MISP: The missing link between extracellular matrix and astral microtubules.

Positioning of the cell division plane is critically important for tissue morphogenesis and architecture. It is therefore not surprising that mitotic spindle orientation must be tightly regulated in living tissues, a phenomenon that is also observed in cells cultured in vitro. Because of the amenability of cultured cells to molecular and physical manipulation, many investigators have used such approaches to identify the unifying rules that control spindle positioning., One long-standing idea is that in tissue culture cells, the long axis of the mitotic spindle aligns with the long axis of the cell. However, many cell types round up during mitosis, which possibly erases pre-mitotic geometric cues. What then, if anything, controls spindle orientation in rounded mitotic cells? Some very elegant studies have recently shown that the extracellular matrix can control spindle positioning via forces that are transmitted through the plasma membrane and which are linked to the retraction fibers formed as the cell rounds up.- On the other (intracellular) side of the plasma membrane, spindle positioning is believed to be controlled by the astral microtubules interacting with the cell cortex via the dynein/dynactin complex. What has been unclear so far is the force transmission mechanism linking extracellular space and astral microtubules. A new study by Maier et al. identifies MISP (mitotic interactor and substrate of Plk1) as the missing link in this network of force-transmitting elements (Fig. 1). The authors had previously identified this protein in a genome-wide siRNA screen for proteins required for centrosome clustering in cancer cells with supernumerary centrosomes. In the present study, the authors used MISP-siRNA and immunohistochemistry to better understand the role of this protein in spindle assembly and function. Key findings of this study include: (1) MISP depletion causes defects in spindle orientation and positioning, and (2) MISP colocalizes with the actin cytoskeleton and focal adhesions (specifically, the focal adhesion kinase, FAK). The authors also find that MISP interacts with the plus-end-tracking protein EB1 and the p150glued subunit of the dynein/dynactin complex, and that cells depleted of MISP display a mitotic arrest/delay. Further insight on the role of MISP in spindle positioning was provided by another recent study also showing that MISP is an actin-associated protein important for spindle positioning. In this study, using live-cell imaging, the Zhu et al. showed that MISP depletion resulted in “unstable” spindle position, in which initially the spindle assembled correctly and the chromosomes aligned properly at the metaphase plate. However, spindle position and chromosome alignment could not be maintained, as the spindles frequently rotated and rocked inside the cell. The authors attributed this behavior to the role of MISP in stabilizing astral microtubules and regulating the cortical distribution of p150glued. This behavior could also explain the presence of BubR1 (indicative of an active mitotic checkpoint) at the kinetochores of MISP-depleted cells., This could happen because the observed instability may alter the balance of forces within the mitotic spindle, and thus reduce the stability of kinetochore-bound microtubules. This would then lead to the generation of unattached or partially unattached kinetochores causing checkpoint re-activation and mitotic arrest. The fact that MISP was initially identified because of its role in centrosome clustering indicates that forces that are important for spindle positioning, are also important for maintenance of spindle structure,,, and that MISP’s function must be finely regulated in mitosis to ensure tight control of both spindle structure and spindle position/orientation. Evidence from these recent studies indicates that such tight regulation occurs via phosphorylation, and that MISP is a CDK1-primed, substrate of Plk1. It is easy to envision how spindle positioning may be important in the context of a living organism/tissue, as abnormal orientation of the cell division plane would lead to disruption of tissue architecture. Equally important is the maintenance of spindle structure, as defects in spindle geometry, even if only transient, are a major cause of chromosome segregation errors, which can, in turn, lead to cell death or transformation. The evidence provided so far convincingly shows that MISP plays a key role in both spindle structure and positioning in tissue culture cells. What remains to be explored is whether this is also true in living organisms/tissues, and this should be the focus of future studies.

Figure 1. Schematic representation of MISP and its interactors in a rounded mitotic cell. Many different cell types round up during mitosis and form retraction fibers. During this process, focal adhesions are lost except at sites where retraction fibers persist, which also correspond to sites of enrichment of cortical actin. MISP (shown in yellow) may promote the orientation of the mitotic spindle, so that the two spindle poles are aligned with these regions of residual focal adhesions and high cortical actin density. A portion of the cell cortex (box) is shown as an enlarged view to the right, and shows that MISP promotes spindle positioning by simultaneously interacting with microtubule-associated proteins, such as dynein and EB1, cortical actin and focal adhesion-specific proteins, such as the focal adhesion kinase (FAK).