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Injury-stimulated Hedgehog signaling promotes regenerative proliferation of Drosophila intestinal stem cells.

Tian A, Shi Q, Jiang A, Li S, Wang B, Jiang J - J. Cell Biol. (2015)

Bottom Line: Inhibition of Hh signaling in the ISC lineage compromised injury-induced ISC proliferation but had little if any effect on homeostatic proliferation.Furthermore, we show that Hh signaling is stimulated by DSS through the JNK pathway and that inhibition of Hh signaling in EBs prevented DSS-stimulated ISC proliferation.Hence, our study uncovers a JNK-Hh-JAK-STAT signaling axis in the regulation of regenerative stem cell proliferation.

View Article: PubMed Central - HTML - PubMed

Affiliation: Department of Developmental Biology and Department of Pharmacology, University of Texas Southwestern Medical Center at Dallas, Dallas, TX 75390.

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Hh signaling is required for DSS-stimulated Upd2 expression. (A–B′) Expression of 10XStat-dGFP in adult midguts expressing Su(H)ts (Con; A and A′) or Su(H)ts>SmoRNAi (B and B′) and treated with Suc or DSS for 1 d. (C–F) Adult female flies expressing esgts or esgts>StatRNAi for 8 d at 29°C were treated with Suc or DSS for 1 d before midguts were dissected out and immunostained for PH3 and DRAQ5. (G) Quantification of PH3+ cells in midguts of the indicated genotypes treated with Suc or DSS after adult flies were raised for 10 d at 29°C. Three independent experiments were performed and 20 guts were examined for each sample. Error bars are standard deviations. **, P < 0.01. (H) Quantification of Upd2 mRNA levels by RT-qPCR in Su(H)ts or Su(H)ts>SmoRNAi midguts treated with Suc or DSS. Three independent experiments were performed and error bars are standard deviations. **, P < 0.01. (I–L) Adult flies expressing Su(H)ts or Su(H)ts>Upd2RNAi for 10 d at 29°C were treated with Suc or DSS for 1 d before midguts were dissected out and immunostained for PH3 and DRAQ5. (M) Quantification of PH3+ cells in midguts of the indicated genotypes treated with Suc or DSS. Three independent experiments were performed and 20 guts were examined for each sample. Error bars are standard deviations. **, P < 0.01. Bars, 50 µm.
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fig6: Hh signaling is required for DSS-stimulated Upd2 expression. (A–B′) Expression of 10XStat-dGFP in adult midguts expressing Su(H)ts (Con; A and A′) or Su(H)ts>SmoRNAi (B and B′) and treated with Suc or DSS for 1 d. (C–F) Adult female flies expressing esgts or esgts>StatRNAi for 8 d at 29°C were treated with Suc or DSS for 1 d before midguts were dissected out and immunostained for PH3 and DRAQ5. (G) Quantification of PH3+ cells in midguts of the indicated genotypes treated with Suc or DSS after adult flies were raised for 10 d at 29°C. Three independent experiments were performed and 20 guts were examined for each sample. Error bars are standard deviations. **, P < 0.01. (H) Quantification of Upd2 mRNA levels by RT-qPCR in Su(H)ts or Su(H)ts>SmoRNAi midguts treated with Suc or DSS. Three independent experiments were performed and error bars are standard deviations. **, P < 0.01. (I–L) Adult flies expressing Su(H)ts or Su(H)ts>Upd2RNAi for 10 d at 29°C were treated with Suc or DSS for 1 d before midguts were dissected out and immunostained for PH3 and DRAQ5. (M) Quantification of PH3+ cells in midguts of the indicated genotypes treated with Suc or DSS. Three independent experiments were performed and 20 guts were examined for each sample. Error bars are standard deviations. **, P < 0.01. Bars, 50 µm.

Mentions: If DSS-mediated tissue damage stimulates ISC proliferation through the Hh signaling pathway, one would predict that feeding adult flies with DSS should induce Udp2 production and activate the JAK–STAT pathway. Indeed, we found that the JAK–STAT pathway reporter gene 10XStat-dGFP was activated in response to DSS treatment and that DSS-induced up-regulation of 10XStat-dGFP was blocked by inactivation of Hh signaling in EBs (Su(H)ts>SmoRNAi; Fig. 6, A–B′). Furthermore, knockdown of STAT in precursor cells (esgts>StatRNAi) blocked DSS-induced ISC proliferation (Fig. 6, C–G), suggesting that DSS-mediated tissue damage stimulates ISC through the JAK–STAT pathway.


Injury-stimulated Hedgehog signaling promotes regenerative proliferation of Drosophila intestinal stem cells.

Tian A, Shi Q, Jiang A, Li S, Wang B, Jiang J - J. Cell Biol. (2015)

Hh signaling is required for DSS-stimulated Upd2 expression. (A–B′) Expression of 10XStat-dGFP in adult midguts expressing Su(H)ts (Con; A and A′) or Su(H)ts>SmoRNAi (B and B′) and treated with Suc or DSS for 1 d. (C–F) Adult female flies expressing esgts or esgts>StatRNAi for 8 d at 29°C were treated with Suc or DSS for 1 d before midguts were dissected out and immunostained for PH3 and DRAQ5. (G) Quantification of PH3+ cells in midguts of the indicated genotypes treated with Suc or DSS after adult flies were raised for 10 d at 29°C. Three independent experiments were performed and 20 guts were examined for each sample. Error bars are standard deviations. **, P < 0.01. (H) Quantification of Upd2 mRNA levels by RT-qPCR in Su(H)ts or Su(H)ts>SmoRNAi midguts treated with Suc or DSS. Three independent experiments were performed and error bars are standard deviations. **, P < 0.01. (I–L) Adult flies expressing Su(H)ts or Su(H)ts>Upd2RNAi for 10 d at 29°C were treated with Suc or DSS for 1 d before midguts were dissected out and immunostained for PH3 and DRAQ5. (M) Quantification of PH3+ cells in midguts of the indicated genotypes treated with Suc or DSS. Three independent experiments were performed and 20 guts were examined for each sample. Error bars are standard deviations. **, P < 0.01. Bars, 50 µm.
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fig6: Hh signaling is required for DSS-stimulated Upd2 expression. (A–B′) Expression of 10XStat-dGFP in adult midguts expressing Su(H)ts (Con; A and A′) or Su(H)ts>SmoRNAi (B and B′) and treated with Suc or DSS for 1 d. (C–F) Adult female flies expressing esgts or esgts>StatRNAi for 8 d at 29°C were treated with Suc or DSS for 1 d before midguts were dissected out and immunostained for PH3 and DRAQ5. (G) Quantification of PH3+ cells in midguts of the indicated genotypes treated with Suc or DSS after adult flies were raised for 10 d at 29°C. Three independent experiments were performed and 20 guts were examined for each sample. Error bars are standard deviations. **, P < 0.01. (H) Quantification of Upd2 mRNA levels by RT-qPCR in Su(H)ts or Su(H)ts>SmoRNAi midguts treated with Suc or DSS. Three independent experiments were performed and error bars are standard deviations. **, P < 0.01. (I–L) Adult flies expressing Su(H)ts or Su(H)ts>Upd2RNAi for 10 d at 29°C were treated with Suc or DSS for 1 d before midguts were dissected out and immunostained for PH3 and DRAQ5. (M) Quantification of PH3+ cells in midguts of the indicated genotypes treated with Suc or DSS. Three independent experiments were performed and 20 guts were examined for each sample. Error bars are standard deviations. **, P < 0.01. Bars, 50 µm.
Mentions: If DSS-mediated tissue damage stimulates ISC proliferation through the Hh signaling pathway, one would predict that feeding adult flies with DSS should induce Udp2 production and activate the JAK–STAT pathway. Indeed, we found that the JAK–STAT pathway reporter gene 10XStat-dGFP was activated in response to DSS treatment and that DSS-induced up-regulation of 10XStat-dGFP was blocked by inactivation of Hh signaling in EBs (Su(H)ts>SmoRNAi; Fig. 6, A–B′). Furthermore, knockdown of STAT in precursor cells (esgts>StatRNAi) blocked DSS-induced ISC proliferation (Fig. 6, C–G), suggesting that DSS-mediated tissue damage stimulates ISC through the JAK–STAT pathway.

Bottom Line: Inhibition of Hh signaling in the ISC lineage compromised injury-induced ISC proliferation but had little if any effect on homeostatic proliferation.Furthermore, we show that Hh signaling is stimulated by DSS through the JNK pathway and that inhibition of Hh signaling in EBs prevented DSS-stimulated ISC proliferation.Hence, our study uncovers a JNK-Hh-JAK-STAT signaling axis in the regulation of regenerative stem cell proliferation.

View Article: PubMed Central - HTML - PubMed

Affiliation: Department of Developmental Biology and Department of Pharmacology, University of Texas Southwestern Medical Center at Dallas, Dallas, TX 75390.

Show MeSH
Related in: MedlinePlus