article published yes Christian F Düllberg author 459064DC-F248-11E8-B48F-1D18A9856A870000-0001-6335-9748 Albert Auer author 3018E8C2-F248-11E8-B48F-1D18A9856A870000-0002-3580-2906 Nikola Canigova author 3795523E-F248-11E8-B48F-1D18A9856A870000-0002-8518-5926 Katrin Loibl author 3760F32C-F248-11E8-B48F-1D18A9856A870000-0002-2429-7668 Martin Loose author 462D4284-F248-11E8-B48F-1D18A9856A870000-0001-7309-9724 MaLo department MiSi department Reconstitution of cell polarity and axis determination in a cell-free system project The differentiation of cells depends on a precise control of their internal organization, which is the result of a complex dynamic interplay between the cytoskeleton, molecular motors, signaling molecules, and membranes. For example, in the developing neuron, the protein ADAP1 (ADP-ribosylation factor GTPase-activating protein [ArfGAP] with dual pleckstrin homology [PH] domains 1) has been suggested to control dendrite branching by regulating the small GTPase ARF6. Together with the motor protein KIF13B, ADAP1 is also thought to mediate delivery of the second messenger phosphatidylinositol (3,4,5)-trisphosphate (PIP3) to the axon tip, thus contributing to PIP3 polarity. However, what defines the function of ADAP1 and how its different roles are coordinated are still not clear. Here, we studied ADAP1’s functions using in vitro reconstitutions. We found that KIF13B transports ADAP1 along microtubules, but that PIP3 as well as PI(3,4)P2 act as stop signals for this transport instead of being transported. We also demonstrate that these phosphoinositides activate ADAP1’s enzymatic activity to catalyze GTP hydrolysis by ARF6. Together, our results support a model for the cellular function of ADAP1, where KIF13B transports ADAP1 until it encounters high PIP3/PI(3,4)P2 concentrations in the plasma membrane. Here, ADAP1 disassociates from the motor to inactivate ARF6, promoting dendrite branching. National Academy of Sciences2021 eng 0027-8424 1091-6490 33443153 00060727010001810.1073/pnas.2010054118 1181 Düllberg, C. F., Auer, A., Canigova, N., Loibl, K., &#38; Loose, M. (2021). In vitro reconstitution reveals phosphoinositides as cargo-release factors and activators of the ARF6 GAP ADAP1. <i>Proceedings of the National Academy of Sciences of the United States of America</i>. National Academy of Sciences. <a href="https://doi.org/10.1073/pnas.2010054118">https://doi.org/10.1073/pnas.2010054118</a> Düllberg CF, Auer A, Canigova N, Loibl K, Loose M. In vitro reconstitution reveals phosphoinositides as cargo-release factors and activators of the ARF6 GAP ADAP1. <i>Proceedings of the National Academy of Sciences of the United States of America</i>. 2021;118(1). doi:<a href="https://doi.org/10.1073/pnas.2010054118">10.1073/pnas.2010054118</a> C.F. Düllberg, A. Auer, N. Canigova, K. Loibl, M. Loose, Proceedings of the National Academy of Sciences of the United States of America 118 (2021). Düllberg, Christian F, Albert Auer, Nikola Canigova, Katrin Loibl, and Martin Loose. “In Vitro Reconstitution Reveals Phosphoinositides as Cargo-Release Factors and Activators of the ARF6 GAP ADAP1.” <i>Proceedings of the National Academy of Sciences of the United States of America</i>. National Academy of Sciences, 2021. <a href="https://doi.org/10.1073/pnas.2010054118">https://doi.org/10.1073/pnas.2010054118</a>. Düllberg, Christian F., et al. “In Vitro Reconstitution Reveals Phosphoinositides as Cargo-Release Factors and Activators of the ARF6 GAP ADAP1.” <i>Proceedings of the National Academy of Sciences of the United States of America</i>, vol. 118, no. 1, e2010054118, National Academy of Sciences, 2021, doi:<a href="https://doi.org/10.1073/pnas.2010054118">10.1073/pnas.2010054118</a>. Düllberg CF, Auer A, Canigova N, Loibl K, Loose M. 2021. In vitro reconstitution reveals phosphoinositides as cargo-release factors and activators of the ARF6 GAP ADAP1. Proceedings of the National Academy of Sciences of the United States of America. 118(1), e2010054118. C. F. Düllberg, A. Auer, N. Canigova, K. Loibl, and M. Loose, “In vitro reconstitution reveals phosphoinositides as cargo-release factors and activators of the ARF6 GAP ADAP1,” <i>Proceedings of the National Academy of Sciences of the United States of America</i>, vol. 118, no. 1. National Academy of Sciences, 2021. 89882021-01-03T23:01:23Z2025-05-14T10:59:29Z