The Role of the Lipid Raft-Associated Protein Flotillin-2 during Development and Progression of Myeloid Leukaemia

Introduction

The role of the bone marrow (BM) microenvironment (BMM) for the regulation of leukaemic stem cells (LSC) and their respective interplay is slowly being elucidated, but knowledge about how membrane structures or adhesion molecules on leukaemia cells may, specifically, interact with the BMM leading to regulation of leukemia cell maintenance and proliferation is limited. Flotillin (flot)-1 and -2 are highly conserved, ubiquitously expressed proteins localised in lipid microdomains in cellular membranes. These dynamic microdomains can serve as platforms for signal transduction, endocytosis and interactions with the actin cytoskeleton, as well as cell adhesion.

Hypothesis

As flotillins in the leukaemia cell membrane are associated with adhesion molecules known to be involved in the engraftment of leukaemia-initiating cells (LIC), we hypothesized that flotillins may play a role in LIC engraftment in the BMM.

Results

Using the murine retroviral transduction/transplantation model of BCR-ABL1-driven chronic myeloid leukaemia (CML) we observed a significant prolongation of survival of recipients of BCR-ABL1⁺, flot-2-deficient bone marrow compared to the respective control. The homing of BCR-ABL1⁺LIC to wild-type recipient bone marrow was reduced. Consistent with the known association of flotillins in the cell membrane with P-selectin glycoprotein ligand-1, a selectin ligand similar to CD44, both of which are known to be involved in the engraftment of CML LIC (Krause DS et al., Nat Med, 2006, and Krause DS et al., Blood, 2014), we demonstrated that flot-2 physically interacts and co-localizes with CD44. Flot-2-deficiency led to a significant reduction of the expression of CD44 and an impairment of the cytoskeleton in BCR-ABL1⁺ cells, as well as impaired migration. Further, Cdc42, a member of the Rho-GTPase family,was differentially distributed in wildtype versus flotillin-2-deficient BCR-ABL1⁺ LSC and may have compromised activity. Consistently, intrafemoral transplantation of flot-2-deficient CML-initiating cells or coexpression of CD44 in flot-2-deficient bone marrow ,rescued' or restored the CML-like disease.

In contrast, in an MLL-AF9-driven model of acute myeloid leukemia (AML) a similar prolongation of survival or reduced homing of AML-initiating cells were not observed. However, expression of CD44, known to play a role for engraftment in this disease (Jin L et al., Nat Med, 2006), in AML-initiating cells was similarly reduced and migration, adhesion and the cytoskeleton was similarly compromised.We demonstrated that this might be due to compensatory upregulation of C-X-C chemokine receptor type 4 (CXCR-4), the receptor for stromal-derived factor (SDF)-1a.

Conclusions

In summary, these data suggest that lipid raft molecules, particularly flotillins, play a previously unknown and possibly leukaemia-specific role in CML progression via modification of the levels and function of CD44 and possible regulation of Cdc42.