Phosphoinositide 3-kinases (PI 3-kinases) are critical regulators of proliferation, motility, and apoptosis. The Class IA PI 3-kinase is a dimer containing distinct regulatory (p85) and catalytic (p110) subunits, both of which are frequently mutated in human cancer. While a large amount of energy in academia and industry has been applied to the study of these enzymes, there are critical aspects of PI 3-kinase regulation that remain poorly understood. For example, although the p110 catalytic subunit has been crystallized, there is little information as to how the distinct domains of p85 are oriented in space, or how mutants of p85 lead to a loss of PI 3-kinase regulation. A specific role of p110 catalytic subunit in the proliferation and tumorigenesis of PTEN null prostate cancer cells has been identified, but the mechanisms that regulate p110 in these cells are not clear. Although novel mutations of p85 have been identified in colon cancer and glioblastoma, the effects of these mutations on tumorigenesis and metastasis in vivo are not known. The current proposal uses both biochemical and genetic approaches to address fundamental questions concerning PI 3-kinase signaling in cancer.
Aim 1 uses single molecule FRET measurements to produce a spatial map of the p85 regulatory subunit, and to determine how the binding of PI 3-kinase activators alters p85 conformation and interactions with the p110 catalytic subunit.
Aim 2 examines the mechanism of p110 activation by G subunits. Sites of p110-G interactions will be defined using genetic and biochemical methods. These sites will then be mutated, in order to test the requirement for G binding in signaling by p110 in PTEN null prostate cancer cells.
Aim 3 will study the signaling by p85 mutants in colon cancer and glioblastoma cell lines, the tumor types in which the recently described mutants were identified. Lentivirus methods will be used to knock down endogenous p85 and rescue with wild type or mutant p85. The effects of these mutants on in vitro cell growth and motility, and in vivo tumorigenesis and metastasis, will be studied. This proposal will define novel aspects of PI 3-kinase signaling and regulation, and will provide a better understanding of structural features that could be exploited as drug targets.

Public Health Relevance

PI 3-kinases are intracellular enzymes that play an important role in cell proliferation and survival, and which frequently contain activating mutations in human cancers. Understanding the mechanisms that regulate their activity in normal and malignant cells is critical for the design of novel and specific drugs to target these enzymes in human disease.

Agency
National Institute of Health (NIH)
Institute
National Institute of General Medical Sciences (NIGMS)
Type
Research Project (R01)
Project #
5R01GM055692-16
Application #
8477199
Study Section
Tumor Cell Biology Study Section (TCB)
Program Officer
Gerratana, Barbara
Project Start
1997-05-01
Project End
2014-05-31
Budget Start
2013-06-01
Budget End
2014-05-31
Support Year
16
Fiscal Year
2013
Total Cost
$333,828
Indirect Cost
$132,727
Name
Albert Einstein College of Medicine
Department
Pharmacology
Type
Schools of Medicine
DUNS #
110521739
City
Bronx
State
NY
Country
United States
Zip Code
10461
Dbouk, Hashem A; Backer, Jonathan M (2013) Novel approaches to inhibitor design for the p110β phosphoinositide 3-kinase. Trends Pharmacol Sci 34:149-53
Dou, Zhixun; Pan, Ji-An; Dbouk, Hashem A et al. (2013) Class IA PI3K p110β subunit promotes autophagy through Rab5 small GTPase in response to growth factor limitation. Mol Cell 50:29-42
Vadas, Oscar; Dbouk, Hashem A; Shymanets, Aliaksei et al. (2013) Molecular determinants of PI3Kγ-mediated activation downstream of G-protein-coupled receptors (GPCRs). Proc Natl Acad Sci U S A 110:18862-7
Salamon, Rachel Schnur; Backer, Jonathan M (2013) Phosphatidylinositol-3,4,5-trisphosphate: tool of choice for class I PI 3-kinases. Bioessays 35:602-11
Dbouk, Hashem A; Khalil, Bassem D; Wu, Haiyan et al. (2013) Characterization of a tumor-associated activating mutation of the p110* PI 3-kinase. PLoS One 8:e63833
Dbouk, Hashem A; Vadas, Oscar; Shymanets, Aliaksei et al. (2012) G protein-coupled receptor-mediated activation of p110β by Gβγ is required for cellular transformation and invasiveness. Sci Signal 5:ra89
Yoon, Mee-Sup; Du, Guangwei; Backer, Jonathan M et al. (2011) Class III PI-3-kinase activates phospholipase D in an amino acid-sensing mTORC1 pathway. J Cell Biol 195:435-47
Backer, Jonathan M (2010) The regulation of class IA PI 3-kinases by inter-subunit interactions. Curr Top Microbiol Immunol 346:87-114
Ciraolo, Elisa; Morello, Fulvio; Hobbs, Robin M et al. (2010) Essential role of the p110beta subunit of phosphoinositide 3-OH kinase in male fertility. Mol Biol Cell 21:704-11
Dbouk, Hashem A; Backer, Jonathan M (2010) A beta version of life: p110β takes center stage. Oncotarget 1:729-33

Showing the most recent 10 out of 19 publications