Functional characterization of the pivotal role of a novel zinc finger protein ZNF206 in regulating early lineage allocation & commitment
Human embryonic stem cells (hESCs) come from a region of the early pre-implantation embryo called the “inner cell mass” (ICM). Understanding the mechanisms by which the ICM remains plastic yet, at the appropriate time, gives rise to a range of specific cell types will provide us not only with a better understanding of how best to use hESCs therapeutically, but also with insights into root causes of normal and abnormal development and reproduction. We have learned that a population of cells, called the primitive endoderm (PE), which helps regulate the function of the ICM in the body, also seems to emerge spontaneously – almost by default – from hESCs in culture. PE cells seem to play the same role in the culture dish that they play in the body – improving the function of ICM-derived cells, i.e., hESCs. Of all of the progeny of the hESC, we actually believe that PE cells are the first to emerge (likely to serve as “helper cells”). We have identified a novel gene that we think regulates the formation of the PE and hence ultimately the health, function, and fate of the ICM and its culture counterpart, the hESC. It works in concert, we believe, with some of the other handful of “stemness” genes previously identified. In this project, we propose to characterize the qualities, molecular partners, and function of this novel gene. With such knowledge in hand, this molecule may be used to manipulate hESCs for use therapeutically as well as for modeling diseases of development and reproduction. It may be a gene that can be monitored during pregnancies to determine early on fundamental developmental flaws.
Finally, in the course of discovering this gene and the PE-like cells that spontaneously emerge from the hESCs, we learned what the minimal essential components are for insuring plasticity (“pluripotency”) and self-renewal of hESCs in culture. From this we devised a culture system that contained only known elements – heretofore an obstacle for the clinical use of hESCs. This culture system derives its efficacy precisely because it induces the unfolding in vitro (within a closed system) of a process that very much emulates the formation & maintenance of the ICM in vivo, including the emergence of a PE which supports these ICM-like cells (i.e., the hESCs). Such a defined system has already allowed us to begin deriving and maintaining hESCs suitable for clinical use. It also will permit the better identification of molecules that permit hESCs to become particular cell types. The hESCs themselves can be used to understand how various cell types during development are related to each other. And, since these PE cells can be isolated and grown as a separate homogenous population, not only can they be used as a product themselves to help hESC function, but the molecules they make -- some of which are known but hard to purify in bulk and some of which are novel – can be used to improve the health and utility of hESCs.
There are more than 4,000 different know birth defects ranging from minor to serious. According to The American College of Obstetricians and Gynecologist (ACOG), out of every 100 babies born in the United States, three have some kind of major birth defect, making birth defects the leading cause of death in the first year. 60%≈70% of birth defects still have unknown causes, and many of them are due to abnormality of early embryonic development. Indeed, the entire process of human development and pregnancy remains poorly understood on a molecular level: despite the identification of many key players, our knowledge of the regulatory cascades originating with the preimplantation embryo and leading to birth of a child barely scratch the surface.
The inner cell mass that gives rise to embryonic stem (ES) cells is at the root of the entire process of human development. The mechanisms that determine ES cells defining properties (self-renewal, pluripotency, and germline competency) are not completely understood, but appear to involved transcriptional regulation, because transcription factors (e.g. OCT4, and NANOG) have been identified that effect these properties. Mutations in these and other transcription factors that are expressed in early development often cause defects in development and reproduction, and are often embryonic lethal as nulls demonstrating their critical importance.
On the basis of our preliminary data, ZNF206 appears likely to be part of a novel transcriptional regulatory hierarchy of ES cell function and/or early development. In this application, we are proposing to determine the molecular, cellular, and physiological functions of ZNF206. Understanding the specific roles(s) it plays has the potential to contribute directly to fewer birth defects and lower incidence of miscarriages and the treatment of birth defects, via either screening or by chemical intervention. Transcription factors are a major category of drug targets, and recent studies suggest that it may also be possible to pharmacologically manipulate the activity of specific DNA sequences (e.g. the binding sites of transcription factors).
Since ZNF206 is expressed specifically in ES cells and may be involved in regulating early primitive endoderm lineage commitment, the results of our research may be applicable not only to the understanding, diagnosing, and treating the causes of birth defects, but also to the treatment of medical disorders. Since ES cells can give rise to different kinds of cells that make our body they are uniquely suited for regenerative medicine. ES cells have been proposed as therapeutic tools for a wide range of debilitating diseases including Parkinson’s and Alzheimer’s diseases, spinal cord injury, stroke, burns, heart disease, and other disorders. ES cells have also been advocated for us in treating birth defects, such as severe combined immunodeficiency disease-X linked recessive (X-SCID), Wiskott-Aldrich syndrome (WAS) and chromosomal