In all mammalian species where cloning has been successful, at best a few percent of nuclear transfer embryos develop to term, and of those many die shortly after birth. This has raised questions of nuclear potency and differentiation that were posed half a century ago with the generation of the first amphibians by transfer of a somatic nucleus into the egg. These issues include whether the potency of a nucleus to direct the formation of an animal is lost with increasing age of the donor cell and whether its state of differentiation influences the type of abnormalities seen in the clone. The main problem of nuclear cloning is thought to be caused by faulty epigenetic reprogramming of the donor nucleus. This proposal seeks to understand molecular mechanisms that are responsible for the low survival rate and abnormal phenotypic characteristics of animals derived by nuclear cloning. We propose the following projects: 1. We will test the potency of nuclei derived from donor embryonic and somatic stem cells by nuclear transfer. 2. We will generate cDNA arrays that will be used to establish molecular criteria for nuclear reprogramming. 3. We will design strategies to improve the epigenetic reprogramming of somatic donor cells. Our goal is to convert the epigenetic state of the somatic cell to one that resembles that of an ES cell. 4. We will clone mice from mature neuronal donor cells. This will test whether alterations occur in the genome of neurons as part of normal brain physiology that would restrict nuclear potency. Embryonic and adult stem cells are thought to offer significant potential for regenerative cell therapy. However, major issues of applying this promising approach are unresolved. For example, the epigenetic state of embryonic and adult stem cell nuclei may be more amenable to reprogramming than that of terminally differentiated cells. It is a central focus of this proposal to establish functional and biological parameters that distinguish the epigenetic state of nuclei from stem cells and differentiated cells. This will serve as a molecular basis for altering the potential of somatic cells so they could be reprogrammed into different cell types that can be used in therapeutic approaches. Indeed, if the molecular mechanisms that are responsible for reprogramming could be understood, it might eventually be possible to manipulate a somatic cell and generate an ES cell-like cell without the need for oocytes and nuclear transfer.

Agency
National Institute of Health (NIH)
Institute
Eunice Kennedy Shriver National Institute of Child Health & Human Development (NICHD)
Type
Research Project (R01)
Project #
1R01HD045022-01
Application #
6686960
Study Section
Cell Development and Function Integrated Review Group (CDF)
Program Officer
Tasca, Richard J
Project Start
2003-07-28
Project End
2008-04-30
Budget Start
2003-07-28
Budget End
2004-04-30
Support Year
1
Fiscal Year
2003
Total Cost
$769,754
Indirect Cost
Name
Whitehead Institute for Biomedical Research
Department
Type
DUNS #
120989983
City
Cambridge
State
MA
Country
United States
Zip Code
02142
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