RNA sequencing of P1 and P14 enriched neurons from wildtype and Zmiz1-KO cortex
Purpose: Zinc Finger MIZ-Type Containing 1 (Zmiz1) is a member of the PIAS family of protein and function as a transcriptional coactivator of Notch, Androgen Receptor (AR), p53, Estrogen Receptor (ER), and Smad3/4 . Despite Zmiz1 mutations association with neurodevelopmental disorders such as ASD, ADHD, and intellectual disability, its role in physiological and pathological neurodevelopment  is significantly unknown. Here, we use murine model to knockout Zmiz1 using Emx1Cre and performed neuron enrichment on P1 and P14 cortex followed by RNA sequencing to profile transcriptional changes upon Zmiz1 deletion. Mice was euthanized using ice (P1) or CO2 (P14) and brain were extracted at P1 or P14. Cortex was dissected and dissociated into single cell suspension using Neural Tissue Dissociation Kit (P) (Miltenyi Biotec, 130-092-628 (P1) or 130-107-677 (P14)) following manufacture instruction. Neurons were magnetically isolated/enriched by negative selection through depletion of non-neuronal cell types using  cocktail of antibodies for non-neuronal cell types (Miltenyi Biotec, 130-115-390). Total RNA was extracted from wildtype and Zmiz1-KO samples. RNA concentration and RNA integrity number were determined. RNA library was prepared, quantified, and verified using TruSeq RNA Library Prep Kit v2, Qubit dsDNA High Sensitivity Assay kit and Bioanalyzer DNA1000 assay kit respectively. Verified samples were sequenced using the NextSeq1000/2000 P2 Reagents (200 Cycles) v3 on a Nextseq1000/2000. RNA-seq data analysis was performed using illumina BaseSpace Sequence Hub. Briefly, sequenced reads were aligned to mouse (mm10) reference genome with RNA-Seq alignment tool (STAR aligner) and differentially expressed genes (DEG) were determined using the RNA-Seq Differential Expression tool (version 1.0.1). Results: For P1 enriched neurons, we found 291 differentially expressed genes of which 124 genes were upregulated while 167 genes were downregulated. Downregulated genes were enriched in biological processes such as neuron development, central nervous system differentiation, and axon development. For P14 enriched neurons,  we found 2,116 differentially expressed genes of which 749 genes were upregulated while 1,367 genes were downregulated. Downregulated genes were enriched in biological processes such as synaptic signalling and organization and neuron development and differentitation. Conclusions: We assessed P1 and P14 cortical neurons transcriptional landscape following Zmiz1 deletion in the cortex. Overall design: P1 and P14 enriched neurons from WT and Zmiz1-KO cortex RNA sequencing