Defining the Contribution of ATR to MC1R-Enhanced DNA Repair in Melanocytes

The melanocyte stimulating hormone (MSH)-melanocortin-1 receptor (MC1R) signaling axis is an inducible pathway that regulates UV responses in melanocytes. Loss-of-function polymorphisms of MC1R signaling, common in fair-skinned UV-sensitive individuals, correlate with up to a four-fold lifetime melanoma risk. Signaling through the MC1R, a Gs-coupled membrane receptor, leads to activation of adenylyl cyclase and generation of the second messenger cAMP. Through cAMP generation, the MC1R pathway enhances the ability of melanocytes to repair UV-damaged DNA. If left unrepaired, UV photodamage causes characteristic “UV signature” mutations that fuel progression of melanocytes into melanoma. As a result, MC1R-defective individuals who have blunted DNA repair responses accumulate more photodamage after UV exposure and are therefore predisposed to melanoma. Our laboratory identified a critical molecular pathway linking MC1R/cAMP signaling to nucleotide excision repair (NER), the genome maintenance pathway responsible for removing UV damaged bases from DNA. Activated by MC1R signaling and cAMP generation, cAMP-dependent protein kinase (PKA) phosphorylates the ataxia and rad3 related (ATR) protein on the S435 residue. This post-translational modification causes ATR to associate with the NER factor xeroderma pigmentosum A (XPA) and accelerate its interaction with nuclear photodamage, thereby enhancing its repair. We have compelling evidence that A-kinase anchoring protein 12 (AKAP12) integrates PKA-ATR-XPA interactions. Experiments proposed in the first Aim will determine how AKAP12 regulates MC1R-enhanced NER in melanocytes. Studies proposed in the second Aim will identify how pS435 ATR impacts NER, focusing on mechanisms of DNA binding and strand incision. Finally, we will define how p-S435 ATR is inactivated in melanocytes since we have determined that dysregulated pS435 signal profoundly sensitizes cells to genotoxic agents. Together, these studies will clarify how MC1R signaling promotes NER and UV resistance and will serve as a platform for the development of rational melanoma-preventive strategies among high-risk MC1R-defective populations.