Abstract LB-081: A nuclear cAMP microdomain functions as a tumor suppressor in melanoma
Authors: Marek M. Drozdz, Jenny Wang, Michael Reilly, Elsbeth Kane, Kelsey Aguirre, Ashley Doane, Jedd D. Wolchok, Olivier Elemento, Garrett Desman, Taha Merghoub, Elena Piskounova, Jonathan Zippin
Journal: Cancer Research (2019)

Abstract Cyclic AMP (cAMP), a ubiquitous second messenger, has a broad impact on cellular processes such as proliferation, differentiation, apoptosis, migration, and gene expression. It has been widely linked to melanomagenesis; however, studies often report contradictory functions for cAMP (e.g., cancer promotion, but also tumor suppression and therapy resistance). Thus, a huge conundrum exists in the field of cAMP signaling and melanoma – one that might be explained by examining cAMP at the level of microdomains. In mammalian cells, two classes of adenylyl cyclases synthesize cAMP: the transmembrane adenylyl cyclases (tmACs) and the soluble adenylyl cyclase (sAC). Unlike tmACs, which are tethered to the plasma membrane, sAC is expressed throughout the cell and within organelles (e.g., nucleus and mitochondria). Thus, sAC and tmACs define distinct sources that generate cAMP in spatially separated cell regions, further restricted by the cAMP catabolizing enzymes phosphodiesterases (PDEs). Since cAMP signaling cascades can be evoked and restricted to small compartments, it is imperative that we interrogate all available cAMP microdomains to truly unravel their role in melanoma. To address this question, we have created a panel of virally-encoded, doxycycline-inducible sAC cDNAs, targeted to specific cellular compartments, i.e. nucleus, cytoplasm or mitochondria. We infected a melanoma sAC knock-out cell line with these cDNAs and have established single clone cell lines where sAC activity can be rapidly restored, only in a specific microdomain, at near pharmacologic time scales. Utilizing these model cell lines, we showed that only the nuclear sAC-dependent cAMP microdomain inhibits melanoma growth, both in vitro and in mice. Neither mitochondrial, nor cytoplasmic sAC cAMP microdomains affected tumor cell growth. Using ATACseq and RNAseq, we discovered that the nuclear cAMP microdomain leads to specific changes in chromatin landscape and a unique gene expression profile, distinct from the canonical MC1R/cAMP-dependent gene expression profile, that encompasses a range of genes with both established and potentially new roles in melanomagenesis. Immunohistochemical analysis of melanoma patient biopsies revealed that sAC localization in melanocytes is not static; while sAC is mostly cytoplasmic in benign melanocytes, it becomes nuclear as melanocytes undergo early malignant transformation (in situ disease and radial growth phase) but is lost from the nucleus upon transition to vertical growth phase. These data suggest that nuclear sAC expression is an early tumor suppressive pathway that is lost in melanoma cells capable of dermal invasion. Thus the pattern of cellular localization of sAC during different stages of melanoma may be a prognostic marker. Taken together, our findings support a novel tumor suppressor pathway that relies on the expression of sAC in specific cellular microdomains. Citation Format: Marek M. Drozdz, Jenny Wang, Michael Reilly, Elsbeth Kane, Kelsey Aguirre, Ashley Doane, Jedd D. Wolchok, Olivier Elemento, Garrett Desman, Taha Merghoub, Elena Piskounova, Jonathan Zippin. A nuclear cAMP microdomain functions as a tumor suppressor in melanoma [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2019; 2019 Mar 29-Apr 3; Atlanta, GA. Philadelphia (PA): AACR; Cancer Res 2019;79(13 Suppl):Abstract nr LB-081.