inx7 is located at position 6E4 on the X chromosome in a cluster of innexin genes including inx2 and ogre. inx7 transcript is detected in the follicle, embryonic gut and epidermis and in the pharate central nervous system (CNS) (Stebbings et al, 2002). The observed protein distribution confirms this expression pattern but also reveals some expression in the embryonic ventral nerve chord (Ostrowski et al, 2008). The protein can be detected in early embryos indicating that it may be maternally inherited. The extensive expression pattern overlaps, at least at some stage in various tissues and developmental timepoints, with all of the other innexins (Stebbings et al, 2002). Inx7 is unlike any of the other innexins studied so far in that it is localised to the nucleus in embryonic epithelial cells and in some midline glia and pioneer neurons, rather than forming plaques at the plasma membrane (Ostrowski et al, 2008). It has been proposed that the anti-Inx7 antibody used by Ostrowski et al, might detect a cleaved C-terminal domain of Inx7 that mediates a role in the nucleus and that other innexins might also be found there, but other groups have examined Inx2 (images showing innexin protein distribution in the wing disc, brain, salivary gland), Inx3 and Zpg/Inx4 (Bohrmann and Zimmermann, 2008) protein distribution using antibodies raised against the C-terminal and have not reported nuclear localisation. It could just be that nobody has looked in the right tissue at the right time with regard to these other innexins…but for now, Inx7 appears to be unique in this respect. Although it is not yet known whether the Inx7 detected in the nucleus represents the whole protein or just a cleaved portion containing the C-tail, it is highly unlikely that the nuclear Inx7 exists in a mulimeric hemichannel form as it is expected to be in the plasma membrane. As innexins are presumably incorporated into hemichannels in the golgi apparatus (such as the case is with connexins) there would have to be a completely different synthetic/transport pathway for nuclear targeted Inx7 compared to that for plasma membrane targeted innexins….unless, of course, the nuclear Inx7 is a cleavage product. Beyond embryonic stage 7, innexin7 protein is detected in the cytoplasm and at the plasma membrane of epithelial cells (…and is absent from the nucleus?..or not?), so despite the unusual cellular localisation, it still has the potential to form gap junction channels. Within the developing ventral nerve chord Inx7 expressed in midline glia and pioneer neurons remains localised to the nucleus (Ostrowski et al, 2008).
Expression of UAS-inx7 in the developing wing produces different phenotypes than those obtained by expression of UAS-ogre or UAS-inx2 - the other two subunits located in the 6E4 innexin gene cluster. UAS-inx7 induces relatively strong Delta-like (extra vein material) phenotypes but weak Serrate-like phenotypes when expressed in the developing wing. UAS-ogre and UAS-inx2 are poor at producing Delta phenotypes but are good at inducing Serrate-like phenotypes. The UAS-inx7 phenotype observed in legs is also qualitatively different than those induced by UAS-ogre and UAS-inx2. Although interpreting UAS-innexin induced phenotypes is not easy given that even the same UAS-innexin can produce widely differing phenotypes depending on expression level (Strong UAS-inx2 expression in the leg disc) and innexin overexpression can have such a dramatic effect of cell structure and survival (Notes on transgenic experiments). Delta-like phenotypes induced by UAS-inx7 in the wing can be weakly enhanced by the presence of Notch or Inx2Dominant mutation-carrying chromosomes. A link has already been reported between inx2 and ligand-receptor signalling pathways (eg Notch) at the transcriptional level (Lechner et al, 2007)….could the nuclear localisation of Inx7 have a role in this mechanism? Inx2 and Inx7 both possess predicted PDZ type II domains at their C-termini and Inx2 is known to interact with a large number of non-innexin molecules; E-cadherin, β-catenin, β-tubulin (Bauer et al, 2004). There is genetic evidence that Inx2 interacts with sodium pump components ….although there’s no indication that it’s a physical interaction. A yeast two hybrid screen suggests an interaction between Inx7 and the α-subunit of the sodium pump (data accessible from www.Flybase.org). Whether the PDZ domain is required for these interactions is not known.
The biological role Inx7 has been explored in the developing nervous system. Expression of a snapback RNAi construct that significantly knocks down Inx7 levels in ventral nerve chord midline glia leads to aberrant targetting of longitudinal commisures, fusion of anterior and posterior segmental commisures, and also holes in the nervous tissue indicative of cell death (Ostrowski et al, 2008). Inx7 protein is localised to the nucleus of midline glia. It would be interesting to see what phenotypes arise when using the snapback RNA-interference construct in cells where Inx7 is localised to the plasma membrane.