Employing techniques such as the GAL4/UAS system (Brand and Dormand, 1995) to express proteins is a common approach used to discover the biological role(s) of molecules. One caveat to the approach is that different UAS-yourfavouritegene lines can express wildly different amounts of protein even when transcriptionally regulated by the same single GAL4 line. Weakly expressing UAS-ogre lines do not appear to destroy cells, yet produce developmental phenotypes and may yield useful information about innexin function. Moderately expressing UAS-ogre lines can lead to accumulation of ogre subunits in the cytoplasm (see webpage - Expressing innexin transgenes). Strongly expressing UAS-ogre lines induce severe developmental phenotypes, but these may be secondary effects due to cell destruction/death (image at top of page) and not directly associated with innexin function. Drawing conclusions based on moderate and strong UAS-innexin expressing fly lines requires caution. Bauer et al, (2004) have reported that overexpression of UAS-Inx2 can lead to changes in epithelial cell polarity and lead to increased levels of cell death in the embryo. Few, if any, innexin-overexpression phenotypes have so far been linked to changes in membrane gap junction channel quantity or activity and it's quite possible that the induced phenotypes are not related to channel activity at all, and instead possibly reflect sequestration of proteins that interact with the overexpressed innexin away from the sites where they are required for normal cell function. It is interesting that high levels of innexin expression may be related to cell death in some way. Incorporation of endogenous innexins Inx2, Inx3 and Ogre into the salivary gland plasma membrane increases significantly between the larval and pupal stages and innexin levels are therefore high in the lead-up to apoptosis (Image: Inx2 protein levels change over time and associated webpage) - whether this serves any role in the process of programmed cell death is unknown.
Using transgenics to study the role of innexins in development
Figure legend: Strong expression of an innexin subunit can destroy cells, complicating interpretation of induced phenotypes iunn developmental studies. A, a wild type wing disc expressing UAS-CD8:GFP in a stripe under the transcriptional control of the dpp-GAL4 enhancer trap line. B, Close-up of wing pouch cells from A. The CD8-GFP reporter incorporates into the plasma membrane, revealing an outline of each cell. C, a whole wing disc from a larva expressing UAS-CD8:GFP and UAS-ogre under the direction of dpp-GAL4. The stripe expression pattern is disrupted. D, A single optical section close-up of the wing pouch region (from C) no-longer reveals cell outlines. Also, the appearance of intensely fluorescent GFP spots indicates that cells may be destroyed by chronic over-expression of wild type Ogre protein.
This is the end of the Innexins and development section