ICA512/IA-2: unraveling the role of its luminal domain in â-cells.

JUHA M. TORKKO, ; M. EVANGELINA PRIMO, ; RONALD DIRKX, ; ANKE SANMEZ, ; ANNE FRIEDRICH, ; ANDREAS MALLER, ; CARLA MANSTER, ; DANIELA RICHTER, ; MAURICIO SICA, ; MARIO ERMACORA ; MICHELE SOLIMENA

Simposio; Symposium Diabetes ? a Threat to Mankind? in Helsinki; 2013

Objective. ICA512/IA-2/RPTPN is a catalytically inactive receptor tyrosine phosphatase enriched in the membrane of insulin secretory granules (SGs) of the pancreatic beta-cells. Previous studies suggested that ICA512 signaling allows -cells to adjust the biogenesis of SGs to their consumption. Structural studies indicated that its mature ectodomain (ME ICA512), similar to mucins, contains a SEA domain-like fold. Interestingly, recombinant ME ICA512 was shown to form dimers in vitro. Here we investigated further the role of the luminal domain of ICA512, including ME ICA512, and its glycosylation on the dimerization and trafficking of ICA512 in insulinoma INS-1 cells. Design. Different luminal domain and ME variants of ICA512 were expressed in insulinoma cells and analysed by immunoprecipitation, immunoblotting and immunocytochemistry in regards to ICA512 trafficking as well as insulin biosynthesis and secretion. To this aim we also developed novel luminal domain and anti-ME ICA512 antibodies. Main outcome. The luminal domain targets ICA512 to SGs. ME ICA512 promotes the dimerization of pro-ICA512 in the ER. Replacement of G553 in one of the two predicted interfaces for ME ICA512 dimerization destabilized pro-ICA512 and prevented its exit from the ER. N-glycosylation of ME ICA512, instead, is not critical for ICA512 stability and trafficking. Conclusions. The luminal domain induces the formation of ICA512 homodimers in the ER. Being critical for protein folding, stability and trafficking, its regulation is likely to regulate ICA512 signaling.