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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

Quantification of stem cells in mex3-1(RNAi) animals.Stem cells were quantified in mid-pharyngeal and tail cross sections of intact worms at 6 and 9 days after RNAi by piwi-1 FISH. Single confocal planes are shown; dorsal, top. Scale bar, 200 μm. *p < 0.05; **p < 0.01 (Student's t-test). 10 animals were counted per RNAi treatment.DOI:http://dx.doi.org/10.7554/eLife.07025.016
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fig5s1: Quantification of stem cells in mex3-1(RNAi) animals.Stem cells were quantified in mid-pharyngeal and tail cross sections of intact worms at 6 and 9 days after RNAi by piwi-1 FISH. Single confocal planes are shown; dorsal, top. Scale bar, 200 μm. *p < 0.05; **p < 0.01 (Student's t-test). 10 animals were counted per RNAi treatment.DOI:http://dx.doi.org/10.7554/eLife.07025.016

Mentions: Given that early and late progeny cell fates failed to be specified and adopted in mex3-1(RNAi) animals despite ongoing stem cell divisions, we hypothesized that there was a loss in stem cell lineage asymmetry in favor of stem cell self-renewal. We sought to determine whether this was the case by quantifying piwi-1+ stem cells in transverse cross sections after RNAi. By 6 days after RNAi, we observed highly significant increases in the number of piwi-1+ stem cells in mex3-1(RNAi) animals compared to controls, which rose to a 50% increase by day 12 (Figure 5C, Figure 5—figure supplement 1). The expansion of piwi-1+ stem cells after mex3-1 knockdown could either reflect a global increase in all stem cell subclasses or reflect an increase in a specific subclass (zeta-, sigma-, and gamma-neoblasts) (van Wolfswinkel et al., 2014). To ascertain whether subclasses were selectively affected, we quantified the number of piwi-1+ stem cells belonging to each subclass using the following probes: zfp-1 for the zeta subclass, hnf4 for the gamma subclass, and a pooled mix of soxP-1 and soxP-2 for the sigma subclass. Assessment at 12 days after RNAi revealed that all three subclasses were significantly increased by approximately 50% in mex3-1(RNAi) animals compared to controls (Figure 5D). Together, these results demonstrated that the failure to specify epithelial progenitors in mex3-1(RNAi) animals was not due to loss of zeta-neoblasts, and suggested that the expansion of the stem cell pools was due to an imbalance in cell fates favoring stem cell self-renewal over differentiation.


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)

Quantification of stem cells in mex3-1(RNAi) animals.Stem cells were quantified in mid-pharyngeal and tail cross sections of intact worms at 6 and 9 days after RNAi by piwi-1 FISH. Single confocal planes are shown; dorsal, top. Scale bar, 200 μm. *p < 0.05; **p < 0.01 (Student's t-test). 10 animals were counted per RNAi treatment.DOI:http://dx.doi.org/10.7554/eLife.07025.016
© Copyright Policy
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC4507787&req=5

fig5s1: Quantification of stem cells in mex3-1(RNAi) animals.Stem cells were quantified in mid-pharyngeal and tail cross sections of intact worms at 6 and 9 days after RNAi by piwi-1 FISH. Single confocal planes are shown; dorsal, top. Scale bar, 200 μm. *p < 0.05; **p < 0.01 (Student's t-test). 10 animals were counted per RNAi treatment.DOI:http://dx.doi.org/10.7554/eLife.07025.016
Mentions: Given that early and late progeny cell fates failed to be specified and adopted in mex3-1(RNAi) animals despite ongoing stem cell divisions, we hypothesized that there was a loss in stem cell lineage asymmetry in favor of stem cell self-renewal. We sought to determine whether this was the case by quantifying piwi-1+ stem cells in transverse cross sections after RNAi. By 6 days after RNAi, we observed highly significant increases in the number of piwi-1+ stem cells in mex3-1(RNAi) animals compared to controls, which rose to a 50% increase by day 12 (Figure 5C, Figure 5—figure supplement 1). The expansion of piwi-1+ stem cells after mex3-1 knockdown could either reflect a global increase in all stem cell subclasses or reflect an increase in a specific subclass (zeta-, sigma-, and gamma-neoblasts) (van Wolfswinkel et al., 2014). To ascertain whether subclasses were selectively affected, we quantified the number of piwi-1+ stem cells belonging to each subclass using the following probes: zfp-1 for the zeta subclass, hnf4 for the gamma subclass, and a pooled mix of soxP-1 and soxP-2 for the sigma subclass. Assessment at 12 days after RNAi revealed that all three subclasses were significantly increased by approximately 50% in mex3-1(RNAi) animals compared to controls (Figure 5D). Together, these results demonstrated that the failure to specify epithelial progenitors in mex3-1(RNAi) animals was not due to loss of zeta-neoblasts, and suggested that the expansion of the stem cell pools was due to an imbalance in cell fates favoring stem cell self-renewal over differentiation.

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