Multiple trkA proteins in PC12 cells bind NGF with a slow association rate.
Rat pheochromocytoma (PC12) cells express two distinct nerve growth factor receptors (NGFRs), p75NGFR and trkA (p140trk). In addition to these receptors, by using 125I-mNGF affinity labeling and BS3 chemical cross-linking of PC12 cell protein, we have identified two additional trkA protein bands with apparent molecular weights of 220,000 and 300,000. These bands contain trkA, but were not immunoprecipitated by p75NGFR-specific antisera, suggesting that they do not represent trkA/p75NGFR protein complexes. The 220-kDa trkA band apparently represents trkA with alternate post-translational modification. The appearance of the 300-kDa trkA band was dependent on cross-linker concentration and could be diminished in the presence of reducing agents, suggesting that it represents a trkA dimer. All trkA bands were phosphorylated on tyrosine residues when bound to mNGF, suggesting that they participate in NGF-induced signal transduction. NGF binding kinetics to all three trkA bands were indistinguishable, with slow dissociation rates, and a slow association rate that required approximately 1 h to reach equilibrium levels at 4 degrees C. All three trkA bands bound the related neurotrophin brain-derived neurotrophic factor and neurotrophin-3 with a profile characteristic of trkA.
D000818:Animals; D015152:Blotting, Northern; D007700:Kinetics; D008970:Molecular Weight; D009414:Nerve Growth Factors / Q000378:metabolism*; D016716:PC12 Cells; D011061:Poly A / Q000235:genetics / Q000302:isolation & purification; D011499:Protein Processing, Post-Translational; D011505:Protein-Tyrosine Kinases / Q000378:metabolism; D011518:Proto-Oncogene Proteins / Q000235:genetics / Q000302:isolation & purification / Q000378:metabolism*; D012313:RNA / Q000235:genetics / Q000302:isolation & purification; D012333:RNA, Messenger / Q000378:metabolism; D020917:Receptor, trkA; D017475:Receptors, Nerve Growth Factor / Q000302:isolation & purification / Q000378:metabolism*
D009414:Nerve Growth Factors; D011518:Proto-Oncogene Proteins; D012333:RNA, Messenger; D017475:Receptors, Nerve Growth Factor; D011061:Poly A; D012313:RNA; D011505:Protein-Tyrosine Kinases; D020917:Receptor, trkA