It is well established that the phorbol esters, natural tumor promoters that mimic the actions of the lipid second messenger diacylglycerol (DAG), activate protein kinase C (PKC), a family of serine-threonine kinases that play important roles in multistage carcinogenesis. The traditional view of PKC as the sole receptor for the phorbol esters has been challenged by the discovery of proteins unrelated to PKC, which have a C1 domain, the phorbol ester binding site in PKC. Indeed, we have established that """"""""chimaerins"""""""", a family of proteins with homology to Rac GAPs (GTPase Activating Proteins) are high affinity receptors for phorbol esters and DAG. Moreover, like PKCs, the beta2-chimaerin isoform is subject to translocation by phorbol esters in a PKC-independent fashion. Importantly, we found that beta2-chimaerin accelerates GTP hydrolysis from Rac1, leading to the inactivation of this GTPase. The central hypothesis of this competing renewal is that chimaerin activation by phorbol ester/DAG will impair Rac-mediated signaling and functions.
In Specific Aim 1 we will explore the complex interrelationship between DAG generation by growth factors, chimaerin translocation and Rac-GTP levels. We predict that translocation of chimaerins to the plasma membrane through its C1 domain will result in the activation of chimaerin Rac-GAP activity and Rac inactivation.
In Specific Aim 2 we will investigate how protein-protein interactions regulate chimaerin function, following analogous models to those described for PKC isozymes. We have isolated several chimaerin interacting proteins that play important roles in controlling intracellular localization and function of chimaerins. A second aspect of this aim will involve the characterization of phosphotyrosine proteins isolated in our laboratory that may associate to the beta2-chimaerin SH2 domain.
In Specific Aim 3 we will focus on the roles of chimaerin isoforms in mitogenic signaling. Since Rac has a central role in the control of mitogenicity, our hypothesis is that chimaerins, by inhibiting Rac function, will impair mitogenic signaling and proliferation. Since Rac is critical for actin cytoskeletal reorganization, Specific Aim 4 will explore if chimaerins regulate key steps of the metastatic cascade, including filament actin membrane structures, cell spreading, migration and invasion. Our research has the potential for delineating novel PKC-independent pathways for the phorbol esters and DAG and their relationship to proliferation, malignant transformation, and metastasis. More importantly, our research challenges the use of phorbol esters as selective PKC activators in cells.

Agency
National Institute of Health (NIH)
Institute
National Cancer Institute (NCI)
Type
Research Project (R01)
Project #
3R01CA074197-09S1
Application #
7232155
Study Section
Chemical Pathology Study Section (CPA)
Program Officer
Ogunbiyi, Peter
Project Start
1997-09-30
Project End
2008-04-30
Budget Start
2006-06-09
Budget End
2007-04-30
Support Year
9
Fiscal Year
2006
Total Cost
$66,106
Indirect Cost
Name
University of Pennsylvania
Department
Pharmacology
Type
Schools of Medicine
DUNS #
042250712
City
Philadelphia
State
PA
Country
United States
Zip Code
19104
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Dipaolo, Brian C; Davidovich, Nurit; Kazanietz, Marcelo G et al. (2013) Rac1 pathway mediates stretch response in pulmonary alveolar epithelial cells. Am J Physiol Lung Cell Mol Physiol 305:L141-53
Gutierrez-Uzquiza, Alvaro; Colon-Gonzalez, Francheska; Leonard, Thomas A et al. (2013) Coordinated activation of the Rac-GAP ?2-chimaerin by an atypical proline-rich domain and diacylglycerol. Nat Commun 4:1849
Caino, M C; Lopez-Haber, C; Kim, J et al. (2012) Proteins kinase C? is required for non-small cell lung carcinoma growth and regulates the expression of apoptotic genes. Oncogene 31:2593-600
Caino, M Cecilia; Lopez-Haber, Cynthia; Kissil, Joseph L et al. (2012) Non-small cell lung carcinoma cell motility, rac activation and metastatic dissemination are mediated by protein kinase C epsilon. PLoS One 7:e31714
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Riccomagno, Martin M; Hurtado, Andrés; Wang, HongBin et al. (2012) The RacGAP ?2-Chimaerin selectively mediates axonal pruning in the hippocampus. Cell 149:1594-606
Wang, Hongbin; Kazanietz, Marcelo G (2010) p23/Tmp21 differentially targets the Rac-GAP beta2-chimaerin and protein kinase C via their C1 domains. Mol Biol Cell 21:1398-408
Griner, Erin M; Caino, M Cecilia; Sosa, Maria Soledad et al. (2010) A novel cross-talk in diacylglycerol signaling: the Rac-GAP beta2-chimaerin is negatively regulated by protein kinase Cdelta-mediated phosphorylation. J Biol Chem 285:16931-41
Notcovich, Cintia; Diez, Federico; Tubio, Maria Rosario et al. (2010) Histamine acting on H1 receptor promotes inhibition of proliferation via PLC, RAC, and JNK-dependent pathways. Exp Cell Res 316:401-11

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