Roundabout 1 exists predominantly as a basal dimeric complex and this is unaffected by binding of the ligand Slit2

Roundabout (Robo) receptors and their Slit polypeptide ligands are known to play key roles in neuronal development and have been implicated in both angiogenesis and cancer development. Like the other family members Robo1 is a large, single transmembrane domain, polypeptide containing a series of well defined extracellular elements. However, the intracellular domain lacks structural definition and little is known about the quaternary structure of Robo receptors or how binding of a Slit might affect this. To address these questions combinations of both autofluorescent protein-based FRET imaging and time-resolved FRET were employed. Both approaches identified oligomeric organization of Robo1 that did not require the presence of the intracellular domain. Spatial Intensity Distribution Analysis of eGFP-tagged forms of Robo1 indicated that for a C-terminally deleted version approximately 2/3rds of the receptor was present as a dimer and 1/3rd as monomer. By contrast, full length Robo1 was present almost exclusively as a dimer. In each case this was unaffected by addition of Slit2 although parallel studies demonstrated the biological activity of Slit2 and its interaction with Robo1. Deletion of both the immunoglobulin and fibronectin type III extracellular repeats prevented dimer formation, with the immunoglobulin repeats providing the bulk of the protein-protein interaction affinity.
Source: BJ Cell - Category: Biochemistry Authors: Tags: BJ Biomolecules Source Type: research