Background: Apoptosis-linked gene 2 (ALG-2) is a calcium-dependent adaptor protein that is recruited to the Sec31A-positive ERES. Results: Physical association between annexin A11 and Sec31A is mediated by ALG-2, and annexin A11 is required for stable association of Sec31A at the ERES. Conclusion: Annexin A11 participates in ER-to-Golgi trafficking. Significance: Annexin A11 is the first annexin shown to modulate the early secretory pathway.

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Apoptosis‐linked gene 2 (ALG‐2), which is a gene product of PDCD6, is a 22‐kDa Ca2+‐binding protein. Accumulating evidence points to a role for ALG‐2 as a Ca2+‐responsive adaptor protein. On binding to Ca2+, ALG‐2 undergoes a conformational change that facilitates its interaction with various proteins. It also forms a homodimer and heterodimer with peflin, a paralog of ALG‐2. However, the differences in cellular roles for the ALG‐2 homodimer and ALG‐2/peflin heterodimer are unclear. In the present study, we found that Trk‐fused gene (TFG) protein interacted with the ALG‐2 homodimer. Immunostaining analysis revealed that TFG and ALG‐2 partially overlapped at endoplasmic reticulum exit sites (ERES), a platform for COPII‐mediated protein transport from the endoplasmic reticulum. Time‐lapse live‐cell imaging demonstrated that both green fluorescent protein‐fused TFG and mCherry‐fused ALG‐2 are recruited to ERES after thapsigargin treatment, which raises intracellular Ca2+ levels. Furthermore, overexpression of ALG‐2 induced the accumulation of TFG at ERES. TFG has an ALG‐2‐binding motif and deletion of the motif decreased TFG binding to ALG‐2 and shortened its half‐life at ERES, suggesting a critical role for ALG‐2 in retaining TFG at ERES. We also demonstrated, by in vitro cross‐linking assays, that ALG‐2 promoted the polymerization of TFG in a Ca2+‐dependent manner. Collectively, the results suggest that ALG‐2 acts as a Ca2+‐sensitive adaptor to concentrate and polymerize TFG at ERES, supporting a potential role for ALG‐2 in COPII‐dependent trafficking from the endoplasmic reticulum.

Clustered protocadherins (Pcdhs), which are cell adhesion molecules, play a fundamental role in self-recognition and non-self-discrimination by conferring diversity on the cell surface. Although systematic cell-based aggregation assays provide information regarding the binding properties of Pcdhs, direct visualization of Pcdh trans interactions across cells remains challenging. Here, we present Förster resonance energy transfer (FRET)-based indicators for directly visualizing Pcdh trans interactions. We developed the indicators by individually inserting FRET donor and acceptor fluorescent proteins (FPs) into the ectodomain of Pcdh molecules. They enabled successful visualization of specific trans interactions of Pcdh and revealed that the Pcdh trans interaction is highly sensitive to changes in extracellular Ca2+ levels. We expect that FRET-based indicators for visualizing Pcdh trans interactions will provide a new approach for investigating the roles of Pcdh in self-recognition and non-self-discrimination processes.