Genesis of the human germline
type: CreativeWork
description: The primary aim of this thesis was to examine the specification of human primordial germ cells (PGCs) and the mechanism regulating the transient gain and loss of competence for germ cell fate. Specification of PGCs in humans occurs during weeks 2 to 3 in gastrulating embryos, which eventually develop into sperm and ova, generate totipotency at fertilisation, and transmit genetic and epigenetic information for development. Since direct studies on early human embryos are not feasible, I used an established human pluripotent stem cell-based in vitro model for early human development. Accordingly, I used human pluripotent stem cells (PSCs) to induce mesendoderm (ME), which at midpoint progresses through a pre-mesendoderm (PreME) state that transiently acquires competence for PGC fate and commits to PGC-like cells (PGCLCs) in response to BMP signalling. To address these questions, I performed highly-resolved transcriptomic profiling and careful epigenomic characterisation of cis-regulatory regions at key steps during the in vitro model. To determine the regulation of the transient gain and loss of competence for germ cell fate, I re-engineered a CRISPR toolkit for activation and inactivation to test candidates for germ cell competence. I found the inhibition of OTX2 increased the efficiency of human PGCLC specification, which is consistent with a transient reduction in OTX2 in the germline competent PreME against rising levels of EOMES. The OTX2 levels continue to increase in ME cells that gain competence for the mesoderm and endoderm fates but are incompetent for germ cell fate. Using the CRISPR toolkit, I further show that SOX17 enhancers are the critical targets of the regulatory circuitry of germline competence. SOX17 and TFAP2C initiate a core germline program, including induction of PRDM1 by SOX17 and maintenance of pluripotency factors, POU5F1 and NANOG. I also explored PGC competence and specification using highly resolved single-cell transcriptomic profiling of …
abstract: The primary aim of this thesis was to examine the specification of human primordial germ cells (PGCs) and the mechanism regulating the transient gain and loss of competence for germ cell fate. Specification of PGCs in humans occurs during weeks 2 to 3 in gastrulating embryos, which eventually develop into sperm and ova, generate totipotency at fertilisation, and transmit genetic and epigenetic information for development. Since direct studies on early human embryos are not feasible, I used an established human pluripotent stem cell-based in vitro model for early human development. Accordingly, I used human pluripotent stem cells (PSCs) to induce mesendoderm (ME), which at midpoint progresses through a pre-mesendoderm (PreME) state that transiently acquires competence for PGC fate and commits to PGC-like cells (PGCLCs) in response to BMP signalling. To address these questions, I performed highly-resolved transcriptomic profiling and careful epigenomic characterisation of cis-regulatory regions at key steps during the in vitro model. To determine the regulation of the transient gain and loss of competence for germ cell fate, I re-engineered a CRISPR toolkit for activation and inactivation to test candidates for germ cell competence. I found the inhibition of OTX2 increased the efficiency of human PGCLC specification, which is consistent with a transient reduction in OTX2 in the germline competent PreME against rising levels of EOMES. The OTX2 levels continue to increase in ME cells that gain competence for the mesoderm and endoderm fates but are incompetent for germ cell fate. Using the CRISPR toolkit, I further show that SOX17 enhancers are the critical targets of the regulatory circuitry of germline competence. SOX17 and TFAP2C initiate a core germline program, including induction of PRDM1 by SOX17 and maintenance of pluripotency factors, POU5F1 and NANOG. I also explored PGC competence and specification using highly resolved single-cell transcriptomic profiling of …
author: Castillo Venzor, Aracely
language: eng
keywords: Germline, human PGC, PGCLC
license: https://www.rioxx.net/licenses/all-rights-reserved/
date published: 2021-12-31
date modified: 2026-07-22T22:24:03Z
provider: Cambridge University
publisher: Networked Digital Library of Theses and Dissertations
type: WebPage
source organization: University of Cambridge

PropertyValue
property ID: DOI
value: 10.17863/CAM.88839