Cytosolic interaction of type III human CD38 with CIB1 modulates cellular cyclic ADP-ribose levels

Significance This study addresses a topological paradox in Ca 2+ signaling mediated by cyclic ADP-ribose (cADPR). The messenger is synthesized by CD38, thought to be a type ll ectoenzyme. Instead, we show that it exists naturally in two opposite membrane orientations and that the type III, with its catalytic domain facing the cytosol, is active in producing cellular cADPR. Its topology was resolved by a technique simultaneously targeting dual epitopes for protein identification, which identifies the intracellular type III CD38 unambiguously. It is also shown that its cADPR-synthesizing activity is regulated by cytosolic interactions with Ca 2+ and integrin-binding protein 1 (CIB1). The results indicate that membrane proteins are not necessarily expressed with only one unique membrane orientation set by sequence motifs as generally believed.