3.3 Structural distribution of cancer-associated GPCR mutations versus natural variants

It is notable that the ”hotspots”— well-defined mutation clusters—are not as common in GPCRs as in those oncogenes such as KRAS and tumor suppressor genes such us TP53, indicating a diverse landscape of genetic alterations (Baeissa, Benstead-Hume, Richardson, & Pearl, 2017). Part of the diversity originates from the non-synonymous natural variants, which represent genetic alterations in GPCRs that result in amino acid changes in healthy people. These variants play a significant role in the functional diversity observed among GPCRs across different individuals and populations. Across all GPCR families, there is a higher prevalence of non-synonymous natural variants in the N-terminus, C-terminus, and transmembrane (TM) domains compared to the extracellular or intracellular loops (A. Lee et al. , 2003). In addition, the highly conserved DRY and NPxxY motifs have been identified in the non-synonymous polymorphism analysis, which indicates that mutations in these structural motifs are inherent features in the diversity of GPCR function across different individuals and populations (Kim, Duc, & Chung, 2018). However, even with correction for natural variants, recent pan-cancer analysis has demonstrated that GPCRs still feature significant accumulation of mutations in some highly conserved structural motifs such as E/DRY, CWxP, NPxxY of class A GPCR, and HETx, GWGxP, PxxG of class B GPCR (Bongers et al. , 2022; Do, Haldane, Levy, & Miao, 2022). Bongers et al. found that conserved residues undergo a higher mutational pressure in cancer patients, which was not observed in natural variants, indicating their importance in cancer progression.
Most of the conserved motifs in GPCRs mediate their inactive conformation, and mutations at these motifs would therefore alter receptor function and stability. For example, the ‘ ’E/DRY’ ’ motif plays a pivotal role in receptor activation and signaling of class A GPCRs (Rovati, Capra, & Neubig, 2007). The ionic lock formed by the aspartic acid and the glutamic acid residue stabilizes the inactive state of the receptor. Upon ligand binding, conformational changes disrupt this ionic lock, allowing the transition to the active state and initiate downstream signaling. Conformational changes caused by mutations in the E/DRY motif could lead to alternations of receptor function, including gain of constitutive activity or loss of function (Huang & Tao, 2014; Römpler, Yu, Arnold, Orth, & Schöneberg, 2006). For example, cancer-associated CCR2 mutations in the DRY motif lead to a reduction or complete absence in G protein activation (den Hollanderet al. , 2023). Similar phenotype has been observed for the muscarinic acid (M1 and M5) receptors (Lu, Curtis, Jones, Pavia, & Hulme, 1997), gonadotropin-releasing hormone (GnRH) receptor (K. K. Arora, Cheng, & Catt, 1997), cannabinoid 2 receptor (CB2R) (Feng & Song, 2003), and the adrenergic receptors (Chung et al. , 2002; Samama, Cotecchia, Costa, & Lefkowitz, 1993). In addition, mutations outside the conserved motifs may also affect receptor function. One example is the N-terminal TSHR mutation found in toxic thyroid adenomas, which resulted in basal activation of the protein kinase A pathway (Nanba et al. , 2012).