Transcription factor Creb3l1 maintains proteostasis in neuroendocrine cells [RNA-seq]

We have shown that the expression of the endoplasmic reticulum stress sensor Creb3l1 increases in magnocellular neurones (MCNs) in the rat hypothalamus in response to increased physiological demands for protein synthesis. Here we adopted a multiomic strategy to investigate specific roles of Creb3l1 in MCN homeostasis. We first performed chromatin immunoprecipitation followed by genome sequencing (ChIP-seq) to identify Creb3l1 genomic targets in the water deprived MCN enriched hypothalamic preparation. We then compared ChIP-seq gene targets with water deprived and Creb3l1 knockdown supraoptic nucleus RNA sequencing transcriptome datasets. This has provided an integrated signalling-gene regulation network for this transcription factor illuminating changes to cell pathways and function, an approach that has led us to understand the physiological changes that occur in MCNs to cope with excessive protein demands. This study was performed on samples collected from Sprague Dawley rats (n=5). Adeno-associated viruses were injected unilaterally into the supraoptic nuclei of the hypothalamus. The control virus was injected into the left supraoptic nucleus and the Creb3l1 knockdown virus into the right supraoptic nucleus of the same animal. A 1 mm punch (Fine Scientific Tools, 18035-01) was used to collect SON samples from 100 μm coronal slices sectioned in a cryostat set at -20°C. Total RNA was isolated from individual SONs (left and right) using phenol-chloroform phase separation method and Zymo Directzol RNA Miniprep kit.