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Full Abstracts – STEM CELLS
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Gene regulatory networks controlling hematopoietic progenitor niche cell production and differentiation in the
Drosophila
lymph gland.
Tsuyoshi
Tokusumi, Yumiko Tokusumi, Douglas A. Shoue, Robert A. Schulz. Department of Biological Sciences, University of Notre Dame, Notre Dame, IN.
The lymph gland is a specialized organ for hematopoiesis, utilized during larval development in
Drosophila
. This tissue is composed of distinct cellular
domains populated by blood cell progenitors (the medullary zone), niche cells that regulate the choice between progenitor quiescence and hemocyte
differentiation [the posterior signaling center (PSC)], and mature blood cells of distinct lineages (the cortical zone). Cells of the PSC express the Hedgehog
(Hh) signaling molecule, which instructs cells within the neighboring medullary zone to maintain a hematopoietic precursor state while preventing hemocyte
differentiation. As a means to understand the regulatory mechanisms controlling Hh production, we characterized a PSC-active transcriptional enhancer that
drives
hh
expression in niche cells. Our findings indicate that the GATA factor Serpent (Srp) is essential for
hh
activation in niche cells, whereas the
Suppressor of Hairless and U-shaped transcriptional regulators prevent
hh
expression in hemocyte progenitors and differentiated hemocytes. Furthermore,
Srp function is required for the proper differentiation of niche cells. With the
hh-GFP
transgene serving as a sensitive marker for the PSC, we initiated an
in
vivo
RNAi screen to discover genes whose function was required for the correct production of niche cells. The Gal4 driver employed was the
P85col-Gal4
strain and it has been used thus far to express 820 RNAi sequences from VDRC. Thorough this screen, we have identified ~100 genes regulating PSC
formation and function. Among them, we focused on two important mechanisms: Brahma complex and Akt1/Tor pathway. Brahma complex, which is an
ATP-dependent chromatin remodeling complex, can genetically interact with
srp
to regulate
hh
gene expression and filopodia formation. On the other hand,
Akt1/Tor pathway can regulate PSC cell growth depended on nutrient conditions. Our results demonstrate novel regulatory mechanisms at work within the
stem-like niche.
40
hedgehog
regulates self-renewal and niche competition in the
Drosophila
testis.
Marc Amoyel
1
, Michael Burel
1
, Erika Bach
1,2
. 1) Pharmacology, New
York University School of Medicine, New York, NY; 2) Kimmel Stem Cell Center.
Stem cells are critical for tissue regeneration during adulthood. The
Drosophila
testis is one of the best characterised stem cell-niche systems, as both the
niche and stem cells are anatomically and molecularly well defined and some signals controlling self-renewal have been established. Somatic hub cells
comprise the niche, which maintains two stem cell populations, germ-line stem cells (GSCs) and somatic cyst stem cells (CySCs). GSC self-renewal requires
activation of Bone Morphogenetic Protein (BMP) receptors by BMP ligands produced by the hub and CySCs, while CySC self-renewal requires activation
of the JAK/STAT pathway by Unpaired (Upd), produced only by hub cells. The requirement of additional signalling pathways in self renewal and niche
occupancy of testis stem cells is not known. While it is known that Hh is expressed in niche cells, a role for Hh in testis stem cells has not been reported. We
find that Hh signalling is detected in CySCs but not in GSCs. Consistent with this we find that Hh signalling is required cell-autonomously in CySCs but not
in GSCs for self-renewal. To determine if Hh affects activity of Stat92E, we examined JAK/STAT pathway readouts in Hh mutant CySCs and vice versa.
We confirmed that each pathway is not affected by disruption of the other. This suggests that Hh and JAK/STAT pathways act in parallel in CySC self-
renewal, which we confirmed by clonal epistasis experiments. We also observed that CySCs with sustained Hh signalling compete with resident CySCs and
GSCs and colonise the niche by displacing both wild-type populations. CySCs with increased Hh proliferate faster than wild-type CySCs. Experimentally
accelerating the cell cycle in testis stem cells alters population dynamics at the niche, leading to preferential accumulation of the highly proliferative
population at the expense of the wild-type. These data support a model in which Hh regulates cell cycle progression of CySCs, which is critical for
regulating homeostasis at the niche.
41
A histone demethylase dUTX regulates crosstalk among different cell types in the
Drosophila
testis stem cell niche.
Xin Chen, Lama Tarayrah. Dept
Biol, Johns Hopkins Univ, Baltimore, MD.
Adult stem cells reside in microenvironments, or niches, that provide signals from surrounding cells to the stem cells to prevent differentiation. The
Drosophila
male germline stem cell (GSC) niche is considered to be one of the best characterized niches in which GSCs associate with two types of somatic
cells: hub cells and cyst stem cells (CySCs). It has been reported that somatic cells play important roles to maintain GSCs through the JAK-STAT signaling
pathway. Here we show that dUTX contributes to the maintenance of male GSC niche by affecting all three cell types in the niche (i.e. GSCs, CySCs and
hub cells). dUTX is the sole
Drosophila
homolog of mammalian UTX, which has been shown to act as a specific demethylase of trimethylation on lysine 27
of histone H3 (H3K27me3)
in vivo
. We found that GSCs from
dUTX
mutant testes have a significantly lower mitotic index and a higher rate of misoriented
centrosomes, suggesting a decrease in GSC activity. In addition, the hub size and structure are abnormal in
dUTX
mutant testis. Interestingly, the hub
phenotype could be recapitulated by knocking-down dUTX in germ cells, suggesting normal function of dUTX is required in germ cells to maintain the hub
architecture. Surprisingly, not only the hub structure changes, a transcription factor zinc finger homeodomain-1(Zfh-1), which is normally turned on in
CySCs and early cyst cells but not in hub cells, is ectopically expressed in
dUTX
mutant hub cells. Further analysis indicated that dUTX acts in CySCs to
maintain the silenced state of Zfh-1 in hub cells. Finally, we found that dUTX may directly regulate the chromatin state of JAK-STAT signaling pathway
components to coordinate behavior of different cell types in the testis niche. Together, our data demonstrate that a chromatin factor regulates the key
signaling pathway and crosstalk among different cell types in the
Drosophila
testis stem cell niche: in addition to the known signaling from hub and CySCs
to GSCs, both GSCs and CySCs can send feedback to hub cells to maintain the integrity and functionality of the niche.