Signaling through cyclic AMP (cAMP) and its effector molecules, such as cAMP-dependent protein kinase (PKA) and exchange proteins activated by cAMP (Epac), regulates a variety of cellular functions including cell growth, proliferation, metabolism, survival and mobility, as well as insulin secretion in the case of pancreatic ? cells. The overall goal of our research is to elucidate the molecular mechanisms and functional roles of spatiotemporal regulation in achieving high specificity in cAMP signaling. Aberrations in the cAMP signaling pathway have implications for clinical conditions such as obesity and type 2 diabetes mellitus, particularly in the context of ?-cell functions. A mechanistic understanding of cAMP signaling specificity is crucial to developing therapeutic strategies for these clinical conditions. The concept of spatial compartmentalization of cAMP effects was proposed 20 years ago, but only in recent years have innovative approaches to studying cAMP signaling in the cellular context become available to provide direct mechanistic evidence. However, despite these recent advances, there are still large gaps in our understanding about the mechanisms underlying the spatiotemporal regulation of cAMP and its effectors. Furthermore, little is known about how the signaling information encoded in the spatiotemporal patterns of activities is translated into specific functional responses. In our preliminary studies, we have developed new molecular tools to monitor and perturb cAMP/PKA activities in living cells with further enhanced spatiotemporal resolution and precision. Furthermore, building on our recent discovery of a Ca2+-cAMP-PKA oscillatory circuit in MIN6 ? cells, we showed that A-Kinase Anchoring Protein 79/150 (AKAP79/150) assembles a PKA-containing signaling complex in these cells and influences the activity dynamics of the Ca2+-cAMP-PKA circuit. In the current proposal, utilizing our new molecular tools and by combining computational modeling and experimental approaches, we will test our hypothesis that the Ca2+-cAMP-PKA oscillatory circuit, further regulated spatially and temporally by AKAPs, allows PKA to achieve high signaling specificity and diversity through frequency modulation.
The specific aims are: 1) developing novel molecular tools to interrogate the spatiotemporal regulation of cAMP/PKA signaling in living cells; 2) elucidating the spatial compartmentalization and frequency control of the Ca2+-cAMP-PKA oscillatory circuit.

Public Health Relevance

Signaling through cAMP regulates a variety of cellular functions such as cell growth, proliferation, metabolism, survival and mobility, as well as insulin secretion in the case of pancreatic ? cells. The overall goal of our research is to achieve a better mechanistic understanding about how this ubiquitous signaling molecule regulates many diverse functions with high signaling specificity. Such an understanding is crucial to developing therapeutic strategies for clinical conditions arising from dysregulated cAMP signaling such as type II diabetes.

Agency
National Institute of Health (NIH)
Institute
National Institute of Diabetes and Digestive and Kidney Diseases (NIDDK)
Type
Research Project (R01)
Project #
5R01DK073368-12
Application #
9312288
Study Section
Special Emphasis Panel (ZRG1)
Program Officer
Silva, Corinne M
Project Start
2006-01-15
Project End
2021-06-30
Budget Start
2017-07-01
Budget End
2018-06-30
Support Year
12
Fiscal Year
2017
Total Cost
Indirect Cost
Name
University of California, San Diego
Department
Pharmacology
Type
Schools of Medicine
DUNS #
804355790
City
La Jolla
State
CA
Country
United States
Zip Code
92093
Ni, Qiang; Mehta, Sohum; Zhang, Jin (2018) Live-cell imaging of cell signaling using genetically encoded fluorescent reporters. FEBS J 285:203-219
Mehta, Sohum; Zhang, Yong; Roth, Richard H et al. (2018) Single-fluorophore biosensors for sensitive and multiplexed detection of signalling activities. Nat Cell Biol 20:1215-1225
Hertel, Fabian; Mo, Gary C H; Dedecker, Peter et al. (2018) Observing the Assembly of Protein Complexes in Living Eukaryotic Cells in Super-Resolution Using refSOFI. Methods Mol Biol 1764:267-277
Ross, Brian L; Tenner, Brian; Markwardt, Michele L et al. (2018) Single-color, ratiometric biosensors for detecting signaling activities in live cells. Elife 7:
Newman, Robert H; Zhang, Jin (2017) Integrated Strategies to Gain a Systems-Level View of Dynamic Signaling Networks. Methods Enzymol 589:133-170
Rodriguez, Erik A; Campbell, Robert E; Lin, John Y et al. (2017) The Growing and Glowing Toolbox of Fluorescent and Photoactive Proteins. Trends Biochem Sci 42:111-129
Mehta, Sohum; Zhang, Jin (2017) Illuminating the Cell's Biochemical Activity Architecture. Biochemistry 56:5210-5213
Gorshkov, Kirill; Mehta, Sohum; Ramamurthy, Santosh et al. (2017) AKAP-mediated feedback control of cAMP gradients in developing hippocampal neurons. Nat Chem Biol 13:425-431
Mo, Gary C H; Ross, Brian; Hertel, Fabian et al. (2017) Genetically encoded biosensors for visualizing live-cell biochemical activity at super-resolution. Nat Methods 14:427-434
Nygren, Patrick J; Mehta, Sohum; Schweppe, Devin K et al. (2017) Intrinsic disorder within AKAP79 fine-tunes anchored phosphatase activity toward substrates and drug sensitivity. Elife 6:

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