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Special Issue on Cell Biology of the Lysosome

The ubiquitin ligases Cbl and Cbl-b regulate macrophage growth by controlling CSF-1R import into macropinosomes

Published Online:https://doi.org/10.1091/mbc.E23-09-0345
new hypothesis

The ubiquitination of transmembrane receptors regulates endocytosis, intracellular traffic, and signal transduction. Bone marrow−derived macrophages from myeloid Cbl–/– and Cbl-b–/– double knockout (DKO) mice display sustained proliferation mirroring the myeloproliferative disease that these mice succumb to. Here, we found that the ubiquitin ligases Cbl and Cbl-b have overlapping functions for controlling the endocytosis and intracellular traffic of the CSF-1R. DKO macrophages displayed complete loss of ubiquitination of the CSF-1R whereas partial ubiquitination was observed for either single Cbl–/– or Cbl-b–/– macrophages. Unlike wild type, DKO macrophages were immortal and displayed slower CSF-1R internalization, elevated AKT signaling, and a failure to transport the CSF-1R into the lumen of nascent macropinosomes, leaving its cytoplasmic region available for signaling. CSF-1R degradation depended upon lysosomal vATPase activity in both WT and DKO macrophages, with this degradation confined to macropinosomes in WT but occurring in distributed/tubular lysosomes in DKO cells. RNA-sequencing comparison of Cbl–/–, Cbl-b–/– and DKO macrophages indicated that while the overall macrophage transcriptional program remained intact, DKO macrophages had alterations in gene expression associated with growth factor signaling, cell cycle, inflammation and senescence. Cbl-b–/– had minimal effect on the transcriptional program whereas Cbl–/– led to more alternations but only DKO macrophages demonstrated substantial changes in the transcriptome, suggesting overlapping but unique functions for the two Cbl-family members. Thus, Cbl/Cbl-b-mediated ubiquitination of CSF-1R regulates its endocytic fate, constrains inflammatory gene expression, and regulates signaling for macrophage proliferation.

  • Macrophages form macropinosomes in response to signaling from the CSF-1R. The CSF-1R is sequestered in the lumen of macropinosomes, however, the mechanism regulating this step and its importance in CSF-1R signaling is unknown.

  • The authors found that overlapping ubiquitination activity of Cbl and Cbl-b are critical for sequestration of the CSF-1R within macropinosomes, shaping CSF-1R signaling and controlling macrophage proliferation and preventing senescence.

  • These findings define a new function for macropinosomes and Cbl and Cbl-b in regulating growth factor signaling with implications for cancer and immunology.