Maximizing tumor cell invasion

Abstract

The Rho family of proteins, comprised of Rho, Rac, and Cdc42 is implicated actomyosin contractility and microtubule arrangement (Ridley, 2001; Scott, 2002). RhoA, B and C activate Rho kinase (ROCK), which causes the formation of actin stress fibers by reinforcing actomysin contractile force. Rac activation, on the other hand, results in lamellipodia formation (Ridley, 2001). Tumor cell migration is highly dependent on cellular actin cytoskeleton dynamics. In order for cancer cells to metastasize, the primary tumor must invade and migrate through tissues so as to reside in a secondary location. The invasion process can either be protease-dependent or -independent. To date, cancer cells exhibit two forms of cellular movement: mesenchymal that involves elongated cells and amoeboid that involves rounded blebbing cells. The mesenchymal mode of invasion engages the extracellular matrix remodeling by proteases, whereas the amoeboid movement is proteases-independent; instead, it relies on extracellular matrix remodeling by force (Sahai and Marshall, 2003). Ameoboid movement is a result of ROCK activation as it requires high actomyosin contractibility. Although aberrant small Rho-GTPase activation has been linked to tumor cell metastasis, little is known about how tumor cells switch from mesenchymal to amoeboid modes of movement or vice versa. Sanz-Moreno et al. (2008) unveiled the factors that control the different modes of cell mobility through a systematic guanine nucleotide exchange factors (GEFs) siRNA screening. The study aimed to identify GEFs that regulate cell morphology and movement in order to understand the interchange between two different modes of movement (mesenchymal and amoeboid movement) in melanoma cells. From the screen, DOCK3 was discovered to control elongated movement. DOCK3 is a Rac specific GEF that, together with NEDD9, regulates Rac and WAVE2 to initiate mesenchymal movement, thus inhibiting amoeboid movement. Conversely, amoeboid movement requires RhoA activation of ROCK and ARHGAP, which in turn inactivates mesenchymal movement. The siRNA based screen was first tested in A375M2 cell line, a low metastatic melanoma cells that are in amoeboid and mesenchymal transition. From the 83 GEFs that were targeted, the suppression of DOCK3 expression resulted in the exhibition of an ameoboid morphology, and the inhibition of the transition from the amoeboid to mesenchymal mode. In this study, Sanz-Moreno et al. also found that inhibition of ROCK activity resulted in a conversion of the round cells to elongated cells, although it did not affect the speed of cancer cell migration. DOCK3’s effect is specific, as other DBL families do not cause cell elongation. In addition, knock-down of DOCK3 or expression of an internal deletion mutant, DELDOCK3, suppressed the conversion of the rounded cells to elongated cells. Similarly, Rac1 inhibition by siRNA or by inhibitor also abolished the elongated morphology. Taken together Sanz-Moreno et al. have demonstrated that Rac signaling is essential for the elongated morphology whereas ROCK activation is essential for the amoeboid phenotype. It was previously reported that DOCK3 forms complexes with NEDD9 (Kim et al., 2006), and NEDD9 ablation results in a similar phenotype to that of DOCK3 suppression. NEDD9 is over-expressed in metastatic human melanomas; hence Rac signaling is amplified in cancer cells. Sanz-Moreno et al. observed that Rac promotes the elongated cell morphology by suppressing actomyosin contractility through WAVE2 in an uncharacterized mechanism. In contrast, ROCK activation activates ARHGAP22, which inhibits Rac activity and results in the amoeboid phenotype. They further studied the role of NEDD9-DOCK3-Rac complex in cell invasion. Inhibition of Rac signaling promotes the cell to invade by amoeboid movement. However, the inhibition of ROCK-ARHGAP22 signaling caused the cell to invade in a mesenchymal fashion. It is well established that the process of metastasis involves the migration of the primary tumor to another location via blood vessels or the lymphatic system to form secondary tumors. Next, the authors compared the metastatic ability of cells with mesenchymal movement versus amoeboid movement. WM1361 melanoma cells were utilized in this study as this cell line has a higher proportion of elongated cells and is more effective in entering the lung after tail vein injection. Tail vain injection of WM1361 cells with depletion of Rac1 or DOCK3 enhanced amoeboid movement favoring colonization in the lung, whereas elevated Rac activity compromised the ability of cells to colonize the lung, which suggests that amoeboid movement is more effective in cell invasion. Collectively, NEDD9-DOCK3-Rac promotes mesenchymal migration and suppresses amoeboid movement. On the other hand, ROCK-ARHGAP22 signaling is able to suppress the mesenchymal migration. As shown in Figure 1, the interplay between the two phenotypes allows cancer cells to invade in different microenvironments and migrate to different tissues. Cancer metastasis is the main cause of cancer death; therefore, drug design that is targeted towards inhibiting the Rac or ROCK pathway might present a new mechanism to cure cancer. Interchange between the mesenchymal-type and amoeboid-type movement. The proteins that are involved in the switch between the two modes of cancer cells movement. The amoeboid-type movement (rounded blebbing cell phenotype) is controlled by the Rho-ROCK-ARHGAP22 activation that results in actomyosin contraction and promotes cell movement through mechanical forces. The mesenchymal-type movement (fibroblast-like elongated cell phenotype) is the result of activation NEDD9-DOCK3-Rac complex and WAVE2, which results in actin polymerization.