Our long-term goal is to understand the regulation of cell growth by cAMP-dependent protein kinase (PKA) by an in-depth investigation of the structure, function and expression of the regulatory (R) and catalytic (C) subunits of PKA. PKA is a key receptor protein for cAMP signals, transducing the signals inside mammalian cells. Importantly, there are two isoforms, type I (PKA-I) and Type II (PKA-II), and expression of these PKA isoforms can change dramatically during cell development, differentiation and transformation. PKA-I and PKA-II contain different R subunits, RI and RII, respectively, but share a common C subunit. We have investigated the following: (1) The functional distinction between PKA-I and PKA-II by structure-function analysis using site-directed mutagenesis and overexpression/suppression of the R and C subunit genes; (2) The cross-talk of PKA with other signaling pathway components in accordance with alteration of PKA expression through overexpression/suppression of the R and C subunit genes; and (3) The cooperative/antagonistic function of PKA with other cellular regulatory proteins and signal receptors by biochemical and immunocytochemical analysis. For functional distinction analysis of PKA isozymes, we targeted the RIa subunit with antisense. LS-174T colon carcinoma and LNCap prostate carcinoma cells mainly express RIa, RIIa and Ca subunits of PKA and the RIIb subunit at undetectable level. The loss of RIa by the antisense resulted in rapid increase in RIIb and PKA-IIb holoenzyme. Pulse-Chase experiments demonstrated that RIIb protein increased its half-life 5.5 fold in antisense treated cells (control, RIIb t 1/2, 2h; antisense treated, RIIb t 1/2, 11h). Thus, RIIb in the holoenzyme complex is stabilized, exhibiting an increased half-life. Through this biochemical adaptation, in the antisense-treated cancer cells, the ratio of PKA-I to PKA-II changes to become similar to that of normal cells. These studies rigorously demonstrated that RIa/PKA-I expression promotes cell proliferation, and that the compensatory stabilization of RIIb protein represents an important biochemical mechanism of RIa antisense that ensures depletion of PKA-I leading to sustained inhibition of tumor cell growth. In summary, our results indicate that RIa/PKA-I is a positive regulator of cell growth while RIIb/PKA-IIb (RIIb- containing PKA) is a growth inhibitory protein, supporting our hypothesis (Cho-Chung, Cancer Res. 50:7093-7100, 1990). Our results also imply that other signaling pathways that regulate cell growth most likely involve PKA chaperone. - Cancer, Differentiation, cAMP-dependent protein kinase, growth arrest, cyclic AMP analog, antisense, Apoptosis, EGF receptor,

Agency
National Institute of Health (NIH)
Institute
National Cancer Institute (NCI)
Type
Intramural Research (Z01)
Project #
1Z01BC005216-28
Application #
6289079
Study Section
Special Emphasis Panel (LTIB)
Project Start
Project End
Budget Start
Budget End
Support Year
28
Fiscal Year
1999
Total Cost
Indirect Cost
Name
National Cancer Institute Division of Basic Sciences
Department
Type
DUNS #
City
State
Country
United States
Zip Code
Nesterova, M; Johnson, N; Cheadle, C et al. (2006) Autoantibody biomarker opens a new gateway for cancer diagnosis. Biochim Biophys Acta 1762:398-403
Cho-Chung, Yoon S (2006) Autoantibody biomarkers in the detection of cancer. Biochim Biophys Acta 1762:587-91
Nesterova, Maria V; Cho-Chung, Yoon S (2004) Antisense protein kinase A RIalpha inhibits 7,12-dimethylbenz(a)anthracene-induction of mammary cancer: blockade at the initial phase of carcinogenesis. Clin Cancer Res 10:4568-77
Cho-Chung, Yoon S (2004) Antisense protein kinase A RI alpha-induced tumor reversion: portrait of a microarray. Biochim Biophys Acta 1697:71-9
Cho-Chung, Yoon S (2003) cAMP signaling in cancer genesis and treatment. Cancer Treat Res 115:123-43
Cho, Yee Sook; Cho-Chung, Yoon S (2003) Antisense protein kinase A RIalpha acts synergistically with hydroxycamptothecin to inhibit growth and induce apoptosis in human cancer cells: molecular basis for combinatorial therapy. Clin Cancer Res 9:1171-8
Kim, Young Hoon; Lim, Do Sun; Lee, Ji Hye et al. (2003) Gene expression profiling of oxidative stress on atrial fibrillation in humans. Exp Mol Med 35:336-49
Cheadle, Chris; Cho-Chung, Yoon S; Becker, Kevin G et al. (2003) Application of z-score transformation to Affymetrix data. Appl Bioinformatics 2:209-17
Cho-Chung, Yoon S (2003) CRE-enhancer DNA decoy: a tumor target-based genetic tool. Ann N Y Acad Sci 1002:124-33
Mani, S; Goel, S; Nesterova, M et al. (2003) Clinical studies in patients with solid tumors using a second-generation antisense oligonucleotide (GEM 231) targeted against protein kinase A type I. Ann N Y Acad Sci 1002:252-62

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