The aim of this second competitive renewal proposal (years 11-15) is to pursue our program identifying the function of c-Src that, in the osteoclast, is required for bone resorption (src- mutants develop osteopetrosis) and is unique to c-Src (i.e., cannot be compensated by other Src family of kinases in this cell). During the first eleven years of this program, we have made very significant progress towards understanding the functions of c-Src in bone resorption. Although the mechanism(s) by which src deletion leads to a decrease in bone resorption is not yet fully characterized, it is now clear, in part through our work, that Src regulates integrin-dependent osteoclast adhesion and motility. Two central questions remain unanswered: 1/ What is the molecular basis for Src's """"""""uniqueness"""""""" in osteoclast function? and 2/ Why would the phosphorylation of Cbl be so important in the osteoclast? For the next funding period, we propose to 1) identify the molecular basis for Src's unique function in osteoclasts, 2) explore the mechanisms by which phosphorylation of Cbl regulates its functions, linking integrins to actin, podosome turnover and osteoclast function and 3) extend these in vitro studies to in vivo models by generating new transgenic mice that express relevant mutated versions of Src or Cbl.
The specific aims of this competitive renewal proposal are: 1) To identify the specific domains of Src that are responsible for the osteoclast phenotype (i.e. cannot be compensated by other members of the Src family of non-receptor tyrosine kinases); 2) To further characterize the mechanisms by which Src-mediated phosphorylation of Cbl regulates osteoclast function, i.e. the ability of Cbl to ubiquitinate and/or interact with other molecules [upstream components (Pyk2) and downstream components (dynamin and PI3-kinase)]; and 3) To translate the in vitro results obtained during the previous funding period and in Aims 1 and 2 into in vivo studies by creating new transgenic mice that express mutated Src or Cbl proteins that alter osteoclast function in vitro. The work proposed here will increase understanding of the regulation of osteoclast function and bone resorption, potentially identifying novel targets for therapeutic intervention to control bone loss related to osteoporosis and other diseases with increased bone resorption such as bone metastasis and periodontal and joint diseases.

Agency
National Institute of Health (NIH)
Institute
National Institute of Arthritis and Musculoskeletal and Skin Diseases (NIAMS)
Type
Research Project (R01)
Project #
5R01AR042927-15
Application #
7454990
Study Section
Skeletal Biology Structure and Regeneration Study Section (SBSR)
Program Officer
Sharrock, William J
Project Start
1994-07-15
Project End
2010-06-30
Budget Start
2008-07-01
Budget End
2009-06-30
Support Year
15
Fiscal Year
2008
Total Cost
$345,483
Indirect Cost
Name
Harvard University
Department
Dentistry
Type
Schools of Dentistry
DUNS #
047006379
City
Boston
State
MA
Country
United States
Zip Code
02115
Destaing, Olivier; Ferguson, Shawn M; Grichine, Alexei et al. (2013) Essential function of dynamin in the invasive properties and actin architecture of v-Src induced podosomes/invadosomes. PLoS One 8:e77956
Purev, Enkhtsetseg; Neff, Lynn; Horne, William C et al. (2009) c-Cbl and Cbl-b act redundantly to protect osteoclasts from apoptosis and to displace HDAC6 from beta-tubulin, stabilizing microtubules and podosomes. Mol Biol Cell 20:4021-30
Bruzzaniti, Angela; Neff, Lynn; Sandoval, Amanda et al. (2009) Dynamin reduces Pyk2 Y402 phosphorylation and SRC binding in osteoclasts. Mol Cell Biol 29:3644-56
Destaing, Olivier; Sanjay, Archana; Itzstein, Cecile et al. (2008) The tyrosine kinase activity of c-Src regulates actin dynamics and organization of podosomes in osteoclasts. Mol Biol Cell 19:394-404
Li, Lei; Hisamoto, Koji; Kim, Kyung Hee et al. (2007) Variant estrogen receptor-c-Src molecular interdependence and c-Src structural requirements for endothelial NO synthase activation. Proc Natl Acad Sci U S A 104:16468-73
Gil-Henn, Hava; Destaing, Olivier; Sims, Natalie A et al. (2007) Defective microtubule-dependent podosome organization in osteoclasts leads to increased bone density in Pyk2(-/-) mice. J Cell Biol 178:1053-64
Bruzzaniti, Angela; Baron, Roland (2006) Molecular regulation of osteoclast activity. Rev Endocr Metab Disord 7:123-39
Aoki, Kazuhiro; Saito, Hiroaki; Itzstein, Cecile et al. (2006) A TNF receptor loop peptide mimic blocks RANK ligand-induced signaling, bone resorption, and bone loss. J Clin Invest 116:1525-34
Sanjay, Archana; Miyazaki, Tsuyoshi; Itzstein, Cecile et al. (2006) Identification and functional characterization of an Src homology domain 3 domain-binding site on Cbl. FEBS J 273:5442-56
Bruzzaniti, Angela; Neff, Lynn; Sanjay, Archana et al. (2005) Dynamin forms a Src kinase-sensitive complex with Cbl and regulates podosomes and osteoclast activity. Mol Biol Cell 16:3301-13

Showing the most recent 10 out of 33 publications