Successful cell division is dependent on the proper formation and precise positioning of the mitotic spindle. The assembly of the mitotic spindle starts in prophase with the nucleation of the microtubules by the centrosomes. Then, aster microtubules grow and extend toward the cell cortex (Lu and Johnston, 2013). In the case of symmetric division, as mitosis progresses, cortical polarity cues position the mitotic spindle in order that the cleavage furrow will bisect the cell in the middle of the central spindle in two equal parts during cytokinesis.

In interphase, RAB11 is associated to recycling endosomes (Grant and Donaldson, 2009). At early stages of mitosis in C. elegans, RAB11, in association with dynein, regulates astral microtubule size, spindle alignment, and the morphology of endoplasmic reticulum (Zhang et al., 2008; Ai and Skop, 2009). The detailed localization of RAB11 in C. elegans at early stages of mitosis has not been described yet. In mammalian cells, a role for RAB11-positive endosomes in spindle pole organization and orientation was recently reported (Hehnly and Doxsey, 2014). Using time-lapse imaging, it was shown that RAB11 endosomes are found localized on the mitotic spindle and at the mitotic spindle poles (Hehnly and Doxsey, 2014; Table 1 and Figure 1). These mitotic recycling endosomes bind to microtubule-nucleating components and to dynein. Astral microtubule disruption, a mitotic delay and a redistribution of spindle poles proteins are observed following RAB11 inhibition. As proposed by the authors, RAB11 endosomes could be part of a dynein-dependent retrograde transport pathway bringing microtubule nucleating factors and spindle pole proteins to mitotic spindle poles (Das et al., 2015).

In interphase, RAB5 is associated with early endosomes. Two studies performed in Drosophila and mammalian cells reported the involvement of RAB5 at an early stage of cell division, where RAB5 acts by modulating the congression and segregation of chromosomes (Capalbo et al., 2011; Serio et al., 2011; Lanzetti, 2012). This function of RAB5 is evolutionary conserved. In both Drosophila and mammalian cells, at early mitosis, RAB5 localizes to endosomes organized around the spindle poles (Capalbo et al., 2011; Serio et al., 2011; Lanzetti, 2012; Table 1 and Figure 1). During Drosophila mitosis, RAB5 is required for proper chromosome alignment before anaphase (Capalbo et al., 2011). RAB5 associates in vivo with nuclear lamin and Mushroom Body Defect (Mud), the Drosophila homolog of the nuclear mitotic apparatus protein (NuMA), which is known to be important for spindle formation and maintenance in mammalian cells. RAB5 is required for the disassembly of the nuclear envelope at mitotic entry and the accumulation of Mud at the spindle poles (Capalbo et al., 2011). In mammalian U2OS cells, RAB5 silencing causes defects in chromosome congression and a marked prometaphase delay, due to a reduction in the localization of the protein CENP-F to kinetochores (Serio et al., 2011). CENP-F is a centromere-associated protein that contributes to the establishment of kinetochore-microtubule interactions. In mitosis, RAB5 can form a complex with CENP-F and regulates the accumulation of CENP-F to kinetochores through the regulation of its kinetic release from kinetochores (Serio et al., 2011). How RAB5 regulates the accumulation of CENP-F to kinetochores remains to be elucidated. Which population of RAB5 is active? Cytosolic or membrane-associated? The accumulation of CENP-F is done via vesicular transport or through the association with a common CENP-F/RAB5 effector? What are the effectors of RAB5 involved in chromosome congression? The identification of specific RAB5 effectors, using proteomic analysis for example, at early mitosis could help resolve these questions.

The other interesting point to address is the existence of RAB5-RAB11 endosomal maturation from prophase to telophase. In interphase cells, RAB5-labeled early endosomes can mature into RAB11-labeled late endosomes (Scott et al., 2014). As mitosis progresses, the amount of RAB5-positive endosomes decreases (Serio et al., 2011; Lanzetti, 2012) while RAB11-positive endosomes accumulate (Hobdy-Henderson et al., 2003).

RAB24 is a member of the RAB GTPase family whose specific function is presently unknown. This atypical RAB is expressed ubiquitously but presents peculiar properties such as a very low intrinsic GTPase activity and inefficient prenylation in comparison with other RABs (Erdman et al., 2000). The description of the localization of RAB24 in interphase and mitosis is mostly based on overexpression of tagged constructs although the distribution of the endogenous protein has also been analyzed by using specific antibodies.

In interphase, RAB24 presents a broad intracellular localization: a perinuclear reticular localization frequently surrounding the nucleus (Olkkonen et al., 1993; Erdman et al., 2000; Munafó and Colombo, 2002), nuclear inclusions (in the case of an “empty mutant”) (Maltese et al., 2002; Wu et al., 2006), a co-localization with ER-Golgi intermediate compartments and the cis-Golgi marker CTR433 (Munafó and Colombo, 2002), a partial overlap with late endosomal markers such as RAB7 (Olkkonen et al., 1993) and finally, after stimulation of autophagy, an association to autophagosomes (Munafó and Colombo, 2002).

During mitosis, RAB24 distribution shows a distinct pattern depending on the stage of mitosis (Militello et al., 2013; Table 1 and Figure 1). Throughout metaphase and anaphase, RAB24 is present at the mitotic spindle. In metaphase, Rab24 overexpression causes chromosomes misalignment with abnormal spindle formation. In addition, a partial overlap of RAB24 with tubulin has also been observed. RAB24's association with microtubules was also demonstrated both in vivo and in vitro (Militello et al., 2013). Thus, several mitotic steps are modulated by RAB24, perhaps via its interaction with microtubules.

However, how RAB24 can associate directly to tubulin has to be elucidated. Is the active RAB24 fraction membrane-associated or cytosolic? Until now, two common RAB24/RAB6 effectors, R6IP1 and GAPCenA have been identified (see detailed discussion below). The identification of specific RAB24 effectors involved in early stages of mitosis would be critical to determine the precise role of this protein at the molecular level.

The functions of many RABs require continuous association and dissociation cytoplasm-membrane cycles. In mitosis, this cycle is disrupted in the case of RAB4. The phosphorylation of RAB4 by the mitotic kinase p34^cdc2 (Bailly et al., 1991) increases the amount of RAB4:GTP in the cytoplasm. This phosphorylation likely results in a less efficient recruitment of RAB4 effectors onto mitotic endosomal membranes and arrest of the endocytic process (Bailly et al., 1991; van der Sluijs et al., 1992; Gerez et al., 2000). RAB4:GTP is maintained in the cytosol through an association with the peptidyl–prolyl isomerase Pin1 (Gerez et al., 2000). The precise function of RAB4 during mitosis remains unknown. The possible phosphorylation of other RABs is discussed in the last section.

Cytokinesis is the terminal stage of eukaryotic cell division. At this stage, in the case of symmetric division, the cytoplasm of the dividing cell is partitioned equally between two daughter cells. Cytokinesis involves complex changes in cell shape. A narrowing acto-myosin contractile ring is formed between the poles of the mitotic spindle and is responsible for ingression of the cleavage furrow (McCollum, 2004). However, this is not the only mechanism that drives cytokinesis. Membrane trafficking is also crucial for abscission (Matheson et al., 2005; Barr and Grüneberg, 2007). Indeed, endosomes constitute a reservoir of new membrane which are incorporated into the cleavage furrow (Boucrot and Kirchhausen, 2007). In addition, a specific endosome-dependent targeting of key proteins involved in the final stage of cytokinesis, implicating RAB11 and RAB35, has also been reported. This may explain why numerous endosome-associated RABs have been implicated in cytokinesis.

In C. elegans, a role for RAB GTPases in late stages of cell division has been documented (Yu et al., 2007). siRNA-mediated depletion of RAB11 leads to cytokinesis defects, including furrow regression and abnormal abscission (Skop et al., 2001). Both RAB11 and its interacting protein RAB11-FIP3 localize to the cleavage furrow during cytokinesis (Horgan et al., 2004; Table 1 and Figure 1). In mammalian cells, RAB11 and RAB11-FIP3 containing recycling endosomes are found accumulated near the cleavage furrow (Wilson et al., 2005).

In interphase, RAB35 localizes to the endocytic recycling pathway, much like RAB11. RAB35 functions at early endosomes in a fast-recycling endocytic pathway prior to the slow recycling endosomal step regulated by RAB11. In cytokinesis, RAB35 is essential for post-furrowing stages (Kouranti et al., 2006). In interphase and mitosis, RAB35 binds to the phosphatase ocucerebrorenal syndrome of Lowe (OCRL). During the post-furrowing cytokinesis stages, OCRL is targeted to the cleavage furrow via RAB35-positive endosomes (Table 1 and Figure 1). There, OCRL regulates PtdIns(4,5)P2 hydrolysis and locally remodels the F-actin cytoskeleton at the intercellular bridge. This event is important for normal cytokinesis abscission (Dambournet et al., 2011). In addition, an ARF6/RAB35 cascade controlling endocytic recycling and successful cytokinesis has been described (Chesneau et al., 2012). EPI64B (a GAP for RAB35) acts as an effector of Arf6 and negatively regulates RAB35 activation. This molecular mechanism controls the RAB35 pathway, including RAB35 localization at the bridge and hence completion of cytokinesis (Chesneau et al., 2012). Regarding the signaling function of RAB11-positive endosomes during cytokinesis, it has been shown that FIP3/RAB11/Arf6 endosomes deliver key proteins involved in late abscission steps, SCAMP2/3 and p50Rho-GAP, to the cleavage furrow (Schiel et al., 2012).

In interphase, RAB21 is an endosomal RAB involved in the regulation of cell adhesion and migration. RAB21 acts through the targeted trafficking of integrins via its association with integrin alpha tails (Pellinen et al., 2006). In cytokinesis, RAB21 targets integrins to the cleavage furrow (Pellinen et al., 2008). In telophase and cytokinesis, RAB21 vesicles are localized to the opposite poles of the daughter cells and to the cleavage furrow (Pellinen et al., 2008). As suggested by Pellinen et al. (2008), these two pools of RAB21 vesicles would have two different functions. The pool of RAB21 vesicles localized to the opposite poles of the daughter cells are associated with β1-intergrin and are forming protrusions at the level of the matrix. These structures would help the mechanical separation of the two daughter cells. The pool of RAB21 vesicles targeted at the cleavage furrow would regulate RhoA activity and consequently the activity of known RhoA effectors. Loss of RAB21 gene expression in human cancer leads to the accumulation of multinucleated cells (Pellinen et al., 2008). In addition, abnormal integrin trafficking was linked with the generation of aneuploidy and cell transformation. In human prostate and ovarian cancer samples, increased malignancy is correlated with downregulation of RAB21. Long-term depletion of RAB21 is sufficient to induce chromosome number aberrations in normal human epithelial cells (Högnäs et al., 2011).

RAB8A has also been implicated in cytokinesis (Table 1, Figure 1). The protein doublecortin domain-containing protein 5 (DCDC5) interacts with cytoplasmic dynein and RAB8 (RAB8A), as well as with the RAB8 nucleotide exchange factor RABin8 (Kaplan and Reiner, 2011). Following DCDC5 knockdown, the duration of metaphase/anaphase transition and cytokinesis, as well as the amount of multinucleated cells, increases. DCDC5 therefore appears to play a role in mediating dynein-dependent transport of RAB8-positive vesicles to coordinate late cytokinesis events.

In cytokinesis, RAB24 is found associated to the midbody (Table 1, Figure 1). In cells overexpressing RAB24 or in RAB24 silenced cells, long chromatin bridges connect cells undergoing cell division (Militello et al., 2013). As a consequence, cells are unable to undergo abscission and cytokinetic furrows eventually retract, leading to the appearance of binucleated and multinucleated cells. It is likely that the observed defects in cell division are due to defects in congression and segregation of chromosomes as explained in the previous section.

Interestingly, RAB24 and RAB6 disruption share a common phenotype: a block in mitosis at the metaphase/anaphase transition and two common effectors, GAPCenA and RAB6IP1. As discussed above, RAB6 and GAPCenA function are required for the metaphase/anaphase transition (Miserey-Lenkei et al., 2006). In the laboratory of M. Colombo, it has been shown that RAB24 interacts with GAPCenA and colocalizes with GAPCenA at the centrosome (Militello et al., 2013). In addition, depletion of R6IP1 leads to a block at the metaphase/anaphase transition and a defect in cytokinesis (Miserey-Lenkei et al., 2007). Preliminary results from the laboratory of M. Colombo indicate that in addition to RAB6 and RAB11, R6IP1 also interacts with RAB24. The existence of these common interactions, with GAPCenA and R6IP1, may in part explain the increased number of cells arrested in metaphase observed in RAB24-depleted cells (Militello et al., 2013). It would now be interesting to address the existence of a dialogue between RAB24, RAB6 in association with GAPCenA and R6IP1 to determine how the function of these proteins is coordinated at the metaphase/anaphase transition.