Scientific RepoRts | 7: 16543 | DOI:10.1038/s41598-017-15187-x
Reprogramming to pluripotency
does not require transition through
a primitive streak-like state
, Moritz Klingenstein
, Anna Möller
, Anett Illing
, Jelena Tosic
, Georg Kuales
, Leonhard Linta
, Thomas Seuerlein
, Sebastian J. Arnold
& Stefan Liebau
Pluripotency can be induced in vitro from adult somatic mammalian cells by enforced expression of
dened transcription factors regulating and initiating the pluripotency network. Despite the substantial
advances over the last decade to improve the eciency of direct reprogramming, exact mechanisms
underlying the conversion into the pluripotent stem cell state are still vaguely understood. Several
studies suggested that induced pluripotency follows reversed embryonic development. For somatic
cells of mesodermal and endodermal origin that would require the transition through a Primitive
streak-like state, which would necessarily require an Eomesodermin (Eomes) expressing intermediate.
We analyzed reprogramming in human and mouse cells of mesodermal as well as ectodermal origin
by thorough marker gene analyses in combination with genetic reporters, conditional loss of function
and stable fate-labeling for the broad primitive streak marker Eomes. We unambiguously demonstrate
that induced pluripotency is not dependent on a transient primitive streak-like stage and thus does not
represent reversal of mesendodermal development in vivo.
During mammalian development, early cell fate decisions during the process of gastrulation lead to the for-
mation of the three germ layers, namely ectoderm, mesoderm and endoderm. e development of mesoderm
and denitive endoderm (DE) is initiated by cell rearrangements of pluripotent, epithelial cells of the posterior
epiblast and a subsequent epithelial-to-mesenchymal transition (EMT) leading to the formation of the primitive
streak (PS). Mesoderm and DE cells are recruited as they migrate through the PS, while epiblast cells which do
not ingress through the streak give rise to neuroectodermal progeny including the epidermis and central nervous
system (reviewed in
). PS formation is initiated and maintained by a complex network of transcription factors
and signaling pathways. Signals include feed forward loops of WNT, TGFβ and BMP factors involving reciprocal
tissue interactions of epiblast and trophectoderm (reviewed in
). Absence or misexpression of these signals in the
epiblast leads to an impaired PS formation followed by disorganization or absence of the mesoderm and endo-
derm germ layers and embryonic lethality. Examples are null mutants for Nodal or Wnt signaling, such as Wnt3
the nuclear receptor Nr5a2
, or the T-box transcription factor Eomesodermin (Eomes)
e specication of dierent PS-derived cell types follows a strict spatio-temporal pattern. e most anterior
PS gives rise to the early transient population of mesendoderm cells that contribute to the DE and axial mes-
oderm. is population is followed by cells ingressing through the anterior third of the streak generating the
anterior mesoderm that gives rise to head and cardiogenic mesenchyme and extraembryonic mesoderm
ingressing at more posterior streak levels are giving rise to paraxial, intermediate and lateral plate mesoderm.
e signaling pathways regulating streak patterning include Nodal- and Wnt-activities. For example, high levels
Institute of Neuroanatomy & Developmental Biology (INDB), Eberhard Karls University Tuebingen, Oesterbergstr.
3, 72074, Tuebingen, Germany.
Department of Internal Medicine I, University Medical Center Ulm, Albert-Einstein-
Allee 23, 89081, Ulm, Germany.
Institute of Experimental and Clinical Pharmacology and Toxicology II, Faculty of
Medicine, University of Freiburg, Freiburg, Germany.
Spemann Graduate School of Biology and Medicine (SGBM),
Albert-Ludwigs-University Freiburg, Freiburg, Germany.
Faculty of Biology, Schänzlestraße 1, Albert-Ludwigs-
University Freiburg, Freiburg, Germany.
BIOSS Centre of Biological Signalling Studies, Albert-Ludwigs-University,
Freiburg, Germany. Stefanie Raab, Moritz Klingenstein and Anna Möller contributed equally to this work. Sebastian J.
Arnold, Alexander Kleger and Stefan Liebau jointly supervised this work. Correspondence and requests for materials
should be addressed to S.J.A. (email: firstname.lastname@example.org) or A.K. (email: alexander.
email@example.com) or S.L. (email: firstname.lastname@example.org)
Received: 28 June 2017
Accepted: 23 October 2017
Published: xx xx xxxx