CRF grant ̈Molecular anatomy and physiology of human cystinosin ̈

Cystinosin is predicted to comprise seven transmembrane domains but its actual topology remains unknown. As a first step to address this issue, we used a classical gene fusion approach in which a topology reporter is fused to various truncated fragments of the membrane protein of interest in order to map the topological orientation of a set of truncation sites along the polypeptide (see Manoil and Beckwith 1986, Science 233:1403-8). A series of C-terminally truncated fragments of human cystinosin linked to a glycosylationbased topology reporter were constructed as depicted in Figure 1.