The regulated expression of cell surface receptors is essential for cellular homeostasis and defective in disease states. Receptor regulation is particularly important as receptor internalization limits the duration and intensity of substrate signaling. Interferon-γ (IFN-γ), a critical antiviral cytokine, transduces its signal through the interferon-γ receptor (IFNGR), a single-pass heterodimeric receptor composed of IFNGR1 and IFNGR2. While the IFN-γ signal transduction pathway is well studied, the cell surface regulation of the IFNGR is less well characterized.

To gain a better understanding of the ligand-independent regulation of cell surface IFNGR, I performed a CRISPR/Cas9 genome-wide genetic screen to identify genes critical for IFNGR regulation. Loss of CHIC2 or STUB1, also known as CHIP, increased IFNGR1 cell surface expression and their role in receptor regulation was further characterized. CHIC2 is a poorly studied, cysteine-rich protein, while the cytosolic E3 ligase, STUB1 is well-recognized for its role in the ubiquitination, internalization and lysosome-mediated degradation of cell surface proteins. To determine if CHIC2 also plays a more generalized role in cell surface protein regulation, I performed ‘plasma membrane profiling’, an unbiased, quantitative proteomic approach to compare the cell surface protein abundance in wildtype vs CHIC2 knockout cells. CHIC2 deficiency increased cell surface expression of 61 proteins in THP-1 cells, 43% of which were also STUB1 regulated, suggesting a critical role for CHIC2 in receptor regulation. In the absence of CHIC2, less cell surface IFNGR1 was internalized and the increased cell surface IFNGR1 expression potentiated IFN-γ-mediated signaling, confirming CHIC2’s functional importance.

Chromosomal translocation of CHIC2 is associated with hematological malignancies, but its endogenous function is unknown. I showed that palmitoylation of CHIC2 is essential for function as CHIC2’s membrane insertion is required for interacting with STUB1 and regulation of the IFNGR.  STUB1 is likely recruited to IFNGR1 through an exposed hydrophobic region that binds Hsp70, an interaction facilitated by CHIC2. Furthermore, CHIC2 is monoubiquitinated by STUB1 and internalized from the plasma membrane in a STUB1-dependent manner. CHIC2’s ubiquitination is essential for its internalization and likely affects IFNGR1 internalization, though the exact details of this interaction remain unclear. I therefore propose a model whereby membrane-associated CHIC2 recruits the soluble STUB1 E3 ligase to the plasma membrane, allowing STUB1 to associate with, ubiquitinate and internalize IFNGR1, and other receptor substrates. CHIC2 therefore plays a novel and essential role in the regulated expression of multiple cell surface receptors.