Lapierre Lynne A, Goldenring James R
Methods Enzymol. 2005;403:715-23. doi: 10.1016/S0076-6879(05)03062-4.
Myosin Vb interacts with Rab11 family members and is the major motor protein identified in association with plasma membrane recycling systems in nonpolarized and polarized cells. Using yeast two-hybrid binary screens, we demonstrated that dominant active Rab25S21V fails to interact with myosin Vb, but does interact with Rab11-FIP2. Transfection of DsRed2-myosin Vb tail in MDCK cell lines stably transfected with wild-type or dominant active forms of Rab11a or Rab25 demonstrated that the distribution of Rab25S21V is only partially altered by expression of the myosin Vb tail. Finally, we demonstrate EGF-dependent sequestration of internalized EGF receptor by EGFP-myosin Vb in A431 cells. Expression of myosin Vb tail represents a powerful provocative test for the trafficking of cargoes through Rab11-containing plasma membrane recycling systems.
肌球蛋白Vb与Rab11家族成员相互作用,是在非极化和极化细胞中与质膜回收系统相关联的主要运动蛋白。通过酵母双杂交二元筛选,我们证明显性活性Rab25S21V不与肌球蛋白Vb相互作用,但确实与Rab11-FIP2相互作用。在稳定转染野生型或显性活性形式的Rab11a或Rab25的MDCK细胞系中转染DsRed2-肌球蛋白Vb尾部,结果表明Rab25S21V的分布仅部分地因肌球蛋白Vb尾部的表达而改变。最后,我们证明在A431细胞中,EGFP-肌球蛋白Vb对内化的表皮生长因子受体(EGF受体)具有EGF依赖性隔离作用。肌球蛋白Vb尾部的表达代表了通过含Rab11的质膜回收系统对货物运输进行有力的激发试验。