Characterization of mechanisms regulating de-differentiation and the re-acquisition of stem cell identity

Characterization of mechanisms regulating de-differentiation and the re-acquisition of stem cell identity

Funding Type: 
New Faculty I
Grant Number: 
RN1-00544-B
Award Value: 
$382,773
Disease Focus: 
Aging
Stem Cell Use: 
Embryonic Stem Cell
Status: 
Active
Public Abstract: 
Statement of Benefit to California: 
Progress Report: 

Year 1

Stem cells are the building blocks during development of organisms as varied as plants and humans. In addition, adult or “tissue” stem cells provide for the maintenance and regeneration of tissues, such as blood and skin throughout the lifetime of an individual. The ability of stem cells to contribute to these processes depends on their unique ability to divide and generate both new stem cells (self-renewal) as well as specialized cell types (differentiation). In some tissues, cells that have already begun to specialize can revert or “de-differentiate” and assume stem cell properties, including the ability to self-renew. De-differentiation of specialized cells could provide a “reservoir” of cells that could act to replace stem cells lost due to wounding or aging. This proposal seeks to uncover the mechanisms that are utilized to regulate the process of de-differentiation and to compare these to the mechanisms that endow stem cells with the ability to self-renew using the fruit fly Drosophila melanogaster as well as pluripotent human cells. In the most recent funding period, we have found that the multiple sex combs (msx) gene in fruit flies encodes a protein that is important for balancing the number of proteins that control DNA packaging. Our research has shown that when a subset of these proteins is present in excess, it causes a low level DNA damage response. In return, it appears that another group of proteins that regulate self-renewal and differentiation are recruited away from their ‘normal’ job and used for DNA repair. As many chemotherapeutic agents and anti-cancer strategies involved using DNA damage to kill proliferating cells, our data suggest that not only will stem cell be susceptible to killing, but the normal differentiation programs might also be affected. Because the function of this gene is conserved in human cells, we speculate that understanding the function of this gene will provide insight In addition, we have continued to characterize the role of human Igf-II mRNA binding protein 1 (hIMP1) in pluripotent human cells and during early neural differentiation. Our primary progress in the current funding period was to identify genome-wide RNA targets for hIMP1, hIMP2, and LIN28 (a protein that associates with IMP proteins). Lastly, we have published a manuscript that shows that specialized stem cell microenvironments (also known as ‘niches’) are able to sustain similar numbers of stem cells, despite damage. This compensatory behavior is important because it suggests that the niche is an important component in sustaining stem cells after tissue damage or during wound repair. We will now use this observation as the basis for a screen to identify additional genes and pathways that regulate maintenance and regeneration of stem cell niches in more complex mammalian tissues.

Year 2

Stem cells are the building blocks during development of organisms as varied as plants and humans. In addition, adult or “tissue” stem cells provide for the maintenance and regeneration of tissues, such as blood and skin throughout the lifetime of an individual. The ability of stem cells to contribute to these processes depends on their unique ability to divide and generate both new stem cells (self-renewal) as well as specialized cell types (differentiation). In some tissues, cells that have already begun to specialize can revert or “de-differentiate” and assume stem cell properties, including the ability to self-renew. De-differentiation of specialized cells could provide a “reservoir” of cells that could act to replace stem cells lost due to wounding or aging. Our original goals were to uncover the mechanisms that are utilized to regulate the process of de-differentiation and to compare these to the mechanisms that endow stem cells with the ability to self-renew using the fruit fly Drosophila melanogaster as well as pluripotent human cells. In the most recent funding period, we have found that the multiple sex combs (msx) gene in fruit flies encodes a protein that is important for balancing the number of proteins that control DNA packaging. Our research has shown that when a subset of these proteins is present in excess, it causes a low level DNA damage response. In return, it appears that another group of proteins that regulate self-renewal and differentiation are recruited away from their ‘normal’ job and used for DNA repair. As many chemotherapeutic agents and anti-cancer strategies involved using DNA damage to kill proliferating cells, our data suggest that not only will stem cell be susceptible to killing, but the normal differentiation programs might also be affected. Because the function of this gene is conserved in human cells, we speculate that understanding the function of this gene will provide insight into how DNA damage as a consequence of environmental assault, drugs, or aging could lead to changes in gene expression that alter stem cell behavior. We published these data in the following publication: S. Landais, C. D’Alterio, and D.L. Jones. (2014) Persistent replicative stress regulates Polycomb phenotypes and tissue homeostasis in Drosophila. Cell Reports 7 (3): 859-870. In addition, we have continued to characterize the role of human Igf2 mRNA binding protein 1 (hIMP1) in pluripotent human cells. This family of RNA binding proteins is widely expressed during development and become re-expressed in many solid tumors. We made significant advances in the current funding period in identifying genome-wide RNA targets for hIMP1 and hIMP2, which could help explain the role of these RNA binding proteins play during development and carcinogenesis. We identified two novel, direct targets: the anti- cell death protein Bcl2 and a cell adhesion protein called Integrin beta 5 (INTß5). These data have been submitted and are currently under review in the journal Cell Stem Cell. A.E. Conway*, M. L. Wilbert*, S. Landais, B.Sundaraman, T. Y. Liang, G. Pratt, A. Essex, D. L. Jones#, G.W. Yeo#. The IGF2BP/IMP family of RNA-binding proteins controls an RNA network in pluripotent stem cells to maintain cell survival. (Under review for Cell Stem Cell). *-equal contribution, #- co-corresponding Lastly, we also embarked to identify and characterize the factors that are involved in regulating de-differentiation of progenitor cells in a well-characterized, genetically amenable stem cell system, the Drosophila male germ line. We have found that somatic stem cells can contribute directly to the stem cell microenvironment, also known as the ‘niche’ (Voog et al., Nature, 2008). Recently, we demonstrated that those supportive niche cells can de-differentiated back to the somatic stem cell lineage upon removal of a single transcription factor. These data were also recently published: J. Voog, S. Sandall, G. Hime, Resende, L.P.F., A. Aslanan, M. Loza-Coll, T. Hunter, M.T. Fuller, and D.L. Jones. (2014) Escargot restricts niche cell-stem cell conversion in the Drosophila testis. Cell Reports 7 (3): 722-734. Altogether, our lab published 9 primary research articles as a result of support from the California Institute of Regenerative Medicine.

© 2013 California Institute for Regenerative Medicine