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A mex3 homolog is required for differentiation during planarian stem cell lineage development.

Zhu SJ, Hallows SE, Currie KW, Xu C, Pearson BJ - Elife (2015)

Bottom Line: In this study, we used transcriptional profiling of irradiation-sensitive and irradiation-insensitive cell populations and RNA interference (RNAi) functional screening to uncover markers and regulators of postmitotic progeny.We also demonstrated the utility of using mex3-1(RNAi) animals to identify additional progenitor markers.These results identified mex3-1 as a cell fate regulator, broadly required for differentiation, and suggest that mex3-1 helps to mediate the balance between ASC self-renewal and commitment.

View Article: PubMed Central - PubMed

Affiliation: Program in Developmental and Stem Cell Biology, The Hospital for Sick Children, Toronto, Canada.

ABSTRACT
Neoblasts are adult stem cells (ASCs) in planarians that sustain cell replacement during homeostasis and regeneration of any missing tissue. While numerous studies have examined genes underlying neoblast pluripotency, molecular pathways driving postmitotic fates remain poorly defined. In this study, we used transcriptional profiling of irradiation-sensitive and irradiation-insensitive cell populations and RNA interference (RNAi) functional screening to uncover markers and regulators of postmitotic progeny. We identified 32 new markers distinguishing two main epithelial progenitor populations and a planarian homolog to the MEX3 RNA-binding protein (Smed-mex3-1) as a key regulator of lineage progression. mex3-1 was required for generating differentiated cells of multiple lineages, while restricting the size of the stem cell compartment. We also demonstrated the utility of using mex3-1(RNAi) animals to identify additional progenitor markers. These results identified mex3-1 as a cell fate regulator, broadly required for differentiation, and suggest that mex3-1 helps to mediate the balance between ASC self-renewal and commitment.

No MeSH data available.


Related in: MedlinePlus

mex3-1 RNAi impairs brain differentiation.(A) Expression of mex3-1 in neural-restricted neoblast progeny by mex3-1 and chat dFISH with PIWI-1 immunolabeling. Percentage of chat+PIWI-1+ cells which express mex3-1 is indicated at top-right of panel. Arrows indicate example triple-labeled cells. Dashed box indicates area of magnified panels. Left scale bar, 50 μm; right scale bar, 10 μm. (B) Animals were administered BrdU at 6 days after RNAi and assessed at 5 days post-BrdU. Scale bar, 100 μm. All images shown are single confocal planes.DOI:http://dx.doi.org/10.7554/eLife.07025.018
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fig6s1: mex3-1 RNAi impairs brain differentiation.(A) Expression of mex3-1 in neural-restricted neoblast progeny by mex3-1 and chat dFISH with PIWI-1 immunolabeling. Percentage of chat+PIWI-1+ cells which express mex3-1 is indicated at top-right of panel. Arrows indicate example triple-labeled cells. Dashed box indicates area of magnified panels. Left scale bar, 50 μm; right scale bar, 10 μm. (B) Animals were administered BrdU at 6 days after RNAi and assessed at 5 days post-BrdU. Scale bar, 100 μm. All images shown are single confocal planes.DOI:http://dx.doi.org/10.7554/eLife.07025.018

Mentions: Previously, it was shown that even by selectively abolishing zeta-neoblasts and epithelial turnover, a regenerative blastema can still form with differentiated tissues such as brain, protonephridia, intestine, and muscle (van Wolfswinkel et al., 2014). Given that mex3-1(RNAi) animals were unable to produce any regenerative blastema, we hypothesized that mex3-1 may have a crucial role in the broad specification of multiple lineages. To examine this possibility, we assessed mex3-1(RNAi) worms for changes in the number of lineage-specified neoblast progeny of other tissue types in various body regions (Figure 6C). As PIWI-1 protein persists in immediate postmitotic stem cell descendants for up to 72 hr (Guo et al., 2006), co-localization of PIWI-1 with tissue-specific markers was used to identify lineage-restricted neoblast descendants, encompassing undifferentiated progenitors and newly differentiating cells. The neural genes, chat and coe, eye-specific transcription factor ovo, pharyngeal marker FoxA, and protonephridial marker six1/2-2 were used as markers to indicate differentiation toward their respective tissues (Scimone et al., 2011; Wagner et al., 2011; Lapan and Reddien, 2012; Cowles et al., 2013; Adler et al., 2014). Quantification of lineage-restricted neoblast progeny in intact mex3-1(RNAi) worms 12 days after RNAi showed significant reductions in all examined cell types (Figure 6D). We observed that virtually all chat+PIWI-1+ cells expressed mex3-1 (Figure 6—figure supplement 1A), suggesting a direct role for mex3-1 in regulating differentiation outside the epithelial lineage. Concordant with decreased production of lineage-restricted neoblast progeny, we also observed that 5 days following labeling with BrdU, the entry of new cells into brain, intestine, and pharynx was significantly decreased in mex3-1(RNAi) animals (Figure 6E, Figure 6—figure supplement 1B). These data demonstrate that the diminished capacity of mex3-1(RNAi) animals to produce differentiated progeny is not restricted to the epidermal lineage but is characteristic of multiple lineages in planarians.


A mex3 homolog is required for differentiation during planarian stem cell lineage development.

Zhu SJ, Hallows SE, Currie KW, Xu C, Pearson BJ - Elife (2015)

mex3-1 RNAi impairs brain differentiation.(A) Expression of mex3-1 in neural-restricted neoblast progeny by mex3-1 and chat dFISH with PIWI-1 immunolabeling. Percentage of chat+PIWI-1+ cells which express mex3-1 is indicated at top-right of panel. Arrows indicate example triple-labeled cells. Dashed box indicates area of magnified panels. Left scale bar, 50 μm; right scale bar, 10 μm. (B) Animals were administered BrdU at 6 days after RNAi and assessed at 5 days post-BrdU. Scale bar, 100 μm. All images shown are single confocal planes.DOI:http://dx.doi.org/10.7554/eLife.07025.018
© Copyright Policy
Related In: Results  -  Collection

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fig6s1: mex3-1 RNAi impairs brain differentiation.(A) Expression of mex3-1 in neural-restricted neoblast progeny by mex3-1 and chat dFISH with PIWI-1 immunolabeling. Percentage of chat+PIWI-1+ cells which express mex3-1 is indicated at top-right of panel. Arrows indicate example triple-labeled cells. Dashed box indicates area of magnified panels. Left scale bar, 50 μm; right scale bar, 10 μm. (B) Animals were administered BrdU at 6 days after RNAi and assessed at 5 days post-BrdU. Scale bar, 100 μm. All images shown are single confocal planes.DOI:http://dx.doi.org/10.7554/eLife.07025.018
Mentions: Previously, it was shown that even by selectively abolishing zeta-neoblasts and epithelial turnover, a regenerative blastema can still form with differentiated tissues such as brain, protonephridia, intestine, and muscle (van Wolfswinkel et al., 2014). Given that mex3-1(RNAi) animals were unable to produce any regenerative blastema, we hypothesized that mex3-1 may have a crucial role in the broad specification of multiple lineages. To examine this possibility, we assessed mex3-1(RNAi) worms for changes in the number of lineage-specified neoblast progeny of other tissue types in various body regions (Figure 6C). As PIWI-1 protein persists in immediate postmitotic stem cell descendants for up to 72 hr (Guo et al., 2006), co-localization of PIWI-1 with tissue-specific markers was used to identify lineage-restricted neoblast descendants, encompassing undifferentiated progenitors and newly differentiating cells. The neural genes, chat and coe, eye-specific transcription factor ovo, pharyngeal marker FoxA, and protonephridial marker six1/2-2 were used as markers to indicate differentiation toward their respective tissues (Scimone et al., 2011; Wagner et al., 2011; Lapan and Reddien, 2012; Cowles et al., 2013; Adler et al., 2014). Quantification of lineage-restricted neoblast progeny in intact mex3-1(RNAi) worms 12 days after RNAi showed significant reductions in all examined cell types (Figure 6D). We observed that virtually all chat+PIWI-1+ cells expressed mex3-1 (Figure 6—figure supplement 1A), suggesting a direct role for mex3-1 in regulating differentiation outside the epithelial lineage. Concordant with decreased production of lineage-restricted neoblast progeny, we also observed that 5 days following labeling with BrdU, the entry of new cells into brain, intestine, and pharynx was significantly decreased in mex3-1(RNAi) animals (Figure 6E, Figure 6—figure supplement 1B). These data demonstrate that the diminished capacity of mex3-1(RNAi) animals to produce differentiated progeny is not restricted to the epidermal lineage but is characteristic of multiple lineages in planarians.

Bottom Line: In this study, we used transcriptional profiling of irradiation-sensitive and irradiation-insensitive cell populations and RNA interference (RNAi) functional screening to uncover markers and regulators of postmitotic progeny.We also demonstrated the utility of using mex3-1(RNAi) animals to identify additional progenitor markers.These results identified mex3-1 as a cell fate regulator, broadly required for differentiation, and suggest that mex3-1 helps to mediate the balance between ASC self-renewal and commitment.

View Article: PubMed Central - PubMed

Affiliation: Program in Developmental and Stem Cell Biology, The Hospital for Sick Children, Toronto, Canada.

ABSTRACT
Neoblasts are adult stem cells (ASCs) in planarians that sustain cell replacement during homeostasis and regeneration of any missing tissue. While numerous studies have examined genes underlying neoblast pluripotency, molecular pathways driving postmitotic fates remain poorly defined. In this study, we used transcriptional profiling of irradiation-sensitive and irradiation-insensitive cell populations and RNA interference (RNAi) functional screening to uncover markers and regulators of postmitotic progeny. We identified 32 new markers distinguishing two main epithelial progenitor populations and a planarian homolog to the MEX3 RNA-binding protein (Smed-mex3-1) as a key regulator of lineage progression. mex3-1 was required for generating differentiated cells of multiple lineages, while restricting the size of the stem cell compartment. We also demonstrated the utility of using mex3-1(RNAi) animals to identify additional progenitor markers. These results identified mex3-1 as a cell fate regulator, broadly required for differentiation, and suggest that mex3-1 helps to mediate the balance between ASC self-renewal and commitment.

No MeSH data available.


Related in: MedlinePlus