Hypoxic induction of AKAP12 variant 2 shifts PKA-mediated protein phosphorylation to enhance migration and metastasis of melanoma cells

Significance Scaffold proteins can serve as critical focal points for association of signaling molecules and downstream pathways that regulate tumor growth and invasion. We demonstrate that low oxygen levels, common in solid tumors, can regulate expression of one member of the AKAP scaffold protein...

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Published in:Proceedings of the National Academy of Sciences - PNAS 2015-04, Vol.112 (14), p.4441-4446
Main Authors: Finger, Elizabeth C., Castellini, Laura, Rankin, Erinn B., Vilalta, Marta, Krieg, Adam J., Jiang, Dadi, Banh, Alice, Zundel, Wayne, Powell, Marianne Broome, Giaccia, Amato J.
Format: Article
Language:English
Subjects:
A Kinase Anchor Proteins - genetics
A Kinase Anchor Proteins - metabolism
Amino Acid Sequence
Animals
Biological Sciences
Cell Cycle Proteins - genetics
Cell Cycle Proteins - metabolism
Cell Hypoxia
Cell Line, Tumor
Cell Movement
Cyclic AMP-Dependent Protein Kinases - metabolism
Gene Expression Regulation, Neoplastic
Humans
Hypoxia
Melanoma - metabolism
Melanoma - pathology
Metastasis
Mice
Molecular Sequence Data
Molecules
Neoplasm Invasiveness
Neoplasm Metastasis
Neoplasm Transplantation
Oxygen - metabolism
Phosphoproteins - metabolism
Phosphorylation
Proteins
Proteomics
Signal Transduction
Skin Neoplasms - metabolism
Tumors
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Summary:Significance Scaffold proteins can serve as critical focal points for association of signaling molecules and downstream pathways that regulate tumor growth and invasion. We demonstrate that low oxygen levels, common in solid tumors, can regulate expression of one member of the AKAP scaffold protein family, AKAP12, in melanoma. Genetic inactivation of AKAP12 leads to decreased migration, invasion, and tumor growth in a mouse model of melanoma. Mechanistically, we discovered a switch in protein kinase A (PKA)-regulated phosphorylations under hypoxia that are dependent on AKAP12 and show that PKA is the critical kinase regulating AKAP12-dependent cellular migration. These results provide novel insight into how the tumor microenvironment modulates signal transduction and biological responses through the regulation of a specific variant of the scaffold protein AKAP12. Scaffold proteins are critical hubs within cells that have the ability to modulate upstream signaling molecules and their downstream effectors to fine-tune biological responses. Although they can serve as focal points for association of signaling molecules and downstream pathways that regulate tumorigenesis, little is known about how the tumor microenvironment affects the expression and activity of scaffold proteins. This study demonstrates that hypoxia, a common element of solid tumors harboring low oxygen levels, regulates expression of a specific variant of the scaffold protein AKAP12 (A-kinase anchor protein 12), AKAP12v2, in metastatic melanoma. In turn, through a kinome-wide phosphoproteomic and MS study, we demonstrate that this scaffolding protein regulates a shift in protein kinase A (PKA)-mediated phosphorylation events under hypoxia, causing alterations in tumor cell invasion and migration in vitro, as well as metastasis in an in vivo orthotopic model of melanoma. Mechanistically, the shift in AKAP12-dependent PKA-mediated phosphorylations under hypoxia is due to changes in AKAP12 localization vs. structural differences between its two variants. Importantly, our work defines a mechanism through which a scaffold protein can be regulated by the tumor microenvironment and further explains how a tumor cell can coordinate many critical signaling pathways that are essential for tumor growth through one individual scaffolding protein.
ISSN:0027-8424
1091-6490
DOI:10.1073/pnas.1418164112