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Testing effects of cell density on NGN2 iN transcriptom

ABSTRACT: Testing effects of cell density on NGN2 iN transcriptom

PROVIDER: PRJNA1142395 | ENA |

REPOSITORIES: ENA

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			Action	DRS
	SRR30049200.fastq.gz	Fastqsanger.gz
	SRR30049201.fastq.gz	Fastqsanger.gz
	SRR30049196.fastq.gz	Fastqsanger.gz
	SRR30049197.fastq.gz	Fastqsanger.gz
	SRR30049198.fastq.gz	Fastqsanger.gz

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Project description:Increased expression of G-coupled Inwardly Rectifying Potassium Channel Subunit 2 (GIRK2) has been linked to an electrophysiological endophenotype for alcohol use disorder. We have increased GIRK2 expression in human glutamatergic (NGN2-induced) neurons using two distinct strategies (CRISPRa targeting of the endogenous gene and ectopic expression with a lentiviral vector) to investigate the interplay between GIRK2 expression levels and neuronal response to alcohol (ethanol). We used six healthy donor cell lines (2 CRISPRa and 4 lenti) to draw comparisons within the same genetic background. Findings from functional data (calcium imaging, electrophysiology, and mitochondrial stress tests) indicate that increased GIRK2 expression, combined with prolonged (21-day) intermittent ethanol exposure (IEE), counteracts IEE-induced changes in glutamatergic activity and cellular respiration. To understand the transcriptomic underpinnings of these functional data, we conducted bulk RNAseq in the same donor cohort. Differential gene expression analysis indicates that both CRISPRa and lenti-increased GIRK2 interacted with IEE to produce unique differential regulation of synaptic genes, including glutamate receptors and voltage-gated ion channels, distinct from the effects of IEE or increased GIRK2 alone. Increased expression of G-coupled Inwardly Rectifying Potassium Channel Subunit 2 (GIRK2) has been linked to an electrophysiological endophenotype for alcohol use disorder. We have increased GIRK2 expression in human glutamatergic (NGN2-induced) neurons using two distinct strategies (CRISPRa targeting of the endogenous gene and ectopic expression with a lentiviral vector) to investigate the interplay between GIRK2 expression levels and neuronal response to alcohol (ethanol). We used six healthy donor cell lines (2 CRISPRa and 4 lenti) to draw comparisons within the same genetic background. Findings from functional data (calcium imaging, electrophysiology, and mitochondrial stress tests) indicate that increased GIRK2 expression, combined with prolonged (21-day) intermittent ethanol exposure (IEE), counteracts IEE-induced changes in glutamatergic activity and cellular respiration. To understand the transcriptomic underpinnings of these functional data, we conducted bulk RNAseq in the same donor cohort. Differential gene expression analysis indicates that both CRISPRa and lenti-increased GIRK2 interacted with IEE to produce unique differential regulation of synaptic genes, including glutamate receptors and voltage-gated ion channels, distinct from the effects of IEE or increased GIRK2 alone.

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Testing effects of cell density on NGN2 iN transcriptom

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