Limits...
The RNA binding protein ESRP1 fine-tunes the expression of pluripotency-related factors in mouse embryonic stem cells.

Fagoonee S, Bearzi C, Di Cunto F, Clohessy JG, Rizzi R, Reschke M, Tolosano E, Provero P, Pandolfi PP, Silengo L, Altruda F - PLoS ONE (2013)

Bottom Line: Esrp1-depleted embryonic stem cells displayed impaired early differentiation in vitro and formed larger teratomas in vivo when compared to control embryonic stem cells.We also show that ESRP1 binds to Oct4 and Sox2 mRNAs and decreases their polysomal loading.ESRP1 thus acts as a physiological regulator of the finely-tuned balance between self-renewal and commitment to a restricted developmental fate.

View Article: PubMed Central - PubMed

Affiliation: Department of Molecular Biotechnology and Health Sciences, Molecular Biotechnology Center, University of Turin, Turin, Italy.

ABSTRACT
In pluripotent stem cells, there is increasing evidence for crosstalk between post-transcriptional and transcriptional networks, offering multifold steps at which pluripotency can be controlled. In addition to well-studied transcription factors, chromatin modifiers and miRNAs, RNA-binding proteins are emerging as fundamental players in pluripotency regulation. Here, we report a new role for the RNA-binding protein ESRP1 in the control of pluripotency. Knockdown of Esrp1 in mouse embryonic stem cells induces, other than the well-documented epithelial to mesenchymal-like state, also an increase in expression of the core transcription factors Oct4, Nanog and Sox2, thereby enhancing self-renewal of these cells. Esrp1-depleted embryonic stem cells displayed impaired early differentiation in vitro and formed larger teratomas in vivo when compared to control embryonic stem cells. We also show that ESRP1 binds to Oct4 and Sox2 mRNAs and decreases their polysomal loading. ESRP1 thus acts as a physiological regulator of the finely-tuned balance between self-renewal and commitment to a restricted developmental fate. Importantly, both mouse and human epithelial stem cells highly express ESRP1, pinpointing the importance of this RNA-binding protein in stem cell biology.

Show MeSH

Related in: MedlinePlus

ESRP1 is expressed in spermatogonial stem cells.A. Spermatogonial stem cells (SSCs) were characterised by qRT-PCR analysis. RQ is relative quantity (n = 4). B. Western blot analysis shows that ESRP1 is expressed in ES cells, in GPSCs as well as in SSCs. ES cells and GPSCs were lysed after 30 minutes’ preplating to remove Mefs, while SSCs were scraped off together with the Mef-feeder layer for lysis. The primordial germ cell marker, MVH was enriched in SSCs versus the other cell lines. Actin was used for normalisation (n = 2). 3T3 fibroblasts were used as negative control. C. Confocal microcroscopy analysis revealed that ESRP1 is both nuclear and cytoplasmic in the SSCs as well. MVH staining was used as positive control. Scale bar is 5 µm.
© Copyright Policy
Related In: Results  -  Collection


getmorefigures.php?uid=PMC3755004&req=5

pone-0072300-g005: ESRP1 is expressed in spermatogonial stem cells.A. Spermatogonial stem cells (SSCs) were characterised by qRT-PCR analysis. RQ is relative quantity (n = 4). B. Western blot analysis shows that ESRP1 is expressed in ES cells, in GPSCs as well as in SSCs. ES cells and GPSCs were lysed after 30 minutes’ preplating to remove Mefs, while SSCs were scraped off together with the Mef-feeder layer for lysis. The primordial germ cell marker, MVH was enriched in SSCs versus the other cell lines. Actin was used for normalisation (n = 2). 3T3 fibroblasts were used as negative control. C. Confocal microcroscopy analysis revealed that ESRP1 is both nuclear and cytoplasmic in the SSCs as well. MVH staining was used as positive control. Scale bar is 5 µm.

Mentions: In order to assess the applicability of these findings to other types of stem cells, we analysed the expression of ESRP1 in the unipotent spermatogonial stem cells or SSCs. As expected, the mouse SSCs express pluripotency markers like Oct4, Sox2, Klf4, Lin28, c-Myc as well as e-cadherin, but not Nanog compared to the pluripotent stem cells, ES and GPSCs [38], [39]. ESRP1 is expressed both at the RNA and protein levels in SSCs (Figure 5A and B). Interestingly, ESRP1 is present both in the nucleus and the cytoplasm of these cells, in accordance with the results obtained on ES cells (Figure 5C and Figure 2A, respectively). Moreover, ESRP1 is also highly expressed in CD133+ human kidney progenitor cells, but downregulated in kidney cancer stem cells (protein extracts kindly provided by B. Bussolati) (Figure S8) [26], [27].


The RNA binding protein ESRP1 fine-tunes the expression of pluripotency-related factors in mouse embryonic stem cells.

Fagoonee S, Bearzi C, Di Cunto F, Clohessy JG, Rizzi R, Reschke M, Tolosano E, Provero P, Pandolfi PP, Silengo L, Altruda F - PLoS ONE (2013)

ESRP1 is expressed in spermatogonial stem cells.A. Spermatogonial stem cells (SSCs) were characterised by qRT-PCR analysis. RQ is relative quantity (n = 4). B. Western blot analysis shows that ESRP1 is expressed in ES cells, in GPSCs as well as in SSCs. ES cells and GPSCs were lysed after 30 minutes’ preplating to remove Mefs, while SSCs were scraped off together with the Mef-feeder layer for lysis. The primordial germ cell marker, MVH was enriched in SSCs versus the other cell lines. Actin was used for normalisation (n = 2). 3T3 fibroblasts were used as negative control. C. Confocal microcroscopy analysis revealed that ESRP1 is both nuclear and cytoplasmic in the SSCs as well. MVH staining was used as positive control. Scale bar is 5 µm.
© Copyright Policy
Related In: Results  -  Collection

Show All Figures
getmorefigures.php?uid=PMC3755004&req=5

pone-0072300-g005: ESRP1 is expressed in spermatogonial stem cells.A. Spermatogonial stem cells (SSCs) were characterised by qRT-PCR analysis. RQ is relative quantity (n = 4). B. Western blot analysis shows that ESRP1 is expressed in ES cells, in GPSCs as well as in SSCs. ES cells and GPSCs were lysed after 30 minutes’ preplating to remove Mefs, while SSCs were scraped off together with the Mef-feeder layer for lysis. The primordial germ cell marker, MVH was enriched in SSCs versus the other cell lines. Actin was used for normalisation (n = 2). 3T3 fibroblasts were used as negative control. C. Confocal microcroscopy analysis revealed that ESRP1 is both nuclear and cytoplasmic in the SSCs as well. MVH staining was used as positive control. Scale bar is 5 µm.
Mentions: In order to assess the applicability of these findings to other types of stem cells, we analysed the expression of ESRP1 in the unipotent spermatogonial stem cells or SSCs. As expected, the mouse SSCs express pluripotency markers like Oct4, Sox2, Klf4, Lin28, c-Myc as well as e-cadherin, but not Nanog compared to the pluripotent stem cells, ES and GPSCs [38], [39]. ESRP1 is expressed both at the RNA and protein levels in SSCs (Figure 5A and B). Interestingly, ESRP1 is present both in the nucleus and the cytoplasm of these cells, in accordance with the results obtained on ES cells (Figure 5C and Figure 2A, respectively). Moreover, ESRP1 is also highly expressed in CD133+ human kidney progenitor cells, but downregulated in kidney cancer stem cells (protein extracts kindly provided by B. Bussolati) (Figure S8) [26], [27].

Bottom Line: Esrp1-depleted embryonic stem cells displayed impaired early differentiation in vitro and formed larger teratomas in vivo when compared to control embryonic stem cells.We also show that ESRP1 binds to Oct4 and Sox2 mRNAs and decreases their polysomal loading.ESRP1 thus acts as a physiological regulator of the finely-tuned balance between self-renewal and commitment to a restricted developmental fate.

View Article: PubMed Central - PubMed

Affiliation: Department of Molecular Biotechnology and Health Sciences, Molecular Biotechnology Center, University of Turin, Turin, Italy.

ABSTRACT
In pluripotent stem cells, there is increasing evidence for crosstalk between post-transcriptional and transcriptional networks, offering multifold steps at which pluripotency can be controlled. In addition to well-studied transcription factors, chromatin modifiers and miRNAs, RNA-binding proteins are emerging as fundamental players in pluripotency regulation. Here, we report a new role for the RNA-binding protein ESRP1 in the control of pluripotency. Knockdown of Esrp1 in mouse embryonic stem cells induces, other than the well-documented epithelial to mesenchymal-like state, also an increase in expression of the core transcription factors Oct4, Nanog and Sox2, thereby enhancing self-renewal of these cells. Esrp1-depleted embryonic stem cells displayed impaired early differentiation in vitro and formed larger teratomas in vivo when compared to control embryonic stem cells. We also show that ESRP1 binds to Oct4 and Sox2 mRNAs and decreases their polysomal loading. ESRP1 thus acts as a physiological regulator of the finely-tuned balance between self-renewal and commitment to a restricted developmental fate. Importantly, both mouse and human epithelial stem cells highly express ESRP1, pinpointing the importance of this RNA-binding protein in stem cell biology.

Show MeSH
Related in: MedlinePlus