Rapid nuclear transit and impaired degradation of amyloid beta and glypican-1-derived heparan sulfate in Tg2576 mouse fibroblasts.

Anhydromannose (anMan)-containing heparan sulfate (HS) derived from S-nitrosylated glypican-1 is generated in endosomes by an endogenously or ascorbate induced S-nitrosothiol-catalyzed reaction. Expression and processing of amyloid precursor protein (APP) is required to initiate formation and endosome-to-nucleus translocation of anMan-containing HS in wild-type mouse embryonic fibroblasts (WT MEF). HS is then transported to autophagosomes and finally degraded in lysosomes. To investigate how APP-derived amyloid beta peptide (Aβ) affects intracellular trafficking of HS we have studied nuclear transit as well as autophagosome/lysosome targeting and degradation in Tg2576 MEF which produce increased amounts of Aβ. Deconvolution immunofluorescence microscopy with an anMan-specific monoclonal antibody showed anMan-staining in the nuclei of Tg2576 MEF after 5 min of ascorbate treatment and after 15 min in WT MEF. There was also greater nuclear accumulation of HS in Tg2576 MEF as determined by (35)S-sulfate labeling experiments. Tg2576 MEF was less sensitive to inhibition of NO production and copper-chelation than WT MEF. By using APP- and Aβ-recognizing antibodies we observed nuclear translocation of Aβ peptide in Tg2576 MEF but not in WT MEF. HS remained in the nucleus of WT MEF for at least 8 h and was then transported to autophagosomes. By 8 h HS had disappeared from the nuclei of Tg2576 MEF but colocalized poorly with the autophagosome marker LC3. Aβ also disappeared rapidly from the nuclei of Tg2576 MEF. Initially it appeared in acidic vesicles and later it accumulated extracellularly. Thus, in Tg2576 MEF there is nuclear accumulation as well as secretion of Aβ and impaired degradation of HS.