Lateralized memory circuit dropout in Alzheimer's disease patients.

Altered connectivity within neuronal networks is often observed in Alzheimer's disease. However, delineating pro-cognitive compensatory changes from pathological network decline relies on characterizing network and task effects together. In this study, we interrogated the dynamics of occipito-temporo-frontal brain networks responsible for implicit and explicit memory processes using high-density EEG and dynamic causal modelling. We examined source-localized network activity from patients with Alzheimer's disease (n = 21) and healthy controls (n = 21), while they performed both visual recognition (explicit memory) and implicit priming tasks. Parametric empirical Bayes analyses identified significant reductions in temporo-frontal connectivity and in subcortical visual input in patients, specifically in the left hemisphere during the recognition task. There was also slowing in frontal left hemisphere signal transmission during the implicit priming task, with significantly more distinct dropout in connectivity during the recognition task, suggesting that these network drop-out effects are affected by task difficulty. Furthermore, during the implicit memory task, increased right frontal activity was correlated with improved task performance in patients only, suggesting that right-hemisphere compensatory mechanisms may be employed to mitigate left-lateralized network dropout in Alzheimer's disease. Taken together, these findings suggest that Alzheimer's disease is associated with lateralized memory circuit dropout and potential compensation from the right hemisphere, at least for simpler memory tasks.

An efficient and scalable pipeline for epitope tagging in mammalian stem cells using Cas9 ribonucleoprotein.

CRISPR/Cas9 can be used for precise genetic knock-in of epitope tags into endogenous genes, simplifying experimental analysis of protein function. However, Cas9-assisted epitope tagging in primary mammalian cell cultures is often inefficient and reliant on plasmid-based selection strategies. Here, we demonstrate improved knock-in efficiencies of diverse tags (V5, 3XFLAG, Myc, HA) using co-delivery of Cas9 protein pre-complexed with two-part synthetic modified RNAs (annealed crRNA:tracrRNA) and single-stranded oligodeoxynucleotide (ssODN) repair templates. Knock-in efficiencies of ~5-30%, were achieved without selection in embryonic stem (ES) cells, neural stem (NS) cells, and brain-tumor-derived stem cells. Biallelic-tagged clonal lines were readily derived and used to define Olig2 chromatin-bound interacting partners. Using our novel web-based design tool, we established a 96-well format pipeline that enabled V5-tagging of 60 different transcription factors. This efficient, selection-free and scalable epitope tagging pipeline enables systematic surveys of protein expression levels, subcellular localization, and interactors across diverse mammalian stem cells.