Evaluation of in vitro models for the study of human X chromosome inactivation

Author(s)

Ana Elisa Ribeiro Orsi

• Document type: Master's Dissertation
• Press: São Paulo.
• Institution: Universidade de São Paulo (USP). Instituto de Biociências (IBIOC/SB)
• Defense date: 2023-08-21

Examining board members

Lygia da Veiga Pereira Carramaschi; João Carlos Setubal

• Advisor: Lygia da Veiga Pereira Carramaschi

Abstract

X chromosome inactivation is a dose compensation mechanism that takes place in the cells of female placental mammals. Human XCI starts around the early blastocyst stage, and its understanding is hampered by the difficulties associated with the use of embryos in research. In order to address this issue, we evaluated different models for the study of XCI in vitro. Primed cells displayed monoallelic expression of the X-linked gene MECP2 and a single XIST cloud. Upon conversion to the naive state, biallelic MECP2 expression was observed, as well as absence of XIST clouds for the first two passages. Evaluation of EPSCs in vitro and in silico suggests that these cells have already undergone XCI. Similar results were observed for cells cultured in 4CL medium. After repriming, naive cells cultured in 5iLA displayed inactivation of the previously inactive MECP2 allele. On the other hand, blastoids generated from cells cultured in PXGL underwent progressive random XCI, although bias for the previously inactive allele was still observed. These results suggest that blastoids could be potentially employed for the study of human XCI in vitro. (AU)

• FAPESP's process: 20/16612-9 - Analysis of gene expression of human preimplantation embryos for the development of a strategy for maintenance of the morulas epigenetic stage
• Grantee: Ana Elisa Ribeiro Orsi
• Support Opportunities: Scholarships in Brazil - Master