Figure 3

Loss of Nkx2.2 affects expression of known endocrine transmembrane proteins, Tmem27, Adra2a, and a novel tetraspanin, Tm4sf4. qRTPCR of (A) Tmem27, (L) Adra2a, and (O) Tm4sf4 mRNA transcript levels were determined at e13.5, e15.5, and e18.5 and all genes compared wild type (grey bars) and Nkx2.2^-/- pancreata (black bars). In situ hybridization comparing wild type and Nkx2.2^-/- pancreas at e12.5, e14.5, and e17.5 of Tmem27 (B, C, D, G, H, and I), e14.5 of Adra2a (M and N), and e12.5, e14.5, and e17.5 of Tm4sf4 (P, Q, R, U, V, and W). Error bars represent SEM and asterisks denote statistical significance < 0.05. To assess co-expression, immunofluoresence staining of Tmem27 (E and J;green) and Ghrelin (F and K;copper) was performed on adjacent sections. Similarly, adjacent sections to the Tm4sf4 in situ hybridizations were stained for ghrelin (T and Y;green) and overlayed (T and Y: inset 2× zoom). DAPI (blue) indicates nuclei. Magnification 20× for e12.5, e14.5, R, S, T, W, X, Y, while 40× for E, F, J, and K. Large black dashed lines outline the pancreas at e12.5 and e14.5. Small solid white lines outline Tmem27 and ghrelin co-expression areas. Dashed white lines outline Tmem27+ cells that do not co-express ghrelin.