Abstract

Heterotrimeric guanine nucleotide-binding proteins (G proteins) of the Gɑ12/13 and Gɑq/11 subfamilies are broadly implicated in cancerous signaling and tumor progression. The primary goal of this study was to understand the amino acid structures within these G proteins that target them for palmitoylation, a lipid modification required for many of their intracellular functions. Cancerous signaling by G proteins requires the enzymatic attachment of the fatty acid palmitate to cysteine amino acids located in their N-terminal regions. Although the role of cysteines as attachment points for lipids is well established, little is known about the role of other, adjacent amino acids in G proteins to allow their recognition by palmitoyl transferase enzymes. A primary question is whether differences in these flanking amino acids bestow different mechanisms of lipid attachment upon different G proteins, such as Gɑq vs. Gɑ12/13. Engineering a “chimeric” Gɑ13 containing the N-terminal amino acids of Gɑq will allow for the use of a Gɑ13-driven oncogenic pathway in cells to study the effects of manipulating Gɑq-specific amino acids. Results from these experiments showed that mutating N-terminal cysteines of Gɑ13, Gɑq, or their chimera fully abolished cancerous signaling, and mutations adjacent to these amino acids had varying effects on this function. In subsequent experiments, additional lipidation sites were engineered in these proteins to assess their ability to “rescue” cancerous signaling that had been lost in these proteins by Cys mutation. Early results suggest that Gɑ13 and Gɑq have different lipid modification requirements. These findings will aid the development of inhibitory compounds that disrupt lipid attachment to specific G proteins in selected tumor types.

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