Schonteich Eric, Pilli Manohar, Simon Glenn C, Matern Hugo T, Junutula Jagath R, Sentz Danielle, Holmes Randall K, Prekeris Rytis
Department of Cellular and Developmental Biology, School of Medicine, University of Colorado Health Sciences Center, 12801 E. 17th Avenue, Aurora, CO 80045, USA.
Eur J Cell Biol. 2007 Aug;86(8):417-31. doi: 10.1016/j.ejcb.2007.05.004. Epub 2007 Jul 12.
Rab11-FIP3 is a Rab11-binding protein that has been implicated in regulating cytokinesis in mammalian cells. FIP3 functions by simultaneously interacting with Rab11 as well as Arf GTPases. However, unlike the interaction between Rab11 and FIP3, the structural basis of FIP3 binding to Arf GTPases has not yet been determined. The specificity of interaction between FIP3 and Arf GTPases remains controversial. While it was reported that FIP3 preferentially binds to Arf6 some data suggest that FIP3 can also interact with Arf5 and even possibly Arf4. The Arf-interaction motif on FIP3 also remains to be determined. Finally, the importance of Arf binding to FIP3 in regulating cell division and other cellular functions remains unclear. Here we used a combination of various biochemical techniques to measure the affinity of FIP3 binding to various Arfs and to demonstrate that FIP3 predominantly interacts with Arf6 in vitro and in vivo. In addition, we identified the motifs mediating Arf6 and FIP3 interaction and demonstrated that FIP3 binds to the Arf6 C-terminus rather than switch motifs. Finally we show that FIP3 and Arf6 binding is required for the targeting of Arf6 to the cleavage furrow during cytokinesis. Thus, we propose that FIP3 is a scaffolding protein that, in addition to regulating endosome targeting to the cleavage furrow, also is required for Arf6 recruitment to the midbody during late telophase.
Rab11 - FIP3是一种与Rab11结合的蛋白质，它参与调控哺乳动物细胞的胞质分裂。FIP3通过同时与Rab11以及Arf GTP酶相互作用来发挥功能。然而，与Rab11和FIP3之间的相互作用不同，FIP3与Arf GTP酶结合的结构基础尚未确定。FIP3与Arf GTP酶之间相互作用的特异性仍存在争议。虽然有报道称FIP3优先结合Arf6，但一些数据表明FIP3也能与Arf5相互作用，甚至可能与Arf4相互作用。FIP3上的Arf相互作用基序也有待确定。最后，Arf与FIP3结合在调控细胞分裂和其他细胞功能中的重要性仍不清楚。在此，我们运用多种生化技术相结合的方法来测量FIP3与各种Arf的结合亲和力，并证明FIP3在体外和体内主要与Arf6相互作用。此外，我们鉴定了介导Arf6与FIP3相互作用的基序，并证明FIP3与Arf6的C末端而非开关基序结合。最后，我们表明在胞质分裂过程中，FIP3与Arf6的结合是Arf6靶向分裂沟所必需的。因此，我们提出FIP3是一种支架蛋白，除了调控内体靶向分裂沟外，在末期后期Arf6募集到中体的过程中也是必需的。
