Beyond the Identification of Transcribed Sequences:
Functional, Evolutionary and Expression Analysis
12th International Workshop
October 25-28, 2002
Washington, DC

List of Abstracts * Speakers * Organizers * Authors * Original Announcement

RNA-Binding Proteins as Reporters of Operational mRNA Networks in Tumors and Complex Tissues

Jack D. Keene, Luiz O. Penalva, Michael Burdick, Hedi Sutterluety and Patrick J. Lager
Department of Molecular Genetics and Microbiology and Center For RNA Biology, Duke University Medical Center, Durham, NC 27710 USA
Telephone: (919) 6845138
Fax: (919) 6848735

Tumors and tissues consist of multiple cell types that are interdependent and interact with one another using secreted factors and other modes of cell-cell communication.  For example, angiogenesis involves the secretion of factors by endothelial cells and by cancer cells that allow them to collaborate in tumor formation. Current methods of genomic analysis account for the steady state levels of messenger RNAs expressed in the whole tumor, but do not measure the expression of mRNAs within each cell type.  In order to understand how individual cells within a society of communicating tumor cells affect the gene expression of surrounding cells, we have devised a new approach to gene expression profiling.  By analyzing RNA-binding proteins within specific cell types that are associated endogenously with messenger ribonucleoproteins we can detect populations of mRNAs that are specific to each cell-type. In addition, we have engineered cells with variously epitope-tagged mRNA-binding proteins using specific promoters and virus-specific receptors, thus allowing us to profile the gene expression state of distinct cells within a mixture of cell types.  Experiments based upon mixed cell populations of tumor cells and endothelial cells will be described showing the recovery of cell type-specific mRNA populations.  In addition, using cDNA arrays we have demonstrated that certain endogenous mRNA-binding proteins can interact with unique subpopulations of mRNAs containing transcripts that encode functionally-related proteins.  These mRNA subsets may be analogous to the polycistronic mRNAs of prokaryotic operons because they can assemble monocistronic mRNAs and coordinate their expression at the posttranscriptional level (Keene and Tenenbaum, Mol Cell, 2002).  We have proposed that these mRNA subsets can function as operational genetic networks in that they contain both unique, as well as overlapping mRNA populations.  Most importantly, these subpopulations have the ability to reassort combinatorially, potentially giving rise to the production of complex phenotypes.  In order to elucidate operational mRNA networks within specific cell types of a tumor or complex tissue, we have immunoprecipitated endogenous mRNA-binding proteins that are tumor cell specific and are not present in the surrounding cells.  These methods should allow precise determination of the mRNA populations within operational networks that control the expression of proteins required for cell-cell communication during tumor formation and in response to anti-tumor therapies.

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List of Abstracts * Speakers * Organizers * Authors * Original Announcement

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