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Published on: September 30, 2010
Generation of cloned mice by direct nuclear transfer from natural killer T cells
Kimiko Inoue1, Hiroshi Wakao, Narumi Ogonuki
1RIKEN Bioresource Center, Tsukuba, Ibaraki 305-0074, Japan.
Abstract:
Cloning mammals by nuclear transfer (NT) remains inefficient. One fundamental question is whether clones have really been derived from differentiated cells rather than from rare stem cells present in donor-cell samples. To date, cells, such as mature lymphocytes, with genetic differentiation markers have been cloned to generate mice only via a two-step NT involving embryonic stem (ES) cell generation and tetraploid complementation [1, 2 and 3]. Here, we show that the genome of a unique T-cell population, natural killer T (NKT) cells, can be fully reprogrammed by a single-step NT. The pups and their placentas possessed the rearranged TCR loci specific for NKT cells. The NKT-cell-cloned embryos had a high developmental potential in vitro: Most (71%) developed to the morula/blastocyst stage, in marked contrast to embryos from peripheral blood T cells (12%; p < 1 x 10(-25)). Furthermore, ES cell lines were efficiently established from these NKT-cell blastocysts. These findings clearly indicate a high level of plasticity in the NKT-cell genome. Thus, differentiation of the genome is not always a barrier to NT cloning for either reproductive or therapeutic purposes, so we can now postulate that at least some mammals cloned to date have indeed been derived from differentiated donor cells.
Insights
Cloning mammals using nuclear transfer (NT) is inefficient. This study demonstrates that natural killer T (NKT) cells can be reprogrammed via single-step NT, indicating genome plasticity and potential for cloning differentiated cells.
Area of Science:
- Reproductive Biology
- Developmental Biology
- Immunology
Background:
- Mammalian cloning via nuclear transfer (NT) faces efficiency challenges.
- A key question is whether clones originate from differentiated cells or rare stem cells.
- Previous cloning of differentiated cells, like lymphocytes, required complex multi-step procedures.
Purpose of the Study:
- To investigate the reprogramming potential of natural killer T (NKT) cells via single-step NT.
- To determine if differentiated NKT cells can serve as effective donors for cloning.
- To assess the developmental capacity of embryos derived from NKT cell cloning.
Main Methods:
- Single-step nuclear transfer (NT) using natural killer T (NKT) cells as donor cells.
- In vitro culture of cloned embryos to assess developmental potential (morula/blastocyst stage).
- Establishment of embryonic stem (ES) cell lines from NKT-cell-derived blastocysts.
Main Results:
- Successful cloning of mice from NKT cells using a single-step NT procedure.
- Cloned pups and placentas retained NKT-cell-specific rearranged TCR loci.
- NKT-cell-derived embryos showed high in vitro developmental potential (71% to morula/blastocyst), unlike peripheral T-cell-derived embryos (12%).
- Efficient derivation of ES cell lines from NKT-cell blastocysts was achieved.
Conclusions:
- The genome of NKT cells exhibits significant plasticity, enabling full reprogramming by single-step NT.
- Differentiation is not an absolute barrier to NT cloning, suggesting cloned mammals may originate from differentiated cells.
- These findings advance possibilities for both reproductive and therapeutic cloning applications.
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