MEDIATION OF NGF-STIMULATED EXTRACELLULAR-MATRIX INVASION BY THE HUMAN-MELANOMA LOW-AFFINITY P75 NEUROTROPHIN RECEPTOR - MELANOMA P75 FUNCTIONS INDEPENDENTLY OF TRKA

Although overexpression of the low-affinity p75 neurotrophin receptor (p75(NTR)) is, frequently associated with advanced stages of human melanoma progression, the functional significance of this finding is unknown. We examined whether the degree of cell surface expression of p75(NTR) in, human melanoma cell variants determines their extent of invasion stimulated by nerve growth factor (NGF). Treatment of MeWo melanoma cells or a metastatic spontaneous wheat germ agglutinin-resistant variant subline (70W) of MeWo cells with 2.5S NGF resulted in a dose-dependent enhancement of invasion through a reconstituted basement membrane. This effect was most pronounced with the 70W subline that exhibits brain-metastasizing potential in nude mice but was not found with a poorly metastatic MeWo variant subline (3S5). The expression of p75(NTR) as determined by Northern blotting and immunoprecipitation analysis of I-125-labeled cell surface proteins correlated with NGF-stimulated invasion. The MeWo melanoma sublines used in this study did not express p140(proto-trkA) mRNA or any p140(proto-trkA) variant transcripts including p70(trkA) as determined by Northern analysis and RT-PCR analysis. Thus, these melanoma cells would not be expected to form functional p75-p140 heterodimers or p140-p140 homodimers capable of transducing an NGF-generated signal to p140(proto-trkA) cytoplasmic substrates. These cells did express authentic p145(trkC) transcripts. However, NGF did not catalytically activate p145(trkC) receptors via increased tyrosine phosphorylation as would be expected if p145(trkC) participated in the signaling established by NGF. Furthermore, a NGF-stimulated purine-analogue-sensitive kinase activity was found to coimmunoprecipitate with p75(NTR). This p75(NTR)-associated kinase may coordinate initial signaling events evoked by p75(NTR) ligand interaction. Addition of 2.5S NGF, at concentrations that should saturate cell surface p75(NTR) to matrix-adherent cultures of human MeWo and 70W but not 3S5 melanoma cells suppressed the expression of 92-kDa type IV collagenase and stimulated the production of 72-kDa type IV collagenase in its fully active 68-kDa form. In the absence of p140(proto-trkA), the matrix-dependent effects of NGF on metalloproteinase expression of brain-metastatic 70W melanoma cells suggest a signaling role for the low-affinity melanoma p75(NTR) receptor and its associated purine-analogue-sensitive kinase in signaling enhanced matrix penetration of NGF-rich stromal microenvironments such as the brain.