AU - Darnay, Bryant G.
AU - Grimm, Elizabeth A.
PY - 1996/1/1
Y1 - 1996/1/1
N2 - Contact between lymphokine-activated killer (LAK) cells and natural killer-resistant tumor targets SK-Mel-1 (human melanoma) or Raji (human lymphoma) stimulates phosphorylation of two Mr 65,000 LAK proteins (pp65a and pp65b) with nearly identical isoelectric points. Phosphoamino acid analysis of pp65a and pp65b detected phosphorylation exclusively on serine residues. Phosphotyrosine could not be detected on either substrate after immunoblotting with an antiphosphotyrosine antibody, and herbimycin A treatment failed to inhibit p65 phosphorylation induced by target contact. However, phorbol myristate acetate treatment alone induced LAK pp65a and pp65b phosphorylation, suggesting phosphorylation may be mediated by protein kinase C or a protein kinase C-regulated kinase. The molecular weight and isoelectric points of pp65a and pp65b are similar to that reported for the human actin-bundling protein, L-plastin (L-fimbrin). On two-dimensional SDS-PAGE gel immunoblots, a peptide specific anti-L-plastin antiserum bound to pp65a and pp65b, suggesting that the phosphoproteins are similar or identical to L-plastin. In addition, two adjacent Mr 65,000 LAK proteins were also detected by the antiserum and may correspond to unphosphorylated forms of L-plastin. On the basis of known properties of phosphorylated L-plastin, it is hypothesized that p65 phosphorylation in LAKs may regulate adhesion to tumor targets.
AB - Contact between lymphokine-activated killer (LAK) cells and natural killer-resistant tumor targets SK-Mel-1 (human melanoma) or Raji (human lymphoma) stimulates phosphorylation of two Mr 65,000 LAK proteins (pp65a and pp65b) with nearly identical isoelectric points. Phosphoamino acid analysis of pp65a and pp65b detected phosphorylation exclusively on serine residues. Phosphotyrosine could not be detected on either substrate after immunoblotting with an antiphosphotyrosine antibody, and herbimycin A treatment failed to inhibit p65 phosphorylation induced by target contact. However, phorbol myristate acetate treatment alone induced LAK pp65a and pp65b phosphorylation, suggesting phosphorylation may be mediated by protein kinase C or a protein kinase C-regulated kinase. The molecular weight and isoelectric points of pp65a and pp65b are similar to that reported for the human actin-bundling protein, L-plastin (L-fimbrin). On two-dimensional SDS-PAGE gel immunoblots, a peptide specific anti-L-plastin antiserum bound to pp65a and pp65b, suggesting that the phosphoproteins are similar or identical to L-plastin. In addition, two adjacent Mr 65,000 LAK proteins were also detected by the antiserum and may correspond to unphosphorylated forms of L-plastin. On the basis of known properties of phosphorylated L-plastin, it is hypothesized that p65 phosphorylation in LAKs may regulate adhesion to tumor targets.
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M3 - Article
C2 - 8548753
AN - SCOPUS:0030069967
SN - 0008-5472
VL - 56
SP - 138
EP - 144
JO - Cancer Research
JF - Cancer Research
IS - 1
ER -