We cloned a gene, BRMS1, which suppresses metastasis in six independently-derived human and murine cell lines without suppressing tumorigenicity. Our objective is to determine the biochemical mechanisms underlying BRMS1 metastasis suppression. Hypothesis 1 - Suppression of metastasis by BRMS1 requires restoration of gap junctional intercellular communication (GJIC). Transfection of BRMS1 restores GJIC along with altered transcription of connexins (Cx), i.e., Cx43 is up-regulated while Cx32 is down-regulated.
Aim 1 will address whether BRMS1 expression and GJIC are both required to suppress metastasis. BRMSl-transfected cells (metastasis-suppressed) will have either BRMS1 (Aim la) or Cx43 (Aim lb) expression selectively down-regulated using siRNA.
Aim lc will test whether decreased Cx32 in metastatic cells decreases metastatic potential while Aim ld will re-express Cx32 in BRMSl-transfected cells and test whether metastasis increases. All transfectants will be tested for GJIC in vitro and metastasis in vivo (Aim 1e). Hypothesis 2 - BRMSl suppresses metastasis via interactions with mSin3:histone deacetylase (HDAC). Using yeast two-hybrid, co-IP and chromatography, we showed that BRMS1 physically interacts with components of large complexes that include mSin3 and HDAC. We will define which BRMSI:HDAC:mSin3 complex(es) are responsible for metastasis suppression.
Aim 2 a will use FPLC, mass spectroscopy, co-IP and Y2H to define BRMS1 complexes and identify the BRMS1 interacting proteins.
Aim 2 b will map BRMS1 domains responsible for specific protein interactions.
Aim 2 c will test the ability of BRMS1 routto (1) restore GJIC, (2) regulate HDAC activity; and (3) suppress metastasis. Hypothesis 3- Brmsl expression will affect metastasis in autochthonous mammary tumors. We propose to develop Brmsl-null and Brine1 transgenic over-expressing mice and test whether endogenous expression alters tumor development and/or metastasis of autochthonous mammary carcinomas.
Aim 3 a will generate Brmsl null mice using conditional Cre-Lox recombination and test the hypothesis that the frequency of metastases from tg: MMTV-PyMT mammary tumors will increase when the mice are crossed.
Aim 3 b will generate mammary-specific and ubiquitous expression Brmsl transgenes that will result in high expression of Brmsl. tg:Brmsl mice will be crossed with tg: MMTV-PYMT and test the hypothesis that metastatic potential will decrease.

Agency
National Institute of Health (NIH)
Institute
National Cancer Institute (NCI)
Type
Research Project (R01)
Project #
5R01CA087728-09
Application #
7393346
Study Section
Tumor Progression and Metastasis Study Section (TPM)
Program Officer
Sathyamoorthy, Neeraja
Project Start
2000-07-01
Project End
2010-04-30
Budget Start
2008-05-01
Budget End
2010-04-30
Support Year
9
Fiscal Year
2008
Total Cost
$303,343
Indirect Cost
Name
University of Alabama Birmingham
Department
Pathology
Type
Schools of Medicine
DUNS #
063690705
City
Birmingham
State
AL
Country
United States
Zip Code
35294
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