Activation of MLC by expression of a phosphomimetic MLC mutant (MLC-DD) enhances directional migration by creating a stable rear with thick actomyosin bundles and large, stable adhesions that inhibit ineffective lamellipodial protrusion. Arrows point to the direction of protrusion.

Image courtesy of Dr Miguel Vicente-Manzanares, University of Virginia, Charlottesville, USA.

How migrating cells establish a front-to-rear polarity axis remains controversial. Myosin-II proteins are actin-based motor proteins that are involved in different aspects of cell migration, from adhesion maturation to retraction of the cell edges. The isoforms MIIA and MIIB are found in most eukaryotic cells. In Journal of Cell Biology, Rick Horwitz and colleagues now shed light on how the polarity axis is established by clarifying the roles that myosin-IIA (MIIA) and myosin-IIB (MIIB) have in defining a cell front and rear, respectively.

The authors first observed that levels of phosphorylated myosin light chain (MLC), which activates myosin-II, were high in cells with a well-defined tail. To address whether activation of MLC and myosin-II cause the formation of an extended tail, a constitutively phosphorylated (active) form of MLC was expressed in cells lacking a prominent rear. Phosphorylated MLC accumulated at confined regions of the cell where it induced the local formation of actin bundles and large stable adhesions. Cell protrusion was inhibited in these regions, which instead formed a stable extended tail. Interestingly, directional cell migration was also enhanced, possibly by limiting the regions where protrusion can occur.

The authors then analysed the contribution of each myosin isoform to the formation of a cell rear. Expression of active MLC induced the formation of an extended rear only when MIIB was co-expressed. Conversely, cells did not form an extended rear upon MIIA activation. Thus, active MLC induces tail formation specifically via MIIB activation.

Visualization of the two myosin isoforms showed that MIIB accumulated at the centre and rear of cells, whereas clusters of MIIA initially appeared in cell protrusions but not at the leading edge. These clusters correlate with the formation of actin bundles and the maturation of adhesion. As cells move forward, this MIIA-rich region incorporates MIIB and becomes the rear. Importantly, large central and rear actin bundles did not form in the absence of MIIA, suggesting that MIIA is involved in the initial formation of actin cables that emerge at the front of migrating cells, whereas MIIB promotes the subsequent formation of thick actin bundles and large adhesions that inhibit protrusion.

Myosins are characterized by an actin-binding head domain and a C-terminal coiled-coiled domain that mediates dimerization. Use of MIIA-MIIB head-tail domain swap chimeras showed that both subcellular localization and specific functions — such as adhesion maturation at the front and formation of a stable tail at the back — relied on the C-terminal domain of MIIA and MIIB, respectively, rather than the actin-binding head domain.

Together this work shows that spatially segregated MIIA and MIIB play an essential role in cell front and rear formation, which is required for directional migration. However, the relationship between myosin-II function and microtubules — known to play a major role in the establishment of cell polarity — remains to be clarified.

Author: Kim Baumann