Structures of Vac8-containing protein complexes reveal the underlying mechanism by which Vac8 regulates multiple cellular processes.

Hyejin Kim, Jihyeon Park, Hyunwoo Kim, Naho Ko, Jumi Park, Eunhong Jang, So Young Yoon, Joyce Anne R Diaz, Changwook Lee, Youngsoo Jun

Proceedings of the National Academy of Sciences of the United States of America 2023 May 02

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Vac8, a yeast vacuolar protein with armadillo repeats, mediates various cellular processes by changing its binding partners; however, the mechanism by which Vac8 differentially regulates these processes remains poorly understood. Vac8 interacts with Nvj1 to form the nuclear-vacuole junction (NVJ) and with Atg13 to mediate cytoplasm-to-vacuole targeting (Cvt), a selective autophagy-like pathway that delivers cytoplasmic aminopeptidase I directly to the vacuole. In addition, Vac8 associates with Myo2, a yeast class V myosin, through its interaction with Vac17 for vacuolar inheritance from the mother cell to the emerging daughter cell during cell divisions. Here, we determined the X-ray crystal structure of the Vac8-Vac17 complex and found that its interaction interfaces are bipartite, unlike those of the Vac8-Nvj1 and Vac8-Atg13 complexes. When the key amino acids present in the interface between Vac8 and Vac17 were mutated, vacuole inheritance was severely impaired in vivo. Furthermore, binding of Vac17 to Vac8 prevented dimerization of Vac8, which is required for its interactions with Nvj1 and Atg13, by clamping the H1 helix to the ARM1 domain of Vac8 and thereby preventing exposure of the binding interface for Vac8 dimerization. Consistently, the binding affinity of Vac17-bound Vac8 for Nvj1 or Atg13 was markedly lower than that of free Vac8. Likewise, free Vac17 had no affinity for the Vac8-Nvj1 and Vac8-Atg13 complexes. These results provide insights into how vacuole inheritance and other Vac8-mediated processes, such as NVJ formation and Cvt, occur independently of one another.

Citation

Hyejin Kim, Jihyeon Park, Hyunwoo Kim, Naho Ko, Jumi Park, Eunhong Jang, So Young Yoon, Joyce Anne R Diaz, Changwook Lee, Youngsoo Jun. Structures of Vac8-containing protein complexes reveal the underlying mechanism by which Vac8 regulates multiple cellular processes. Proceedings of the National Academy of Sciences of the United States of America. 2023 May 02;120(18):e2211501120