"In medio stat virtus": Insights into hybrid E/M phenotype attitudes

Abstract

Epithelial-mesenchymal plasticity (EMP) refers to the ability of cells to dynamically interconvert between epithelial (E) and mesenchymal (M) phenotypes, thus generating an array of hybrid E/M intermediates with mixed E and M features. Recent findings have demonstrated how these hybrid E/M rather than fully M cells play key roles in most of physiological and pathological processes involving EMT. To this regard, the onset of hybrid E/M state coincides with the highest stemness gene expression and is involved in differentiation of either normal and cancer stem cells. Moreover, hybrid E/M cells are responsible for wound healing and create a favorable immunosuppressive environment for tissue regeneration. Nevertheless, hybrid state is responsible of metastatic process and of the increasing of survival, apoptosis and therapy resistance in cancer cells. The present review aims to describe the main features and the emerging concepts regulating EMP and the formation of E/M hybrid intermediates by describing differences and similarities between cancer and normal hybrid stem cells. In particular, the comprehension of hybrid E/M cells biology will surely advance our understanding of their features and how they could be exploited to improve tissue regeneration and repair.

Keywords: cancer; collective migration; hybrid E/M phenotype; immune evasion; immune suppression; metabolic reprograming; partial EMT; stem cell.

FIGURE 1

Partial EMT and Hybrid E/M Phenotypes. During partial EMT are generated several intermediated phenotypes which bear mixed epithelial (E) and mesenchymal (M) features. These states are called hybrid E/M phenotype and display particular surface markers such as CD106, CD51 and CD61. The loss of E phenotype coincides with an increase of stemness and plasticity that reaches its peak during the formation of hybrid states and decrease again when EMT is complete. Plasticity and stemness are at the base of hybrid cells peculiar features: collective migration, development and differentiation, regeneration, microenvironmental organization, metabolic plasticity, immune evasion, immune suppression and therapy resistance.

FIGURE 2

Gene regulatory networks (GRN) govern the fate of hybrid E/M cells. Schematic overview of the different GNR involved in the modulation of hybrid E/M phenotype decisions. (A) EMP circuit (yellow) is formed by the interaction of two mutually inhibitory feedback loops (miR-200/ZEB and miR-34/SNAIL) and regulates EMT/MET decisions. Stemness circuit (blue) integrate EMP circuit with another mutually inhibitory feedback loop formed by LIN28/let-7 and regulates the expression of stemness genes. Migration circuit (red) involves Rac1/RhoA interactions and is also integrated with EMP circuit thus regulating the switch between collective and individual migration. (B) Phenotypic Stability Factors (PSF) such as OVOL, GRHL2 and Nrf2 maintain the hybrid E/M phenotype, thus avoiding a complete EMT. PSF circuit (purple) is completely integrated in the EMP circuit and their interactions regulate the cell fate. (C) Coupling EMP circuit with Notch circuit (light green) which regulates cell-cell communication and cell asymmetry. (D) Notch-Delta signaling induce different fate in neighboring cells: one becomes Sender (high Delta/low Notch) and the other becomes Receiver (high Notch/low Delta). This behavior causes lateral inhibition and promotes the formation of the so-called “salt and pepper” pattern. Notch-Jagged signaling induces the same fate in neighboring cells which become Sender/Receiver (high Notch/high Jagged). This behavior causes lateral induction and promotes the formation of the similar pattern among neighboring cells. (E) Therefore, during lateral inhibition (up) cells in partial EMT might not be spatially close, thus promoting single cell asymmetry. Conversely, during lateral induction (down) cells in partial EMT can mutually stabilize the hybrid E/M phenotype, thus generating cluster symmetry (Figures 2C–E have been adjusted from Bocci et al., 2017).

FIGURE 3

Migratory plasticity. Primary epithelial tissue through partial EMT (pEMT) and complete EMT can give rise to motile cells capable of invasion via collective migration of hybrid E/M cells or individual migration of mesenchymal cells, respectively. Afterwards, specific microenvironment conditions govern reversible cell fates. On one hand, cluster of hybrid E/M cells can undergo collective-individual transition either by completing EMT process and becoming mesenchymal cells or via collective-amoeboid transition (CAT) and giving rise to amoeboid cells. The reverse process is called amoeboid-collective transition (ACT). Amoeboid and mesenchymal cells can reversibly interchange phenotype via amoeboid-mesenchymal (AMT) and mesenchymal-amoeboid transition (MAT), respectively. Partial AMT and partial MAT give rise to hybrid A/M cells with mixed amoeboid and mesenchymal features. On the other hand, mesenchymal cells via pEMT can either undergo individual-collective migration giving rise to a cluster of migratory hybrid E/M cells or revert their phenotype via MET and becoming epithelial cells. All the transitions rely on the levels of two small GTPases, RhoA and Rac1, which can be upregulated (green) or downregulated (red) depending on the cell phenotype.

FIGURE 4

Immune escape and immune suppression mechanisms of hybrid E/M cells. Immune evasion is a mechanism thereby hybrid E/M cells can elude immune surveillance and killing by cytotoxic T lymphocyte (CTL) and Natural killer cells (NK). The main mechanisms involved include the downregulation of T cell receptor (TCR) and NKG2D ligands on hybrid E/M cell surface, the inhibition of the antigen presenting machinery and proteasome protein expression (light yellow box). On the other hand, hybrid E/M cells perform the immune suppression by increasing the release of immunosuppressive soluble mediators, upregulating the expression of immunosuppressive membrane proteins and inducing the differentiation of immune cells toward their immunosuppressive phenotypes such as anti-inflammatory M2 macrophages, regulatory T cells (Treg) and myeloid-derived suppressor cells (MDSC) (light blue box). These cells, in turn, have the ability to damp the immune response by inhibiting the activation of immune stimulatory cells (e.g., M1 macrophages, CTL), thus promoting the formation of an immunosuppressive microenvironments.