Mechanisms Underlying the Diverse Functions of STAT3 in Embryonic Stem Cell Fate Regulation

Mechanisms Underlying the Diverse Functions of STAT3 in Embryonic Stem Cell Fate Regulation

Funding Type: 
New Faculty II
Grant Number: 
RN2-00938
Award Value: 
$2,261,174
Stem Cell Use: 
Embryonic Stem Cell
Status: 
Active
Public Abstract: 
Embryonic stem cells (ESCs) are derived from very early stage embryos. ESCs can be maintained in culture indefinitely while retain the ability to make any type of cell in the body. These properties make ESCs a very powerful tool to address basic biology questions. ESCs also offer an important renewable resource for future cell replacement therapies for many diseases such as Parkinson’s disease, spinal cord injury, etc. However, before the full potential of ESCs can be exploited in the clinic, we need to understand more about their biological properties so that we can control their fate towards either self-renewal or differentiation into a specific cell type required for cell replacement therapy. STAT3 is a major player in controlling the fates of a variety of cell types including ESCs. Recently we demonstrated that STAT3 has diverse and distinct roles in regulating cell fate in both mouse and human ESCs. In mouse ESCs, STAT3 is involved in cell adhesion, cell growth/survival and maintenance of self-renewal. Interestingly, STAT3 seems to have opposite roles in human ESCs. It induces growth arrest and differentiation of human ESCs. Why does the same factor play such diverse and contradictory roles between these very similar cells? The answer may lie on how STAT3 is in action. STAT3 is present in every type of cell. It contains six distinct functional regions. STAT3 can directly induce the expression of many genes. STAT3 can also cooperate with other proteins to regulate gene expression. We recently derived STAT3-/- ES cells in which the STAT3 gene was removed. These cells will provide us a powerful tool to dissect STAT3 function. We will first determine the role of each of its six functional regions. Then we will try to understand why they function differently. Is it because they induce different sets of genes, or because they cooperate with different partners? Understanding how STAT3 works is important for us to control the fate of ESCs, and for their eventual clinical application.
Statement of Benefit to California: 
Human embryonic stem cells (hESCs) can reproduce themselves in a culture dish. They can also give rise to every cell type in the body. In the future, hESCs may hold the key to replacing cells lost in many devastating diseases such as Parkinson’s disease, spinal cord injury, etc. Before hESCs can be used clinically, however, we must learn more about how to control their fate. STAT3 is a key player in regulating ES cell fate. STAT3 is also involved in the pathogenesis of diverse human cancers. In this proposed research, we will use a unique tool developed by us to understand STAT3's function. Our work will lead to a better understanding how hESC fate is regulated, which will be an important step towards achieving the therapeutic potential of hESCs. We also expect that our research will have a great implication in developing effective cancer therapies against novel STAT3 targets identified in this study.
Progress Report: 

Year 1

STAT3 is a major player in controlling the fates of a variety of cell types including embryonic stem cells. In this project, we proposed to investigate how STAT3 regulates embryonic stem cell fate. We found that STAT3 play diverse roles in regulating embryonic stem cell properties including self-renewal, cell adhesion and cell growth/survival. In the past reporting period, we have made the following progress: 1. Constructed nine different mutant forms of STAT3 and performed preliminary functional rescue experiments. 2. Established an inducible gene expression system which allows us to efficiently control the expression of different STAT3 mutants. 3. Performed preliminary experiments on identifying STAT3 target genes by microarray and real-time PCR analysis; 4. Examined the effect of STAT3 activation on human embryonic stem cell self-renewal. Based on the above results we have obtained and the tools we have developed, we are currently investigating the basic mechanisms how STAT3 regulates the fate of embryonic stem cells.

Year 2

STAT3 play a major role in controlling the fates of a variety of cell types including embryonic stem cells (ESCs). In this project, we proposed to investigate how STAT3 regulates ESC fate. In the past reporting period, we made the following progress: 1. Established and applied an inducible system in the mouse ESCs. This system allows us to tightly control the expression of different STAT3 mutants in mouse ESCs. 2. Discovered that phosphorylation of both Tyrosine 705 and Serine 727 is required for STAT3’s function in mouse ESCs. 3. Performed microarray analysis and identified 20 STAT3 target genes. 4. Further confirmed that STAT3 can enhance human ESC cell adhesion, but does not support human ESC self-renewal. Based on the above results we have obtained and the tools we have developed, we are currently investigating the basic mechanisms how STAT3 regulates the fate of ESCs.

Year 3

The overall goal of this project is to understand how STAT3 regulates the fate of mouse and human embryonic stem cells. In the past reporting period, we made the following major advancements: 1. The discovery that activation of STAT3 through phosphorylation at Tyrosine 705 can promote the conversion of mouse epiblast stem cells into a naïve embryonic stem cell state when Serine 727 phosphorylation of STAT3 is simultaneously blocked. 2. Identification of Gbx2 and En2 as the STAT3 downstream targets that play a critical role in sustaining mouse embryonic stem cell self-renewal. 3. Development of a small-molecule based culture condition that can maintain mouse epiblast stem cell and human embryonic stem cell self-renewal. Our findings suggest that human embryonic stem cells share defined features with mouse epiblast stem cells, but not with mouse embryonic stem cells. Based on these findings, we plan to investigate whether STAT3 regulates the fate of embryonic stem cells and epiblast stem cells through distinct mechanisms.

Year 4

The overall goal of this project is to understand how STAT3 regulates the fate of mouse and human embryonic stem cells. In the past reporting period, we focused on identifying and characterizing the genes that are induced by STAT3 and can promote embryonic stem cell self-renewal. Below are the major advancements we made in the past year: (1) The discovery that Gbx2, a gene directly induced by STAT3, can promote reprogramming to and retention of the pluripotent embryonic stem cell state. (2) Identification of Tfcp2l1 (Transcription factor CP2-like 1) as a STAT3 downstream target that plays an important role in sustaining mouse embryonic stem cell self-renewal. (3) Optimization of a small-molecule based culture condition for the maintenance of human embryonic stem cells. We further elucidated the mechanism by which these small molecules promote human embryonic stem cell self-renewal. Our findings will advance our efforts to better control stem cell fate, which is critical to the future of regenerative medicine.

Year 5

The overall goal of this project was to understand how STAT3 regulates the fate of mouse and human embryonic stem cells. In the past reporting period, we found that knockdown of Crif1, a protein associated with STAT3, impairs mouse embryonic stem cell self-renewal mediated by LIF/STAT3 signaling. We also identified and characterized two genes that are regulated by LIF/STAT3 signaling and involved in regulating embryonic stem cell fate. The first is Lef1. We found that Lef1 is negatively regulated by LIF/STAT3 signaling and knockdown of Lef1 promotes mouse embryonic stem cell self-renewal. The other gene is Cdx2 which is induced when STAT3 is hyperactivated. Induction of Cdx2 induces trophoblast differentiation of mouse embryonic stem cells. These findings provide an expanded understanding of embryonic stem cell fate regulation and are important for the future of cell replacement therapy using cells derived from embryonic stem cells.

© 2013 California Institute for Regenerative Medicine