N-glycosylation by Mgat5 imposes a targetable constraint on immune-mediated tumor clearance.

The regulated glycosylation of the proteome has widespread effects on biological processes that cancer cells can exploit. Expression of N-acetylglucosaminyltransferase V (encoded by Mgat5 or GnT-V), which catalyzes the addition of ß1,6-linked N-acetylglucosamine to form complex N-glycans, has been linked to tumor growth and metastasis across tumor types. Using a panel of murine pancreatic ductal adenocarcinoma (PDAC) clonal cell lines that recapitulate the immune heterogeneity of PDAC, we found that Mgat5 is required for tumor growth in vivo but not in vitro. Loss of Mgat5 results in tumor clearance that is dependent on T cells and dendritic cells, with NK cells playing an early role. Analysis of extrinsic cell death pathways revealed Mgat5-deficient cells have increased sensitivity to cell death mediated by the TNF superfamily, a property that was shared with other non-PDAC Mgat5-deficient cell lines. Finally, Mgat5 knockout in an immunotherapy-resistant PDAC line significantly decreased tumor growth and increased survival upon immune checkpoint blockade. These findings demonstrate a role for N-glycosylation in regulating the sensitivity of cancer cells to T cell killing through classical cell death pathways.

The dopamine D2 receptor can directly recruit and activate GRK2 without G protein activation.

The dopamine D2 receptor (D2R) is a G protein-coupled receptor (GPCR) that is critical for many central nervous system functions. The D2R carries out these functions by signaling through two transducers: G proteins and ß-arrestins (ßarrs). Selectively engaging either the G protein or ßarr pathway may be a way to improve drugs targeting GPCRs. The current model of GPCR signal transduction posits a chain of events where G protein activation ultimately leads to ßarr recruitment. GPCR kinases (GRKs), which are regulated by G proteins and whose kinase action facilitates ßarr recruitment, bridge these pathways. Therefore, ßarr recruitment appears to be intimately tied to G protein activation via GRKs. Here we sought to understand how GRK2 action at the D2R would be disrupted when G protein activation is eliminated and the effect of this on ßarr recruitment. We used two recently developed biased D2R mutants that can preferentially interact either with G proteins or ßarrs as well as a ßarr-biased D2R ligand, UNC9994. With these functionally selective tools, we investigated the mechanism whereby the ßarr-preferring D2R achieves ßarr pathway activation in the complete absence of G protein activation. We describe how direct, G protein-independent recruitment of GRK2 drives interactions at the ßarr-preferring D2R and also contributes to ßarr recruitment at the WT D2R. Additionally, we found an additive interaction between the ßarr-preferring D2R mutant and UNC9994. These results reveal that the D2R can directly recruit GRK2 without G protein activation and that this mechanism may have relevance to achieving ßarr-biased signaling.