Bradykinin B₂ receptor-mediated transport into intact cells: anti-receptor antibody-based cargoes.

Endocytosis of the bradykinin-stimulated B(2) receptors is parallel to the transport and subsequent degradation of the ligand. To implement biotechnological applications based on receptor-mediated transport, one strategy is to conjugate the agonist ligand to a cargo. Alternatively, we studied whether the B(2) receptor can transport large antibody-based cargoes into intact cells and characterized the ensuing endosomal routing. Myc-tagged B(2) receptors (coded by the vector myc-B(2)R) and a truncated construction devoid of the Ser-Thr phosphorylation domain (myc-B(2)R(trunc) vector) were coupled to anti-myc monoclonal antibodies that did not impair bradykinin binding or elicit calcium signaling in intact cells. Anti-myc antibodies, conjugated or not with secondary antibodies optionally coupled to Qdot nanomaterials, were transported into early endosome autoantigen 1-, and β-arrestin-positive vesicles in bradykinin-stimulated intact cells expressing receptors encoded by myc-B(2)R. Antibody-conjugated cargoes progressed into late-endosomes-lysosomes within 3h without evidence of autophagy. Receptors encoded by myc-B(2)R(trunc) did not support the ligand-controlled endocytosis of anti-myc antibodies. Aside from small ligand-conjugated cargoes, very large antibody-based cargoes can be transported by agonist-stimulated B(2) receptors into intact cells. The latter type of cargo requires a receptor competent for interaction with β-arrestins, enters the degradation pathway separately from the receptor as a function of time and has the potential to confer a qualitatively novel function to a receptor.