Turner syndrome is caused by complete or partial loss of the second sex chromosome, which results in females with only one X chromosome (45,X). Turner females have a wide variety of symptoms, which include premature ovarian failure (POF) and infertility. POF in Turner females typically results from either the inability to form oocytes or their accelerated loss, leading to infertility. Yet, little is known about the gene and pathways underlying the requirement for two X chromosomes for human germ cell development. Moreover, mouse models do not assist in understanding the biology since mice with one X chromosome have few or no reproductive deficits. Because of the inaccessibility of human germ cells during development, a cell-based genetically tractable system to identify genes on the X chromosome is necessary to identify genes on the X chromosome that are necessary for human germ cell development. The hypothesis of this research proposal is that two copies of the human X chromosome are essential for early germ cell development by maintaining appropriate X-linked gene function. My research plan combines our understanding of human development and the genetic pathways involved in human cell fate, with our ability to produce human primordial germ cells (PGCs) from human embryonic stem cells (hESC) and induced pluripotent stem cells (iPSCs). Approximately15% of Turner syndrome females are mosaic for both a 45,X and a normal 46,XX cell population, and the remaining women exhibit varying degrees of X chromosome loss. Fibroblasts obtained from mosaic Turner females offer a source of genetically-matched 45,X and 46,XX somatic cells for derivation of iPSCs.
In Aim 1, I propose the derivation of at least two genetically-matched sets of iPSCs from mosaic Turner syndrome female fibroblasts.
In Aim 2, these iPSC lines, along with other 45,X and 46,XX iPSCs (from non mosaic sources) and a control 46,XX hESC line can be differentiated to the primordial germ cell lineage to uncover the necessity of two X chromosomes for proper germ cell development. To analyze the sufficiency of these X-linked genes, my third aim is to introduce X chromosome fragments into Turner syndrome iPSCs to rescue the deficiencies in germ cell development. With the completion of these studies I will determine if germ cell development relies on the presence of two functional X chromosomes. I will also determine which X-linked genes must be expressed in two copies for proper early germ cell development. I hope these studies will form the foundation for clinical applications to preserve and/or restore fertility in not only Turner syndrome women, but women with ovarian failure, cancer, and autoimmune disorders associated with failure to maintain pluripotent germ cells.
With the completion of these studies, I will determine if human female oocyte (egg) development relies on the presence of two functional X chromosomes. I will also determine which genes located on the X chromosome must be expressed in two copies for proper early oocyte development. Ultimately, these studies will form the foundation for clinical applications to preserve and/or restore fertility in women with ovarian failure, cancer or autoimmune disorders associated with failure to maintain pluripotent germ line cells.