Neurofibrillary tangle is one of the hallmarks of Alzheimer's disease (AD) and is composed of paired helical filament (PHF) of cytosolic protein tau. Previously we reported the occurrence of N-glycosylation of PHF tau prepared from the AD brain and we found that structures of N-glycans of PHF tau were mainly high-mannose type (Sato et al. 2001 FEBS Lett. 496, 152-160). N-glycosylation of PHF tau was thought to be responsible for the maintenance of PHF structure (Wang et al. 1996 Nat. Med. 2, 871-875). These results suggest that AD may be related to aberrant N-glycosylation of tau and that cytosolic protein may be abnormally N-glycosylated in AD brain. Because the aging is one of the risk factors of AD, we examined whether N-glycosylated proteins of brain cytosol increased in the aged process. We analyzed glycoprotein-expression level in the brain soluble fraction by 2D-PAGE in combination with concanavalin A (Con A) staining. In cortex, it was found that eight spots in aged rat (30-month-old) were more reactive to Con A than in young-adult (9-week-old). Among them, reactivations of three spots in the aged-rat were also observed in hippocampus, cerebellum and spinal cord. These results suggested that the increase of Con A-reactivity of soluble proteins was the common phenomenon in the central nervous system. Because the reactivations were diminished by Endo H digestion, these glycoproteins had high-mannose type N-glycans. However, the appearance of cytosolic N-glycosylated proteins in the aged brain is unclear because N-glycosylation and its processing proceeded in the lumen of endoplasmic reticulum (ER) and Golgi apparatus. Recently it was reported that misfolded N-glycosylated proteins were degraded by ER associated degradation (ERAD) system. If the deterioration of ERAD occurred in the aged rat, N-glycosylated proteins may be accumulated in the cytosol. We are now trying to identify these accumulated glycoproteins.
