Missing-in-metastasis protein downregulates CXCR4 by promoting ubiquitylation and interaction with small Rab GTPases

Abstract

Surface expression of chemokine receptor CXCR4 is downregulated by missing-in-metastasis protein (MIM; also known as MTSS1), a member of the inverse BAR (I-BAR)-domain protein family that recognizes and generates membranes with negative curvature. Yet, the mechanism for the regulation is unknown. Here, we show that MIM forms a complex with CXCR4 by binding to E3 ubiquitin ligase AIP4 (also known as ITCH) in response to stromal cell-derived factor 1 (SDF-1; also known as CXCL12). Overexpression of MIM promoted CXCR4 ubiquitylation, inhibited cellular response to SDF-1, caused accumulation and aggregation of multivesicular bodies (MVBs) in the cytoplasm, and promoted CXCR4 sorting into MVBs in a manner depending on binding to AIP4. In response to SDF-1, MIM also bound transiently to the small GTPase Rab5 at 5 min and to Rab7 at 30 min. Binding to Rab7 requires an N-terminal coiled-coil motif, deletion of which abolished MIM-mediated MVB formation and CXCR4 internalization. Our results unveil a previously unknown property of MIM that establishes the linkage of protein ubiquitylation with Rab-guided trafficking of CXCR4 in endocytic vesicles.

Differential interactions of missing in metastasis and insulin receptor tyrosine kinase substrate with RAB proteins in the endocytosis of CXCR4

“…S1A). Taken together, these data are consistent with our previous finding based on HeLa cells and mouse cells that MIM down-regulates CXCR4-mediated chemotaxis (27).…”

“…MIM-GFP or GFP, colocalization of CD63 and CXCR4 was low and not significantly impacted by siRAB7. MIM promotes the formation of late endosomes (27), as indicated by increased CD63 puncta ( Fig. 2F).…”

“…We recently demonstrated that MIM promotes CXCR4 internalization in either primary mouse bone marrow cells or HeLa cells (25,27). As aged MIM-deficient mice were prone to lymphatic malignancies (4), we were interested in whether MIM plays a similar role in human lymphatic cells and analyzed the internalization of CXCR4 in several lymphocytic malignant cells, including Reh (acute lymphocytic leukemia), Raji (Burkitt's lymphoma), and Daudi (Burkitt's lymphoma) cells, and patient-derived human B lymphatic (PBL) cells.…”

“…Although the detailed mechanism of CXCR4 internalization remains elusive, the major events following exposure to its ligand SDF-1 include phosphorylation at the C terminus of the receptor and activation of the ubiquitin E3 ligase AIP4, which further leads to endocytic sorting of the receptor from early to late endosomes (26). Significantly, MIM interacts with the complex of AIP4 and CXCR4 upon SDF-1 stimulation and promotes CXCR4 ubiquitination and its sorting into late endosomes (27). Also, MIM associates sequentially with RAB5 and RAB7 during the response to SDF-1.…”

Missing-in-Metastasis/Metastasis Suppressor 1 Regulates B Cell Receptor Signaling, B Cell Metabolic Potential, and T Cell-Independent Immune Responses

“…It also directly interacts with and regulates actin via its C-terminal WASp homology 2 (WH2) domain (6,9) and indirectly via interactions with other actin regulatory proteins, such as cortactin and Rac1 GTPase (6,7,10,11). MIM has also been shown to regulate bone marrow and lymphoid cell trafficking presumably through regulation of CXCR4 internalization as seen in cancer cell lines (12)(13)(14)(15). Importantly, MIM has been linked to various cancers, either as a putative tumor metastasis suppressor or promoter (16).…”