Depletion of Mageb16 induces differentiation of pluripotent stem cells predominantly into mesodermal derivatives

John Antonydas Gaspar,Poornima Sureshkumar,Symeon Papadopoulos,Sureshkumar Perumal Srinivasan,Jürgen Hescheler,Michael Xavier Doss,Agapios Sachinidis

doi:10.1038/s41598-017-14561-z

John Antonydas Gaspar, Poornima Sureshkumar + Show 5 more

Open Access

https://doi.org/10.1038/s41598-017-14561-z

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Journal: Scientific Reports	Publication Date: Oct 27, 2017
Citations: 7	License type: open-access

Affiliation: University of Cologne

Abstract

The Melanoma-associated Antigen gene family (MAGE) generally encodes for tumour antigens. We had identified that one of the MAGE gene members, Mageb16 was highly expressed in undifferentiated murine embryonic stem cells (ESCs). While the role of Mageb16 in stemness and differentiation of pluripotent stem cells is completely unknown, here, in our current study, we have demonstrated that Mageb16 (41 kDa) is distributed in cytosol and/or in surface membrane in undifferentiated ESCs. A transcriptome study performed at differentiated short hairpin RNA (shRNA)-mediated Mageb16 knockdown (KD) ESCs and scrambled control (SCR) ESCs until a period of 22 days, revealed that Mageb16 KD ESCs mainly differentiated towards cells expressing mesodermal and cardiovascular lineage - gene markers. Gene markers of other mesoderm-oriented biological processes such as adipogenesis, osteogenesis, limb morphogenesis and spermatogenesis were also significantly enriched in the differentiated Mageb16 KD ESCs. The expression levels of contractile genes were higher in differentiated Mageb16 KD ESCs when compared to differentiated SCR and wild ESCs, suggesting a higher cardiomyogenic potential of Mageb16 depleted ESCs. Further analysis indicates that regulative epigenetic networks and nucleocytoplasmic modifications induced by the depletion of Mageb16, may play a probable role in differentiation.

Highlights

Melanoma-associated Antigen family (MAGE) of genes are located in X chromosome and encodes for tumour antigens which normally bind to T-lymphocytes in cancer patients[1]
Authors have reported very high expression levels of Mageb[16] in testis of 3 to 18 day postnatal mice embryos and though slightly decreased, the expression level was still high in testis of 4 month old adult mice[6]
Maximal expression of Mageb[16] has been observed in undifferentiated SCR embryonic stem cells (ESCs) and the expression started declining at 4-day old embryoid bodies (EBs) (Fig. 1A) (Microarray data)

Summary

Introduction

Melanoma-associated Antigen family (MAGE) of genes are located in X chromosome and encodes for tumour antigens which normally bind to T-lymphocytes in cancer patients[1]. SiRNA knockdown of Mageb[16] in undifferentiated ESCs cultured under monolayer conditions resulted in a remarkable upregulation of mesodermal, ectodermal and endodermal marker genes after 48 h of differentiation as compared to untreated and control scrambled (SCR)-oligonucleotide treated ESCs7. These results suggested a crucial key role of Mageb[16] for maintenance of pluripotency and differentiation of ESCs. In the present study, this hypothesis has been evaluated by generating a transgenic mESC line in which Mageb[16] was permanently silenced by knockdown (KD) of Mageb[16] using a shRNA directed to Mageb[16] mRNA. The cellular localization of Mageb[16] has been determined in ESCs

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Conclusion