Transient transcriptional activation of gastrin during sodium butyrate-induced differentiation of islet cells

Transient expression of pancreatic gastrin corresponds to a period of rapid islet cell development. After birth gastrin expression silencing is coincidental with islet cell terminal differentiation, while persistent expression is accompanied with nesidioblastosis and reexpression observed in islet cell tumors. Experiments with transgenic animals suggested that gastrin might act synergistically with growth factors to stimulate islet cell development. The present study intended to establish an in vitro cell culture model to analyse the molecular events controlling gastrin gene activation and repression dependent on islet cell differentiation. Sodium butyrate, a proliferation-arresting compound has previously been shown to differentiate insulinoma cells while increasing insulin production. The present paper demonstrates concomitant transient increase in gastrin mRNA, intracellular and secreted gastrin during sodium butyrate treatment. Increased gastrin expression was due to activation or derepression of gastrin promoter activity as revealed by promoter analyses. This in vitro model mimics the expression pattern of gastrin and insulin observed during fetal islet cell development and provides an excellent tool to analyse the molecular mechanisms controlling gastrin gene activation and selective repression during islet cell differentiation.     

Modulation of Rac1 and ARF6 activation during epithelial cell scattering

Epithelial cell scattering encompasses the dissolution of intercellular junctions, cell-cell dissociation, cell spreading, and motility. The Rac1 and ARF6 GTPases have been shown to regulate one or more of these aforementioned processes. In fact, activated Rac1 has been shown to promote cell-cell adhesion as well as to enhance cell motility, leading to conflicting reports on the effect of Rac1 activation on epithelial cell motility. In this study, we have examined the activation profiles of endogenous Rac1 and ARF6 during the sequential stages of epithelial cell scattering. Using Madin-Darby canine kidney cells treated with hepatocyte growth factor/scatter factor or cell lines stably expressing activated v-Src, we show that Rac1 and ARF6 exhibit distinct activation profiles during cell scattering. We have found that an initial ARF6-dependent decrease in the levels of Rac1-GTP is necessary to induce cell-cell dissociation. This is followed by a steady increase in Rac1 and ARF6 activation and cell migration. In sum, this study documents the progression of ARF6 and Rac1 activities during epithelial cell scattering.     

Prolonged activation of S6K1 does not suppress IRS or PI-3 kinase signaling during muscle cell differentiation

Background  

Myogenesis in C2C12 cells requires the activation of the PI3K/mTOR signaling pathways. Since mTOR signaling can feedback through S6K1 to inhibit the activation of PI3K, the aim of this work was to assess whether feedback from S6K1 played a role in myogenesis and determine whether siRNA mediated knockdown of S6K1 would lead to an increased rate of myotube formation.     

Dynamic and selective interactions of the transcriptional corepressor TIF1 beta with the heterochromatin protein HP1 isotypes during cell differentiation

Cell differentiation is a multi-step process marked by progressive silencing of gene expression through mechanisms believed to involve heterochromatin. We have previously shown that interaction between the Krüppel associated box-containing zinc finger proteins (KRAB-ZFP) corepressor TIF1β and the heterochromatin proteins HP1 is essential for progression through differentiation of embryonal carcinoma F9 cells. This analysis clearly demonstrated the link between gene specific repressors, components of heterochromatin and cell differentiation. In mammals, there are three HP1 isotypes, HP1α, β, and γ, that appear to be involved in both eu- and heterochromatin, but whose individual functions are still poorly defined. Therefore, the aim of the present study was to determine in vivo (i) which HP1 isotypes interact with TIF1β, (ii) in which sub-nuclear compartments these interactions occur and (iii) whether these interactions are regulated during cell differentiation. To address these questions, we established stable F9 cell lines co-expressing TIF1β fused to the ECFP fluorophore and HP1α, β, or γ fused to the EYFP fluorophore. Using the Föster resonance energy transfer (FRET) technology, we map the physical interaction between TIF1β-CFP and the different HP1-YFP isotypes in living F9 cells. We demonstrate that in non-differentiated cells, TIF1β-CFP/HP1-YFP interaction occurs only within euchromatin and involves selectively HP1β-YFP and HP1γ-YFP, but not HP1α-YFP. Furthermore, in differentiated cells, TIF1β-CFP selectively associates with HP1β-YFP within heterochromatin, while TIF1β-CFP/HP1γ-YFP is exclusively present within euchromatin. No physical TIF1β-CFP/HP1α-YFP interaction is detected in neither non differentiated nor differentiated cells. These results support the notion that, in vivo, HP1 isotypes have specific nonredundant functions and provide evidence for HP1β playing an essential role in the shuttling of TIF1β from eu- to heterochromatin during cell differentiation.