Myosin-I's motor and actin assembly activation activities are modular and separable in budding yeast clathrin-mediated endocytosis.

The myosin-Is, Myo3 and Myo5 in budding yeast, are implicated in force generation and actin assembly during clathrin-mediated endocytosis (CME). The myosin-Is have motor activity, bind the plasma membrane, and activate the Arp2/3 complex to promote branched actin assembly. We reveal that Myo5 's force-generating motor activity and nucleation-promoting factor (NPF) activity each must be coupled to membrane binding for successful CME. However, the motor and NPF activities are modular and separable, showing that these activities function independently rather than in an obligatorily integrated manner to provide myosin-I's essential functions in actin network assembly and force generation during budding yeast CME.