Witucki_2023_J.Alzheimers.Dis_95_1735

BACKGROUND: Bleomycin hydrolase (BLMH), a homocysteine (Hcy)-thiolactone detoxifying enzyme, is attenuated in Alzheimer's disease (AD) brains. Blmh loss causes astrogliosis in mice while the loss of histone demethylase Phf8, which controls mTOR signaling, causes neuropathy in mice and humans. OBJECTIVE: To examine how Blmh gene deletion affects the Phf8/H4K20me1/mTOR/autophagy pathway, amyloid-beta (Abeta) accumulation, and cognitive/neuromotor performance in mice. METHODS: We generated a new mouse model of AD, the Blmh-/-5xFAD mouse. Behavioral assessments were conducted by cognitive/neuromotor testing. Blmh and Phf8 genes were silenced in mouse neuroblastoma N2a-APPswe cells by RNA interference. mTOR- and autophagy-related proteins, and AbetaPP were quantified by western blotting and the corresponding mRNAs by RT-qPCR. Abeta was quantified by western blotting (brains) and by confocal microscopy (cells). RESULTS: Behavioral testing showed cognitive/neuromotor deficits in Blmh-/- and Blmh-/-5xFAD mice. Phf8 was transcriptionally downregulated in Blmh-/- and Blmh-/-5xFAD brains. H4K20me1, mTOR, phospho-mTOR, and AbetaPP were upregulated while autophagy markers Becn1, Atg5, and Atg7 were downregulated in Blmh-/- and Blmh-/-5xFAD brains. Abeta was elevated in Blmh-/-5xFAD brains. These biochemical changes were recapitulated in Blmh-silenced N2a-APPswe cells, which also showed increased H4K20me1-mTOR promoter binding and impaired autophagy flux (Lc3-I, Lc3-II, p62). Phf8-silencing or treatments with Hcy-thiolactone or N-Hcy-protein, metabolites elevated in Blmh-/- mice, induced biochemical changes in N2a-APPswe cells like those induced by the Blmh-silencing. However, Phf8-silencing elevated Abeta without affecting AbetaPP. CONCLUSIONS: Our findings show that Blmh interacts with AbetaPP and the Phf8/H4K20me1/mTOR/autophagy pathway, and that disruption of those interactions causes Abeta accumulation and cognitive/neuromotor deficits.