Abstract
Rho-dependent proteins control assembly of the cytokinetic contractile ring, yet it remains unclear how those proteins guide ring closure and how they promote subsequent formation of a stable midbody ring. Citron kinase is one important component required for midbody ring formation but its mechanisms of action and relationship with Rho are controversial. Here, we conduct a structure-function analysis of the Drosophila Citron kinase, Sticky, in Schneider's S2 cells. We define two separable and redundant RhoGEF/Pebble-dependent inputs into Sticky recruitment to the nascent midbody ring and show that each input is subsequently required for retention at, and for the integrity of, the mature midbody ring. The first input is via an actomyosin-independent interaction between Sticky and Anillin, a key scaffold also required for midbody ring formation. The second input requires the Rho-binding domain of Sticky, whose boundaries we have defined. Collectively, these results show how midbody ring biogenesis depends on the coordinated actions of Sticky, Anillin, and Rho.