Analysis of protein dynamics within the septate junction reveals a highly stable core protein complex that does not include the basolateral polarity protein Discs large

Abstract

Barrier junctions prevent pathogen invasion and restrict paracellular leakage across epithelial sheets. To understand how one barrier junction, the septate junction (SJ), is regulated in vivo, we used fluorescence recovery after photobleaching (FRAP) to examine SJ protein dynamics in Drosophila. Most SJ-associated proteins, including Coracle, Neurexin IV and Nervana 2, displayed similar, extremely immobile kinetics. Loss of any of these components resulted in dramatically increased mobility of all others, suggesting that they form a single, highly interdependent core complex. Immobilization of SJ core components coincided with formation of the morphological SJ but occurred after their known role in maintaining epithelial polarity, suggesting that these functions are independent. In striking contrast to the core components, the tumor suppressor protein Discs large was much more mobile and its loss did not affect mobility of core SJ proteins, suggesting that it is not a member of this complex, even though it colocalizes with the SJ. Similarly, disruption of endocytosis affected localization of SJ core components, but did not affect their mobility. These results indicate that formation of a stable SJ core complex is separable from its proper subcellular localization, and provide new insights into the complex processes that regulate epithelial polarity and assembly of the SJ.

Axonal wrapping in theDrosophilaPNS is controlled by glia-derived neuregulin homolog Vein

“…As Gliotactin is an important constituent of tricellular septate junctions and Neuroglian is a core component of general septate junctions (Genova and Fehon, 2003;Oshima and Fehon, 2011;Schulte et al, 2003), this corroborates the pivotal function of the EGF receptor in organizing septate junctions in the SPG. A possible link between EGF receptor activation and septate junction formation might either stem from a direct transcriptional activation or it could be due to changes in alternative splicing (Edenfeld et al, 2006;Oshima and Fehon, 2011). Cell type-specific splicing of neurexinIV, encoding another core component of septate junctions, is brought about by the splicing factor How, the activity of which can be regulated through phosphorylation by Raf/MAP kinase (Nir et al, 2012;Rodrigues et al, 2012).…”

“…Likewise, it was shown that expression of Neuroglian is dramatically reduced in egfr mutants (Sepp and Auld, 2003b). As Gliotactin is an important constituent of tricellular septate junctions and Neuroglian is a core component of general septate junctions (Genova and Fehon, 2003;Oshima and Fehon, 2011;Schulte et al, 2003), this corroborates the pivotal function of the EGF receptor in organizing septate junctions in the SPG. A possible link between EGF receptor activation and septate junction formation might either stem from a direct transcriptional activation or it could be due to changes in alternative splicing (Edenfeld et al, 2006;Oshima and Fehon, 2011).…”

Macroglobulin complement-relatedencodes a protein required for septate junction organization and paracellular barrier function inDrosophila

“…Similar to mutations in other genes encoding core SJ components (Oshima and Fehon, 2011) (Fig. 5A,B; supplementary material Fig.…”

“…Behr et al, 2003;Genova and Fehon, 2003;Paul et al, 2003;Faivre-Sarrailh et al, 2004;Tiklová et al, 2010). Fluorescence recovery after photobleaching (FRAP) experiments have revealed that SJ proteins are essentially fixed in the membrane by stage 14 of embryogenesis (Laval et al, 2008;Oshima and Fehon, 2011). Mutations in any core SJ gene increases the mobility of other SJ proteins at stage 14, further demonstrating the highly stable and interdependent nature of the SJ complex.…”

A somatic permeability barrier around the germline is essential forDrosophilaspermatogenesis

“…We found that the core septate junction components Neurexin-IV (Nrx-IV) and Coracle (Cora) (Oshima and Fehon, 2011) both localized around the germline throughout spermatogenesis (Fig. 7A).…”