| First Author | Srivastava AK | Year | 2022 |
| Journal | Proc Natl Acad Sci U S A | Volume | 119 |
| Issue | 34 | Pages | e2204618119 |
| PubMed ID | 35969745 | Mgi Jnum | J:340362 |
| Mgi Id | MGI:7437414 | Doi | 10.1073/pnas.2204618119 |
| Citation | Srivastava AK, et al. (2022) Serine 408 phosphorylation is a molecular switch that regulates structure and function of the occludin alpha-helical bundle. Proc Natl Acad Sci U S A 119(34):e2204618119 |
| abstractText | Occludin is a tetramembrane-spanning tight junction protein. The long C-terminal cytoplasmic domain, which represents nearly half of occludin sequence, includes a distal bundle of three alpha-helices that mediates interactions with other tight junction components. A short unstructured region just proximal to the alpha-helical bundle is a phosphorylation hotspot within which S408 phosphorylation acts as molecular switch that modifies tight junction protein interactions and barrier function. Here, we used NMR to define the effects of S408 phosphorylation on intramolecular interactions between the unstructured region and the alpha-helical bundle. S408 pseudophosphorylation affected conformation at hinge sites between the three alpha-helices. Further studies using paramagnetic relaxation enhancement and microscale thermophoresis indicated that the unstructured region interacts with the alpha-helical bundle. These interactions between the unstructured domain are enhanced by S408 phosphorylation and allow the unstructured region to obstruct the binding site, thereby reducing affinity of the occludin tail for zonula occludens-1 (ZO-1). Conversely, S408 dephosphorylation attenuates intramolecular interactions, exposes the binding site, and increases the affinity of occludin binding to ZO-1. Consistent with an increase in binding to ZO-1, intravital imaging and fluorescence recovery after photobleaching (FRAP) analyses of transgenic mice demonstrated increased tight junction anchoring of enhanced green fluorescent protein (EGFP)-tagged nonphosphorylatable occludin relative to wild-type EGFP-occludin. Overall, these data define the mechanisms by which S408 phosphorylation modifies occludin tail conformation to regulate tight junction protein interactions and paracellular permeability. |
