Using Xenopus to Understand
23 Pluripotency and to Reprogram
Cells for Therapeutic Use
Meghana S. Oak and Eva Hörmanseder
CONTENTS
23.1. Introduction .............................................................................................................................................................. 325
23.2. Inception of Nuclear Reprogramming ..................................................................................................................... 325
23.2.1. Early SCNT Experiments Establish the Reprogramming Field ............................................................... 326
23.2.2. Nuclear Reprogramming Using Xenopus laevis Oocytes, Eggs, and Egg Extracts .................................. 327
23.3. Nuclear Reprogramming Takes a Leap from Frog to Mammals and Clinical Application ..................................... 327
23.3.1. Mammalian Nuclear Transfer and Generation of Pluripotent Stem Cell Lines ........................................ 327
23.3.2. Use of Alternative Methods for Nuclear Reprogramming ........................................................................ 328
23.4. Molecular Insights into the Process of Nuclear Reprogramming Gained in Xenopus laevis .................................. 329
23.4.1. Reprogramming and DNA Replication ..................................................................................................... 329
23.4.2. Change of Gene Expression Patterns during Reprogramming ................................................................. 330
23.4.3. Oocyte Factors Promote Nuclear Reprogramming ................................................................................... 330
23.4.4. DNA Methylation as a Barrier to Nuclear Reprogramming ..................................................................... 332
23.4.5. Histone Modifcations and Resistance to Nuclear Reprogramming ......................................................... 332
23.5. Conclusion and Future Perspectives ........................................................................................................................ 332
References ............................................................................................................................................................................ 333
23.1. INTRODUCTION
Every multicellular organism possesses a variety of cells
of different types. Differentiated cells are extremely
stable and, once defned, do not ordinarily change their
fate. We now know that differentiation of cells takes place
through distinct transcription patterns and is maintained
with the help of epigenetic modifcations and chromatin
structure, even though the exact mechanistic details are
still not fully understood. It has been shown using different techniques that the nucleus of a differentiated cell
can be reverted to a state mimicking that of an undifferentiated, totipotent cell, which can itself develop into
an entire organism, a process commonly referred to as
“nuclear reprogramming.”
One method that achieves reprogramming of nuclei is a
technique called somatic cell nuclear transfer (SCNT), in
which a somatic nucleus is transplanted into the enucleated
egg of an organism. The early attempts at SCNT were performed on amphibians, and much of the seminal work that
proved for the frst time that nuclear reprogramming to totipotency is possible in differentiated somatic nuclei was done
on Xenopus laevis. The use of Xenopus in developmental
biology, although fortuitous, is tremendously advantageous.
The early accomplishments seen using this model organism
eventually led to successful cloning in mammalian systems
and new reprogramming techniques being developed using
in vitro models. The clinical applications of nuclear reprogramming and cellular plasticity are many, including therapeutical cloning, cell replacement therapies, and modeling
of human diseases in vitro. The establishment of nuclear
reprogramming in more advanced model systems has
allowed this technology to make its way into clinical trials. The relevance of nuclear reprogramming studies using
Xenopus as a model system was recognized in 2012, when
the Nobel Prize in Physiology or Medicine was awarded to
Sir John B. Gurdon, together with Shinya Yamanaka, for
their discovery that mature cells can be reprogrammed to
become pluripotent.
23.2. INCEPTION OF NUCLEAR
REPROGRAMMING
Do all cell types house the same set of genes?
The totipotent zygote divides and differentiates into a number of different cell types to give rise to a live, fully developed organism. Differentiated cells rarely, if ever, reverse
or change their state of terminal differentiation. This begs
the question—do all cell types house the same set of genes?
The question of how cell differentiation is established and
maintained paved the way to the cell fate reprogramming
DOI: 10.1201/9781003050230-26
325
23 Pluripotency and to Reprogram
Cells for Therapeutic Use
Meghana S. Oak and Eva Hörmanseder
CONTENTS
23.1. Introduction .............................................................................................................................................................. 325
23.2. Inception of Nuclear Reprogramming ..................................................................................................................... 325
23.2.1. Early SCNT Experiments Establish the Reprogramming Field ............................................................... 326
23.2.2. Nuclear Reprogramming Using Xenopus laevis Oocytes, Eggs, and Egg Extracts .................................. 327
23.3. Nuclear Reprogramming Takes a Leap from Frog to Mammals and Clinical Application ..................................... 327
23.3.1. Mammalian Nuclear Transfer and Generation of Pluripotent Stem Cell Lines ........................................ 327
23.3.2. Use of Alternative Methods for Nuclear Reprogramming ........................................................................ 328
23.4. Molecular Insights into the Process of Nuclear Reprogramming Gained in Xenopus laevis .................................. 329
23.4.1. Reprogramming and DNA Replication ..................................................................................................... 329
23.4.2. Change of Gene Expression Patterns during Reprogramming ................................................................. 330
23.4.3. Oocyte Factors Promote Nuclear Reprogramming ................................................................................... 330
23.4.4. DNA Methylation as a Barrier to Nuclear Reprogramming ..................................................................... 332
23.4.5. Histone Modifcations and Resistance to Nuclear Reprogramming ......................................................... 332
23.5. Conclusion and Future Perspectives ........................................................................................................................ 332
References ............................................................................................................................................................................ 333
23.1. INTRODUCTION
Every multicellular organism possesses a variety of cells
of different types. Differentiated cells are extremely
stable and, once defned, do not ordinarily change their
fate. We now know that differentiation of cells takes place
through distinct transcription patterns and is maintained
with the help of epigenetic modifcations and chromatin
structure, even though the exact mechanistic details are
still not fully understood. It has been shown using different techniques that the nucleus of a differentiated cell
can be reverted to a state mimicking that of an undifferentiated, totipotent cell, which can itself develop into
an entire organism, a process commonly referred to as
“nuclear reprogramming.”
One method that achieves reprogramming of nuclei is a
technique called somatic cell nuclear transfer (SCNT), in
which a somatic nucleus is transplanted into the enucleated
egg of an organism. The early attempts at SCNT were performed on amphibians, and much of the seminal work that
proved for the frst time that nuclear reprogramming to totipotency is possible in differentiated somatic nuclei was done
on Xenopus laevis. The use of Xenopus in developmental
biology, although fortuitous, is tremendously advantageous.
The early accomplishments seen using this model organism
eventually led to successful cloning in mammalian systems
and new reprogramming techniques being developed using
in vitro models. The clinical applications of nuclear reprogramming and cellular plasticity are many, including therapeutical cloning, cell replacement therapies, and modeling
of human diseases in vitro. The establishment of nuclear
reprogramming in more advanced model systems has
allowed this technology to make its way into clinical trials. The relevance of nuclear reprogramming studies using
Xenopus as a model system was recognized in 2012, when
the Nobel Prize in Physiology or Medicine was awarded to
Sir John B. Gurdon, together with Shinya Yamanaka, for
their discovery that mature cells can be reprogrammed to
become pluripotent.
23.2. INCEPTION OF NUCLEAR
REPROGRAMMING
Do all cell types house the same set of genes?
The totipotent zygote divides and differentiates into a number of different cell types to give rise to a live, fully developed organism. Differentiated cells rarely, if ever, reverse
or change their state of terminal differentiation. This begs
the question—do all cell types house the same set of genes?
The question of how cell differentiation is established and
maintained paved the way to the cell fate reprogramming
DOI: 10.1201/9781003050230-26
325