Nerve Growth Factor-Mediated Regulation of Low Density Lipoprotein Receptor-Related Protein Promoter Activation

Abstract

The mechanisms by which nerve growth factor (NGF) increases the level of low density lipoprotein receptor-related protein (LRP1) are not known. Administration of nitric oxide synthase (NOS) inhibitors modulates several of the neurotrophic actions of NGF, including TrkA signalling pathway activation, increases in gene expression and neurite outgrowth. The present study investigated whether NGF regulates the transcription of LRP1 as well as the role of NO and the individual TrkA signalling pathways in this action of NGF. PC12 cells were transfected with luciferase reporter constructs containing various sized fragments of the LRP1 promoter and treated with NGF (50 ng/mL) to establish whether NGF altered LRP transcription. NGF significantly increased luciferase activity in all LRP1 promoter construct-transfected cells with the NGF-responsive region of the promoter identified to be present in the first 1000 bp. The non-selective NOS inhibitor N(ω)-nitro-L-arginine methylester (L-NAME; 20 mM) had no effect on the NGF-mediated increase in luciferase activity, while the inducible NOS selective inhibitor s-methylisothiourea (S-MIU; 2 mM) attenuated the NGF-induced activation of the LRP1 promoter. Pretreatment of PC12 cells with 10 μM bisindolylmaleimide 1 (BIS-1) prevented the NGF-mediated increase in LRP1 promoter activation while 50 μM U0126 partially inhibited this response. In combination with S-MIU, all of the TrkA signalling pathway inhibitors blocked the ability of NGF to increase LRP1 transcription. These data suggest the NGF-mediated increase in LRP1 levels occurs, at least in part, at the level of transcription and that NO and the TrkA signalling pathways cooperate in the modulation of LRP1 transcription.