IMMUNOLOCALIZATION OF THIMET OLIGOPEPTIDASE IN CHICKEN EMBRYONIC FIBROBLASTS

An antiserum was raised against chicken thimet oligopeptidase (TOP), a nd specific antibodies were isolated by immunoadsorption. The specific ity of the antibodies was confirmed by immunoblot analysis with the ce ll extract of cultured chicken fibroblasts. Subcellular localization o f TOP in normal and Rous sarcoma virus-transformed chicken embryonic f ibroblasts was investigated by indirect immunofluorescence staining wi th affinity-purified, monospecific anti-TOP antibodies. In addition to the diffuse reaction expected for a cytosolic enzyme, a punctate, cyt oplasmic localization was observed. The appearance of the organelles c ontaining TOP did not correspond to that expected for secondary lysoso mes, mitochondria, endoplasmic reticulum, or Golgi apparatus, but did resemble that of endosomes. When cells were allowed to internalize Luc ifer yellow before fixation, some of the organelles containing TOP wer e observed to contain Lucifer yellow. Moreover, those organelles conta ining TOP were positively immunolabeled with the anti-transferrin rece ptor antibody. We conclude that part of the TOP in chicken embryonic f ibroblasts is located in endosomes and discuss the implications of thi s. (C) 1995 Academic Press, Inc.