Npas4 is expressed in β-cells. Compared with other PAS domain factors, Npas4 was highly expressed in freshly isolated mouse islets (n = 4) (A). Pancreatic Npas4 gene expression was induced late in development. Embryonic pancreatic tissue was dissected between e10.5 and e18.5, RNA was
extracted and reverse transcribed, and Taqman PCR was carried out for Npas4 (n = 3–4) (B). Npas4 was restricted to the endocrine pancreas (C and D). Confocal imaging (C) demonstrated that Npas4 (green) colocalized with DNA (blue; DAPI) within glucagon (red)-expressing cells. Widefield immunofluorescence
demonstrated that Npas4 (red) colocalized with both DNA (blue) and insulin (green) within pancreatic β-cells (D). There was significant variability in the percentage of cells showing Npas4 immunoreactivity within the nucleus in islets
within one pancreas, indicating that there is likely nuclear-cytoplasmic shuttling in response to metabolic demands. Scale
bars, 50 μm. Gcg, glucagon; RPKM, Reads Per Kilobase of transcript per Million mapped reads.