Tremble et al

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Key publications:

Tremble KC, Stirparo GG, Bates LE, Maskalenka K, Stuart HT, Jones HT, Andersson-Rolf A, Radzisheuskaya A, Koo BK, Bertone P, Silva JCR. Sox2 modulation increases naïve pluripotency plasticity. iScience (2021). 24(3):102153. doi: 10.1016/j.isci.2021.102153.

In this study we found that perturbation of Sox2 expression elicits a naïve pluripotent stem cell with the ability to both acquire an embryonic or extraembryonic lineage fate.

Stuart HT,  Stirparo G, Lohoff T, Bates L, Kinoshita M, Lim CY, Sousa E, Maskalenka K, Radzisheuskaya A, Malcolm A, Alves MRP, Lloyd R, Nestorowa S, Humpreys P, Mansfield W, Reik W, Bertone P, Nichols J, Göttgens B, Silva JCR. Distinct molecular trajectories converge to induce naive pluripotency. Cell Stem Cell (2019). 25(3):388-406. doi: 10.1016/j.stem.2019.07.009.

Here we report distinct routes of reprogramming to naïve pluripotency. These differ in their transcriptional trajectories, mechanistic requirements and developmental parallels, thus demonstrating considerable flexibility for a given cell identity transition to occur. However, distinct routes converge on precise Oct4 expression, which is necessary and sufficient for naïve pluripotency induction.


Sousa EJ, Stuart HT, Bates LE, Ghorbani M, Nichols J, Dietmann S, Silva JCR. Exit from Naive Pluripotency Induces a Transient X Chromosome Inactivation-like State in Males. Cell Stem Cell. (2018) 22(6):919-928. doi:10.1016/j.stem.2018.05.001.

Here we report that the initiation of X chromosome inactivation takes place in males and on both X chromosomes in females. This is transient and rapid and is triggered by downregulation of naive pluripotent transcription factors during the onset of differentiation.

****highlighted in Science:

Santos R, Tosti L, Radzisheuskaya A, Caballero I, Kaji K, Hendrich B, Silva JCR. Mbd3/NuRD facilitates transcription factor induced pluripotency in a context dependent manner. Cell Stem Cell. (2014) 15(1):102-10. doi:10.1016/j.stem.2014.04.019.

Here we show that Mbd3/NuRD plays a positive role in reprogramming in certain contexts and that overexpression of Mbd3 facilitates Nanog-mediated reprogramming.


Stuart HT, van Oosten AL, Radzisheuskaya A, Martello G, Miller A, Dietmann S, Nichols J, Silva JCR. NANOG amplifies STAT3 activation and they synergistically induce the naïve pluripotent program. Current Biology. (2014) 24(3):340-6. doi: 10.1016/j.cub.2013.12.040.

In this study, we connect NANOG with STAT3 signalling, two key mechanisms driving naïve pluripotency acquisition. Our finding that NANOG modulates signal transduction of extracellular cues adds a new dimension to the interplay between external environment and nuclear control networks to instate and reinforce cellular identity.


Radzisheuskaya A, Chia GLB, Santos R, Theunissen TW, Castro LFC, Nichols J, Silva JCR. Defined Oct4 level governs cell state transitions of pluripotency entry and differentiation into all embryonic lineages. Nature Cell Biology. (2013) 15(6):579-90. doi:10.1038/ncb2742.

We significantly expanded and re-defined our previous understanding of the biological role of Oct4 from one important for pluripotency to one also actively controlling cell state transitions taking place during the entry to and exit from the naive pluripotent cell state.


Costa Y, Ding J, Theunissen TW, Faiola F, Hore TA, Shliaha PV, Fidalgo M, Saunders A, Lawrence M, Dietmann S, Das S, Levasseur DN, Li Z, Xu M, Reik W, Wang J#, Silva JCR#. Nanog-dependent function of Tet1 and Tet2 in establishment of pluripotency. Nature. (2013) 495(7441):370-4. doi:10.1038/nature11925.​​

This work describes for the first time a direct molecular mechanism by which Nanog works in the induction of naive pluripotency. This occurs via the epigenetic regulators Tet1 and Tet2 that we now also associate with a transcription factor.

Radzi sheuskaya A, Pas que V, Gillich A, Halley-Stott RP, Panamarova M, Zernicka-Goetz M, Surani MA, Silva JCR. Histone variant macroH2A marks embryonic differentiation in vivo and acts as an epigenetic barrier to induced pluripotency. Journal of Cell Science. (2012) 125(Pt 24):6094-104. doi: 10.1242/jcs.113019

Our study identified for the first time a link between an epigenetic mark and cell fate restriction during somatic cell differentiation, which helps to maintain cell identity and antagonises induction of a pluripotent stem cell state.

 van Oosten AL , Costa Y, Smith A, Si lva JCR. Jak/Stat3 signalling is sufficient and dominant over antagonistic cues for the establishment of naïve pluripotency. Nature Communications. (2012) 3:817. doi: 10.1038/ncomms1822.

This study attributes key properties to JAK/STAT3 that position it as one of the most potent reprogramming factors for the induction of naïve pluripotency. Increased JAK/STAT3 activation was found to be sufficient and dominant over antagonistic cues to enable the induction of a naïve pluripotent state.

  Theunissen TW  ,  Costa Y ,  Radzisheusk aya A , van Oosten AL, Lav ial F, Pain B, Castro, LFC, Silva JCR. Reprogramming capacity of Nanog is functionally conserved in vertebrates and resides in a unique homeodomain. Development. (2011) 138(22):4853-65. doi:10.1242/dev.068775.

We demonstrated that the reprogramming capacity of Nanog is functionally conserved in vertebrates and resides in its unique homeodomain. This is of particular significance when taking into account that Nanog is the only reprogramming factor showing poor protein sequence conservation, implying that a naive pluripotent cell state is a feature of all vertebrates.

 Theunissen TW , van Oosten AL, Castelo-Bra nco G, Hall J, Smith A, Silva JCR. Nanog overcomes reprogramming barriers and induces pluripotency in minimal conditions. Current Biology. (2011) 21(1):65-71. doi:10.1016/j.cub.2010.11.074.

This paper revealed that Nanog has the capacity to overcome multiple barriers to reprogramming and also to induce pluripotency in minimal culture conditions. The latter characteristic provides a strategy for imposing naive pluripotency in mammalian cells independently of species-specific culture requirements.

  Silva J#  , Nic hols J,  Theunissen TW , Guo G,   van Oosten AL , Barrandon O, Wray J, Cham bers I, Yamanak a S, Smith A#. Nanog is the Gateway to the Pluripotent Ground State. Cell. (2009) 138(4):722-37.

This paper demonstrates that Nanog is required for the establishment of naive pluripotency both in vitro and in vivo, which draws a parallel between in vitro-induced pluripotency and embryonic development.

 Silva J# , Barrandon O, Nichol s J, Theunissen TW, Kawaguchi J, Smith A#. Promotion of Reprogramming to Ground State Pluripotency by Signal Inhibition. PLoS Biology. (2008) 6(10):e253. doi:10.1371/journal.pbio.0060253.

We identified reprogramming intermediate cells, which were thought to be by-products of induced pluripotency but are instead cells trapped in a proliferative cell state. Upon medium switch to one containing inhibitors of the signalling pathways of Mek/Erk and Gsk3b combined with LIF, they undergo relatively fast and efficient reprogramming to a naive pluripotent cell state.

Silva J, Chambers I, Pollard S, Smith A. Nanog promotes transfer of pluripotency after cell fusion. Nature. (2006) 441(7096):997-1001. Epub 2006 Jun 14.

This paper precedes Yamanaka’s work and reports the first ever identified gene with nuclear reprogramming activity in the conversion of a somatic epigenome back into a pluripotent epigenome.


# Joint corresponding/senior authors. Silva lab members are indicated in italicised bold.

Other publications:


Stirparo GG, Kurowski A, Yanagida A, Bates LE, Strawbridge SE, Hladkou S, Stuart HT, Boroviak TE, Silva JCR, Nichols J. OCT4 induces embryonic pluripotency via STAT3 signaling and metabolic mechanisms. Proc Natl Acad Sci U S A. (2021) 118(3):e2008890118. doi: 10.1073/pnas.2008890118.

Lawrence M, Theunissen T, Lombard P, Adams D, Silva JCR. ZMYM2 inhibits NANOG mediated reprogramming. Welcome Open Research (2019). Jul 4:88. doi: 10.12688/wellcomeopenres.15250.1.

Peñalosa-Ruiz G, Bousgouni V, Gerlach JP, Waarlo S, van de Ven JV, Veenstra TE, Silva JCR, van Heeringen SJ, Bakal C, Mulder KW, Veenstra GJC. WDR5, BRCA1, and BARD1 Co-regulate the DNA Damage Response and Modulate the Mesenchymal-to-Epithelial Transition during Early Reprogramming. Stem Cell Reports. (2019) 12(4):743-756. doi: 10.1016/j.stemcr.2019.02.006.

Kleine-Brüggeney H, van Vliet LD, Mulas C, Gielen F, Agley CC, Silva JCR, Smith A, Chalut K, Hollfelder F. Long-Term Perfusion Culture of Monoclonal Embryonic Stem Cells in 3D Hydrogel Beads for Continuous Optical Analysis of Differentiation. Small. (2018) 15(5):e1804576.. doi: 10.1002/smll.201804576.

Bates LE, Silva JC. Reprogramming human cells to naïve pluripotency: how close are we? Current Opinion in Genetics & Development. (2017) 46:58-65. PMID: 28668635

Andersson-Rolf A, Mustata RC, Merenda A, Kim J, Perera S, Grego T, Andrews K, Tremble K, Silva JC, Fink J, Skarnes WC, Koo BK. One-step generation of conditional and reversible gene knockouts. Nat Methods. (2017) 14(3):287-289. doi: 10.1038/nmeth.4156.

Silva JC, Pera RA. Editorial overview: cell reprogramming, regeneration and repair. Current Opinion in Genetics & Development. (2014) 28:v-vi. doi:10.1016/j.gde.2014.11.003.

Schwarz BA, Bar-Nur O, Silva JCR, Hochedlinger K. Nanog is dispensable for the generation of induced pluripotent stem cells. Current Biology. (2014) 24(3):347-50. doi: 10.1016/j.cub.2013.12.050.


Christophorou MA, Castelo-Branco G, Halley-Stott R, Oliveira CS, Loos R, Radzisheuskaya A, Mowen KA, Bertone P, Silva JCR, Zernicka-Goetz M, Nielsen ML, Gurdon J, Kouzarides T. Citrullination regulates pluripotency and histone H1 binding to chromatin. Nature. (2014) 507(7490):104-8. doi:10.1038/nature12942.


Radzisheuskaya A and Silva JC. Do all roads lead to Oct4? The emerging concepts of induced pluripotency. Trends Cell Biol. (2013) 24(5):275-84. doi: 10.1016/j.tcb.2013.11.010.


Silva J. Deciphering Reprogramming. Cell Stem Cell. (2012) 11(6):742.

Fidalgo M, Faiola F, Pereira CF, Ding J, Saunders A, Gingold J, Schaniel C, Lemischka IR, Silva JCR, Wang J. Zfp281 Mediates Nanog Autorepression through Recruitment of the NuRD Complex and Inhibits Somatic Cell Reprogramming. Proc Natl Acad Sci U S A. (2012) 109(40):16202-7. doi: 10.1073/pnas.1208533109. Epub 2012 Sep 17.

 Theunissen TW  and Silva JCR. Somatic cel l reprogramming: role of homeodomain protein Nanog. Stem Cells and Cancer Stem Cells, 2012 Volume 6, DOI 10.1007/978-94-007-2993-3_33. Edited by M.A. Hayat. Springer Press.

 Theunissen TW , Silva JCR. Switching o n pluripotency: a perspective on the biological requirement of Nanog. Philos Trans R Soc Lond B Biol Sci. (2011) 366(1575):2222-9. doi:10.1098/rstb.2011.0003.

Yang J,   van Oosten AL  ,  Theunissen TW , Guo G, Silva JCR, S  mith A. Stat3 activation is limiting for reprogramming to ground state pluripotency. Cell Stem Cell. (2010) 7(3):319-28. doi:10.1016/j.stem.2010.06.022.

Nichols J, Silva J, Roode M, Smith A. Suppression of Erk signaling promotes ground state pluripotency in the mouse embryo. Development. (2009) 136(19):3215-22. doi:10.1016/j.cell.2009.07.039.

Banito A, Rashid ST, Acosta JC, Li S, Pereira CF, Geti I, Pinho S, Silva JC, Azuara V, Walsh M, Vallier L, Gil J. Senescence impairs successful reprogramming to pluripotent stem cells. Genes Dev. (2009) 23(18):2134-9. doi:10.1101/gad.1811609

Buehr M, Meek S, Blair K, Yang J, Ure J, Silva J, McLay R, Hall J, Ying Q, Smith A. Capture of Authentic Embryonic Stem Cells from Rat Blastocysts. Cell. (2008) 135(7):1287-98. doi:10.1016/j.cell.2008.12.007.

Silva J, Smith A. Capturing pluripotency. Cell. (2008) 132(4):532-6. doi:10.1016/j.cell.2008.02.006.

Chambers I, Silva J, Colby D, Nichols J, Robertson M, Nijmeijer B, Vrana J, Grotewold L, Smith A. Nanog safeguards pluripotency and mediates germline development. Nature. (2007) 450(7173): 1230-4.

de Napoles M, Mermoud JE, Wakao R, Tang YA, Endoh M, Appanah R, Nesterova TB, Silva J, Otte AP, Vidal M, Koseki H, Brockdorff N. Polycomb group proteins Ring1A/B link ubiquitylation of histone H2A to heritable gene silencing and X inactivation. Developmental Cell. (2004) 7(5):663-76.

Silva J, Mak W, Zvetkova I, Appanah R, Nesterova TB, Webster Z, Peters AH, Jenuwein T, Otte AP, Brockdorff N. Establishment of histone H3 methylation on the inactive X chromosome requires transient recruitment of Eed-Enx1 polycomb group complexes. Developmental Cell. (2003) 4(4):481-95.

Mak W, Baxter J, Silva J, Newall AE, Otte AP, Brockdorff N. Mitotically stable association of polycomb group proteins Eed and Enx1 with the inactive X chromosome in trophoblast stem cells. Current Biology. (2002) 12(12):1016-20.

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Sox2 modulation increases naïve pluripotency plasticity.