Commitment to cytokinetic furrowing requires the coordinate activity of microtubules and Plk1

Abstract

At anaphase, spindle microtubules (MTs) position the cleavage furrow and trigger actomyosin assembly by localizing the small GTPase RhoA and the scaffolding protein anillin to a narrow band along the equatorial cortex. Using vertebrate somatic cells we examined the temporal control of furrow assembly. Although its positioning commences at anaphase onset, furrow maturation is not complete until ~10-11 min later. The maintenance of the RhoA/anillin scaffold initially requires continuous signaling from the spindle; loss of either MTs or polo-like kinase 1 (Plk1) activity prevents proper RhoA/anillin localization to the equator, thereby disrupting furrowing. However, we find that at ~6 min post-anaphase, the cortex becomes committed to furrowing; loss of either MTs or Plk1 after this stage does not prevent eventual furrowing, even though at this point the contractile apparatus has not fully matured. Also at this stage, the RhoA/anillin scaffold at the equator becomes permanent. Surprisingly, concurrent loss of both MTs and Plk1 activity following the commitment to furrowing stage results in persistent, asymmetric half-furrows, with only one cortical hemisphere retaining RhoA/anillin, and undergoing regression. This phenotype is reminiscent of asymmetric furrows caused by a physical block between spindle and cortex, or by acentric spindle positioning. The formation of these persistent half-furrows suggests a potential feedback mechanism between the spindle and the cortex that maintains cortical competency along the presumptive equatorial region prior to the commitment to furrowing stage of cytokinesis, thereby ensuring the eventual ingression of a symmetric cleavage furrow.