CRISPR screening by AAV episome-sequencing (CrAAVe-seq) is a highly scalable cell type-specific screening platform.

There is a significant need for scalable CRISPR-based genetic screening methods that can be applied directly in mammalian tissues while enabling cell type-specific analysis. To address this, we developed an adeno-associated virus (AAV)-based CRISPR screening platform, CrAAVe-seq, that incorporates a Cre-sensitive sgRNA construct for pooled screening within targeted cell populations in mouse tissues. We demonstrate the utility of this approach by screening two distinct large sgRNA libraries, together targeting over 5,000 genes, in mouse brains to create a robust profile of neuron-essential genes.

An amino-terminal fragment of apolipoprotein E4 leads to behavioral deficits, increased PHF-1 immunoreactivity, and mortality in zebrafish.

Although the increased risk of developing sporadic Alzheimer's disease (AD) associated with the inheritance of the apolipoprotein E4 (APOE4) allele is well characterized, the molecular underpinnings of how ApoE4 imparts risk remains unknown. Enhanced proteolysis of the ApoE4 protein with a toxic-gain of function has been suggested and a 17 kDa amino-terminal ApoE4 fragment (nApoE41-151) has been identified in post-mortem human AD frontal cortex sections. Recently, we demonstrated in vitro, exogenous treatment of nApoE41-151 in BV2 microglial cells leads to uptake, trafficking to the nucleus and increased expression of genes associated with cell toxicity and inflammation.

Fragmentation of Apolipoprotein E4 is Required for Differential Expression of Inflammation and Activation Related Genes in Microglia Cells.

The apolipoprotein E4 () allele represents the single greatest risk factor for late-onset Alzheimer's disease (AD) and accumulating evidence suggests that fragmentation with a toxic-gain of function may be a key molecular step associated with this risk. Recently, we demonstrated strong immunoreactivity of a 151 amino-terminal fragment of apoE4 (E4-fragment) within the nucleus of microglia in the human AD brain. In vitro, this fragment led to toxicity and activation of inflammatory processes in BV2 microglia cells.