Scientists at the Institute of Cancer Research (ICR) in London have discovered that melanoma cells spread by harnessing a gene normally involved in the development of the nervous system.

The gene, ARHGEF9, encodes collybistin, a neuronal synaptic protein. When hijacked by melanoma cells, collybistin promotes the formation of membrane protrusions or filopodia that act as "molecular drills," attaching to and then punching holes through surrounding cells and structures.

Inhibiting ARHGEF9 destabilized the filopodia and prevented melanoma cells from spreading, while upregulating the gene led to the growth of more and longer filopodia.

The researchers say ARHGEF9 is likely to be involved in the spread of other types of cancer and suggest it represents a drug target for preventing metastases. They also found a further 13 genes that were involved in regulating the morphology of melanoma cells. The findings were published in iScience on August 8, 2022.

"This research has made a fundamental discovery about how melanoma cells manipulate their shape to become invasive – the first step towards metastasis," said Clare Isacke, dean of academic and research affairs at the ICR. "Although it is early research, and more work needs to be done, by understanding more about how skin cancer spreads, we could open up new avenues for developing treatments," she said.

During metastasis, melanoma cells invade tissue of diverse rigidities through dynamic alterations in their shape. However, to date most understanding of how cell shape is determined comes from 2D studies on extremely stiff substrates, which the researchers say may not provide physiologically relevant matrices.

To investigate the effect of matrix rigidity, the researchers compared the effects on cell morphology of silencing Rho GTP exchange factors and Rho GTPase activating proteins that are known to be involved in a range of cellular functions, including cell polarity, adhesion and cell motility, in cells cultured in 3D on soft collagen hydrogels, with cells cultured in 2D rigid matrices.

They found that WM266-4 metastatic human melanoma cells cultured in 2D and 3D matrices form extensive filopodia that mark sites of focal adhesions by which the cytoskeleton of a cell connects to the extracellular matrix.

Assessing the impact of selective gene silencing on melanoma cell morphology, the researchers found nine genes that regulated shape on soft matrix and five that regulated shape on stiff matrix. Depletion of ARHGEF9 was particularly effective in altering cell shape in melanoma cells on soft matrices. Only ARHGEF9 and intersectin 2 (ITSN2), a gene involved in actin remodelling, regulated melanoma cell shape in both soft and stiff matrices.

Further investigation of the role of ARHGEF9 showed it is essential for filopodia formation, focal adhesion assembly, contractility and cell protrusion in melanoma cells.

In the absence of ARHGEF9-mediated filopodia formation, focal adhesion morphogenesis is disrupted, causing adhesions to lose traction.

The researchers propose that melanoma cells use ARHGEF9 to promote filopodia-driven adhesion during morphogenesis in environments with varying rigidity and geometry.

Conserved function

To confirm this mechanism they performed further experiments in mouse melanoma cutaneous cells, observing ARHGEF9-depleted mouse melanoma cells were devoid of filopodia, and were far less contractile than control cells. In addition, ARHGEF9 inhibition in mouse cells significantly blocked their ability to invade collagen matrices.

They conclude that ARHGEF9 has a conserved function in promoting actin reorganization, adhesion and contractility across mouse and human cells.

"ARHGEF9 may thus confer melanoma cells with the ability to invade diverse tissue during metastasis, in a similar manner to how endothelial or neuronal cells migrate through the body during development," the researchers say.

Malignant melanoma is highly metastatic. Patients with advanced disease have a median survival of 6-10 months and a 5-year survival of less than 5%. Understanding the mechanisms controlling melanoma metastasis is essential to the development of new treatments.

"Our work shows that melanoma cells borrow use of ARHGEF9 from nerve cells to change shape, branch out and invade new tissues," said Chris Bakal, professor of cancer morphodynamics at the ICR. The next step will be to look at the broader impacts of inhibiting ARHGEF9 to explore whether it could be suitable drug target.

Defects inARHGEF9in the nervous system can lead to epilepsy and neurodevelopmental disorders including hyperactivity, anxiety and autism. The researchers thinkARHGEF9also could play a role in the development of the childhood cancer neuroblastoma, which develops from neuroblasts left behind following the development of the central nervous system in the fetus.