Regulation of membrane remodelling during cell division

The Endosomal Sorting Complex Required for Transport (ESCRT) machinery constitutes a family of highly conserved heteromeric protein complexes (ESCRT-0, ESCRT-I, ESCRT-II and ESCRT-III), which orchestrate membrane remodelling processes throughout the cell [1]. Together ESCRTs mediate key events during cell division such as mitotic reformation of the nuclear envelope and cytokinetic abscission [2]. Central to ESCRT-mediated membrane remodelling processes is the action of ESCRT-III, a filament forming complex which constricts membranes [3].

Historically, the ESCRT complexes were first characterised for their role in coordinating endosomal sorting of ubiquitinated proteins [4]. However, operation of ESCRT-III in Archaea, which lack endomembrane structures, suggests the archetypal role of ESCRTs was to function during cell division [5].

During cell division the metazoan microtubule-organising centre (MTOC), the centrosome, coordinates assembly of microtubules and generates opposing poles of the bipolar mitotic spindle which together facilitate DNA segregation [6]. During mitosis, duplicated DNA is separated by microtubule fibres of the mitotic spindle, which requires dynamic remodelling of the nuclear envelope to provide access to chromosomes [7]. Mitotic reformation of the nuclear envelope occurs following segregation of chromosomes to opposite poles of the cell, which is facilitated by ESCRT-III and the microtubule-severing protein spastin [8].

During the final stages of cytokinesis, daughter cells remain connected at the midbody, a transient bridge packed tightly with antiparallel microtubules derived from the mitotic spindle [9]. Here, microtubule severing is mediated by spastin while recruitment of ESCRT-III to the midbody and its subsequent nucleation into helical filaments directs cytokinetic abscission at a secondary constriction site adjacent to the midbody [10, 11].

Therefore, central to orderly reformation of the nuclear envelope and timely cytokinesis abscission are the action of the centrosome and the mitotic spindle which require the action of ESCRT-III to regulate their maintenance and assist with their function [12]. Furthermore, depletion of ESCRT-III subunits alters centrosome and spindle pole numbers, resulting in defects in chromosome segregation materialising in aberrant nuclear morphologies [12].



Recently we showed that CC2D1B, a poorly characterised member of the CC2D1/Lgd family of proteins, regulates ESCRT-III polymerisation at the reforming nuclear envelope during mitosis, preventing premature recruitment which would otherwise lead to failure in the membrane sealing mechanism and removal of penetrating spindle microtubules [13]. CC2D1A, a related protein, has also been shown to prevent ESCRT-III polymerisation in vitro [14]. CC2D1A undergoes mitotic phosphorylation by CDK1, a master regulator of mitosis, and prevents premature centriole splitting during the cell cycle [15, 16].

We have preliminary evidence that suggests CC2D1A acts as a spatiotemporal regulator of ESCRT-III polymerisation and activity during cell division. We found that CC2D1A localises to the cytoplasm and specifically to the poles of the mitotic spindle during mitosis. In CC2D1A-depleted cells, we observed ectopic localisation of ESCRT-III subunits around the mitotic spindle in addition to dysregulated microtubule growth, which coincided with aberrant cellular morphologies. Furthermore, we found that depletion of CC2D1A was associated with delays in abscission timing, but we could not conclude any impact on ESCRT-III polymerisation from our preliminary investigation.
We hypothesise that CC2D1A functions as a spatiotemporal regulator of ESCRT-III activity around the mitotic spindle during cell division by coordinating ESCRT-III polymerisation. Furthermore, we postulate that conformational changes, potentially induced by CDK1 phosphorylation, mediate CC2D1A function.