The effects of H3.3 mutation on ES stem cell self renewal, pluripotency and differentiation

• Main Author: Sim, Wei Xiong
• Other Authors: Lim Chin Yan
• Format: Final Year Project
• Language: English
• Published: 2017
• Subjects: DRNTU::Science
• Online Access: http://hdl.handle.net/10356/72050
• Institution: Nanyang Technological University

Recent studies have shed light on mutations of H3.3 N-terminal tail and the role it plays in neoplastic diseases such as paediatric diffuse intrinsic pontine glioma (DIPG) and chondroblastomas. However, little is known on how these H3.3 single amino acid substitutions can affect embryonic stem cell characteristics. In this study, we show that H3.3 Glycine position X may be necessary for ES cell survival, self-renewal, and pluripotency. Multiple ES cell lines overexpressing different H3.3 proteins were generated to study effects of mutant H3.3 expression. From the two attempts to generate the ES cell lines; we observed a poor ES colony survival in GXR construct transfected ES cells. Furthermore, most surviving H3.3 GXR expressing cell colonies appear unhealthy and differentiated. The transient expression of mutant H3.3 in ES cultures confirmed that the expression of mutant H3.3 GXR compromises ES cell survival, self-renewal and pluripotency, showing a significant reduction in number of surviving cells only in ES cells expressing GXR H3.3. Furthermore the lineage marker analysis of the transfected ES cells revealed a slight drop in the expression of pluripotent genes and a significant increase in the expression of differentiation markers of ES cells transiently expressing GXR H3.3. All in all, our study revealed a potential novel role of Glycine Position X in the Histone Variant H3.3.