GIST Repository
College of Life Sciences and Medical Engineering
Department of Life Sciences
3. Theses(Master)
Functional analysis of Gm11545 in lineage specification during preimplantation embryogenesis in mice
- Abstract
- After fertilization, embryos undergo continuous cleavage, leading to the first cell fate decision, known as the first lineage specification. This process is tightly regulated by various molecular mechanisms, including the Hippo signaling pathway. Despite extensive research, the exact regulatory mechanisms governing the first lineage specification remain unclear. In this study, I investigated the role of an unnamed gene, designated Gm11545, in early embryogenesis, particularly its involvement in the Hippo signaling pathway during the first cell lineage specification in mouse embryos. Knockdown (KD) of Gm11545 led to an increase in cytoplasmic F-actin meshwork in the outer blastomeres of the morula, which caused the cytoplasmic localization of angiomotin (AMOT) and a reduction in nuclear yes-associated protein (YAP) localization. Consequently, caudal type homeobox 2 (CDX2) expression, a critical transcription factor for trophectoderm differentiation, was downregulated, ultimately leading to failure in blastocyst formation. Additionally, in blastocysts developed from chimeric morulae of which each embryo contains both normal and KD blastomeres, Gm11545 KD blastomeres were predominantly localized in the inner cell mass (ICM) region, suggesting that Gm11545 plays a significant role in promoting trophectoderm (TE) specification. These findings highlight Gm11545 as a crucial regulator of asymmetric Hippo signaling between inner and outer blastomeres, influencing cell fate decisions during the first lineage specification in embryogenesis. Keywords Embryo, lineage specification, Gm11545, Hippo pathway
- Issued Date
- 2025
- Type
- Thesis
- Alternative Author(s)
- Jaeho Song
- Department
- 대학원 생명과학부
- Advisor
- Cho, Chung Hee
- Table Of Contents
- ABSTRACT ⅰ
CONTENTS ⅱ
LIST OF FIGURES ⅳ
1. INTRODUCTION 1
2. MATERIALS AND METHODS 3
2. 1. Mouse embryo collection and culture 3
2. 2. Mouse oocyte collection and In vitro maturation 3
2. 3. RNA extraction and RT-PCR 3
2. 4. Real-Time PCR 3
2. 5. Generation of dsRNA 4
2. 6. Generation of mRNA 4
2. 7. Microinjection 4
2. 8. Immunofluorescence staining (IF) 5
2. 9. Imaging and quantification 5
2. 10. Morula aggregation assay 5
2. 11. Statistical analysis 6
3. RESULTS 7
3. 1. Localization of Gm11545 during embryogenesis 7
3. 2. Confirmation of impaired blastocyst formation in Gm11545 KD embryos 7
3. 3. Expression of lineage marker genes in Gm11545 KD embryos 7
3. 4. Gm11545 KD decreases CDX2 expression in outer blastomeres. 8
3. 5. Gm11545 KD disrupts asymmetric Hippo signaling pathway. 9
3. 6. Gm11545 KD did not affect apical F-actin stability. 10
3. 7. Gm11545 KD promotes ICM lineage specification. 11
4. DISCUSSION AND CONCLUSION 31
5. REFERENCES 35
6. ABSTRACT IN KOREAN 39
7. ACKNOWLEDGEMENT 40
LIST OF FIGURES & TABLES
Table 1. List of primers used for qRT-PCR, RT-PCR. 12
Figure 1. Localization of mCherry tagged Gm11545 during embryogenesis 13
Figure 2. Knockdown analysis with microinjection of Gm11545 dsRNA during preimplantation
embryogenesis. 15
Figure 3. Knockdown analysis of Gm11545 with microinjection of Gm11545 dsRNA during oocyte
maturation. 17
Figure 4. Expression of ICM, TE marker, and YAP-TEAD4 target genes in morula stage of Gm 11545
KD embryos. 19
Figure 5. CDX2 and OCT4 expression in morula stage of Gm11545 KD embryos. 21
Figure 6. YAP, AMOT and F-actin expression in the morula stage of Gm11545 KD embryos 23
Figure 7. FRAP analysis of apical F-actin complex 25
Figure 8. Morula aggregation assay 27
Figure 9. Schematic diagram of main results 29
- Degree
- Master
- Appears in Collections:
- Department of Life Sciences > 3. Theses(Master)
- 파일 목록
-
-
관련 파일이 존재하지 않습니다.
-
Items in Repository are protected by copyright, with all rights reserved, unless otherwise indicated.