1. Developmental Biology

Numb provides a fail-safe mechanism for intestinal stem cell self-renewal in adult Drosophila midgut

  1. Mengjie Li
  2. Aiguo Tian
  3. Jin Jiang  Is a corresponding author
  1. Department of Molecular Biology, University of Texas Southwestern Medical Center at Dallas, United States
  2. Department of Pharmacology, University of Texas Southwestern Medical Center at Dallas, United States
https://doi.org/10.7554/eLife.104723.3
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Cite this article
  1. Mengjie Li
  2. Aiguo Tian
  3. Jin Jiang
(2025)
Numb provides a fail-safe mechanism for intestinal stem cell self-renewal in adult Drosophila midgut
eLife 14:RP104723.
https://doi.org/10.7554/eLife.104723.3
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7 figures, 1 table and 1 additional file

Figures

Figure 1
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BMP signaling inhibits Dl-independent N pathway activity to promote intestinal stem cell (ISC) self-renewal.

(A) A scheme for the ISC lineage in Drosophila midgut. (B-E’’) Representative images of Control guts (B–B’’), midguts expressing UAS-Put-RNAi (C–C’’), UAS-Dl-RNAi (D–D’’), or UAS-Put-RNAi+UAS-Dl-RNAi…

Figure 1—source data 1

Source data for the quantification in Figure 1.

https://cdn.elifesciences.org/articles/104723/elife-104723-fig1-data1-v1.xlsx
Figure 2
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Numb is important for intestinal stem cell (ISC) maintenance when BMP pathway activity is attenuated.

(A-D’’) Representative images of adult midguts expressing UAS-mCherry-RNAi (Control) (A–A’’), UAS-Mad-RNAi (B–B’’), UAS-Numb-RNAi (C–C’’), and UAS-Mad-RNAi+UAS-Numb-RNAi (D–D’’) with esg-Gal4ts, UAS-…

Figure 2—source data 1

Source data for the quantification in Figure 2E–G.

https://cdn.elifesciences.org/articles/104723/elife-104723-fig2-data1-v1.xlsx
Figure 3
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numb and mad double mutations resulted in loss of intestinal stem cell (ISC).

(A-H’) Representative images of adult midguts containing MARCM clones (green) of FRT40 (Control) (A, A’, E, E’), mad1-2 (B, B’, F, F’), numb4 (C, C’, G, G’), and mad1-2, numb4 (D, D’, H, H’) and …

Figure 3—source data 1

Source data for the quantification in Figure 3L.

https://cdn.elifesciences.org/articles/104723/elife-104723-fig3-data1-v1.xlsx
Figure 4
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Depletion of both Numb and Mad leads to more symmetric intestinal stem cell (ISC) divisions that produce two enteroblasts (EBs).

(A) Scheme of an ISC division that produces differentially labeled daughter cells (RFP+ GFP− and RFP− GFP+) through FRT-mediated mitotic recombination. Adapted from Tian and Jiang, 2014. (B) Scheme …

Figure 4—source data 1

Source data for the quantification in Figure 4P.

https://cdn.elifesciences.org/articles/104723/elife-104723-fig4-data1-v1.xlsx
Figure 5 with 1 supplement
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numb mutant clones exhibit weak stem cell loss phenotype.

(A-C’) Representative images of adult midguts containing MARCM clone (green) of FRT40 (Control) (A, A’), numb4 (B, B’), and numb15 (C, C’) and immunostained for Dl (red), GFP (green), and DAPI …

Figure 5—source data 1

Source data for the quantification in Figure 5E.

https://cdn.elifesciences.org/articles/104723/elife-104723-fig5-data1-v1.xlsx
Figure 5—figure supplement 1
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Numb is required for enteroendocrine (EE) fate determination.

(A-C’) MARCM clone (green) of control (A, A’), numb4 (B, B’), and numb15 (C, C’) are stained for Pros (red) at 14 days after clone induction. Representative clone in control guts (A, A’) contains EE …

Figure 6
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Numb is critical for intestinal stem cell (ISC) maintenance during regeneration.

(A–I’) Adult flies of indicated genotype were treated with sucrose, bleomycin, or dextran sodium sulfate (DSS) for 24 hr at 14 days after clone induction and recovered for another 4 days before …

Figure 6—source data 1

Source data for the quantification in Figure 6J.

https://cdn.elifesciences.org/articles/104723/elife-104723-fig6-data1-v1.xlsx
Figure 7
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Model for Numb and BMP signaling in intestinal stem cell (ISC)/enteroblast (EB) fate decision.

(A) During asymmetric ISC division, the basal ISC daughter transduces higher level of BMP signaling and inherits higher level of Numb activity than the apical one. Inhibition of N by BMP signaling …

Tables

Key resources table
Reagent type (species) or resourceDesignationSource or referenceIdentifiersAdditional information
Genetic reagent (D. melanogaster)esg-Gal4Jiang et al., 2009FLYB: FBti0013268FlyBase symbol: P{GawB}NP5130
Genetic reagent (D. melanogaster)Dl-lacZZeng et al., 2010FLYB: FBti0004778FlyBase symbol: P{PZ}Delta[05151]
Genetic reagent (D. melanogaster)Su(H)-lacZZeng et al., 2010FLYB: FBtp0014034FlyBase symbol: P{Ddc.E(spl)m8-HLH-lacZ.Gbe}
Genetic reagent (D. melanogaster)E(spl)mβCD2Bloomington Drosophila Stock CenterBDSC 83353
FLYB: FBst0083353
RRID:BDSC_83353		FlyBase symbol: w[*]; l(2)*[*]/CyO, P{ry[+t7.2]=en1}wg[en11]; P{w[+mC]=E(spl)mbeta-HLH-CD2.dC}T6
Genetic reagent (D. melanogaster)Tub-Gal80tsJiang et al., 2009FLYB: FBti0027796FlyBase symbol: P{tubP-GAL80ts}
Genetic reagent (D. melanogaster)UAS-Put RNAiVienna Drosophila Resource CenterVDRC 107071
FLYB: FBst0478894
RRID:Flybase_FBst0473060		FlyBase symbol: P{KK102676}VIE-260B
Genetic reagent (D. melanogaster)UAS-Dl RNAiBloomington Drosophila Stock CenterBDSC 28032
FLYB: FBst0028032
RRID:BDSC_28032		FlyBase symbol: y (1) v(1); P{TRiP.JF02867}attP2
Genetic reagent (D. melanogaster)UAS-Mad RNAiVienna Drosophila Resource CenterVDRC 12635
FLYB:
FBst0450590
RRID:Flybase_FBst0450590		FlyBase symbol: w1118; P{GD4121}v12635
Genetic reagent (D. melanogaster)UAS-Numb RANiBloomington Drosophila Stock CenterBDSC 35045
FLYB: FBst0035045 RRID:BDSC_35045		FlyBase symbol: y(1) sc[*] v(1) sev(21); P{y[+t7.7] v[+t1.8]=TRiP.HMS01459}attP2
Genetic reagent (D. melanogaster)UAS-mCherry RANiBloomington Drosophila Stock CenterBDSC 35785
FLYB: FBst0035785
RRID:BDSC_35785		FlyBase symbol: y(1) sc[*] v(1) sev(21); P{y[+t7.7] v[+t1.8]=VALIUM20-mCherry.RNAi}attP2
Genetic reagent (D. melanogaster)numb4Skeath and Doe, 1998FLYB: FBal0090215FlyBase symbol: numb(4)
Genetic reagent (D. melanogaster)numb15Sallé et al., 2017FLYB: FBal0146969FlyBase symbol: numb(15)
Genetic reagent (D. melanogaster)mad1-2Bloomington Drosophila Stock CenterBDSC 7323
FLYB: FBst0007323
RRID:BDSC_7323		FlyBase symbol: w*; Mad(; 1-2) P{neoFRT}40A/CyO
Genetic reagent (D. melanogaster)FRT 40ABloomington Drosophila Stock CenterBDSC 8212
FLYB: FBst0008212
RRID:BDSC_8212		FlyBase symbol: w[1118]; P{ry[+t7.2]=neoFRT}40A/CyO; P{ry[+t7.2]=ey-FLP.N}6, ry[506]
Genetic reagent (D. melanogaster)yw, hs-FLPBloomington Drosophila Stock CenterBDSC 1929
FLYB: FBst0001929
RRID:BDSC_1929		FlyBase symbol: P{ry[+t7.2]=hsFLP}12, y(1) w[*]; sna[Sco]/CyO
Genetic reagent (D. melanogaster)FRT 40A, tub-GAL80Bloomington Drosophila Stock CenterBDSC 5192
FLYB: FBst0005192
RRID:BDSC_5192		FlyBase symbol: y(1) w[*]; P{w[+mC]=tubP-GAL80}LL10 P{ry[+t7.2]=neoFRT}40A/CyO
Genetic reagent (D. melanogaster)UAS-GFPBloomington Drosophila Stock CenterBDSC 5130
FLYB: FBst0005130
RRID:BDSC_5130		FlyBase symbol: y(1) w[*]; betaTub60D[Pin-Yt]/CyO; P{w[+mC]=UAS-mCD8::GFP.L}LL6
Genetic reagent (D. melanogaster)FRT 19ABloomington Drosophila Stock CenterBDSC 1709
FLYB: FBst0001709
RRID:BDSC_1709		FlyBase symbol: P{ry[+t7.2]=neoFRT}19A; ry[506]
Genetic reagent (D. melanogaster)Ubi-GFPnlsChen and Schüpbach, 2006FLYB: FBti0015575FlyBase symbol: P{Ubi-GFP(S65T)nls}X
Genetic reagent (D. melanogaster)FRT 19A, ubi-mRFPnlsBloomington Drosophila Stock CenterBDSC 31418
FLYB: FBst0031418
RRID:BDSC_31418		FlyBase symbol: P{w[+mC]=Ubi-mRFP.nls}1, w[*], P{ry[+t7.2]=hsFLP}12P{ry[+t7.2]=neoFRT}19A
Antibodyanti-GFP (chicken polyclonal)AbcamCat#: ab13970; RRID:AB_300798IF (1:1000)
Antibodyanti-β-Galactosidase (rabbit polyclonal)MP BiomedicalsCat#: 08559761
RRID:AB_3675281		IF (1:1000)
Antibodyanti-Rat CD2 (mouse monoclonal)Bio-RadCat#: MCA154GA
RRID:AB_566608		IF (1:2000) (Formerly AbD Serotec)
Antibodyanti-Dl extracellular domain (mouse monoclonal)DSHBCat#: c594.9b
RRID:AB_528194		IF (1:20)
Antibodyanti-Prospero (mouse monoclonal)DSHBCat#: Prospero
RRID:AB_528440		IF (1:20)
Antibodyanti-Pdm1
(rabbit polyclonal)		Dr. Xiaohang YangIF (1:1000)
Antibodyanti-Chicken Alexa Fluor 488 (goat polyclonal secondary)Thermo Fisher ScientificCat#: A-11039
RRID:AB_2534096		IF (1:1000)
Antibodyanti-Mouse Alexa Flour 546 (goat polyclonal secondary)Thermo Fisher ScientificCat#: A-11030
RRID:AB_2737024		IF (1:1000)
Antibodyanti-Rabbit Alexa Flour 546 (goat polyclonal secondary)Thermo Fisher ScientificCat#: A-11035
RRID:AB_2534093		IF (1:1000)
Antibodyanti-Mouse Cy5 (goat polyclonal)Jackson ImmunoResearch LabsCat#: 115-175-166
RRID:AB_2338714		IF (1:500)
Antibodyanti-Rabbit Cy5 (goat polyclonal)Jackson ImmunoResearch LabsCat#: 111-175-144
RRID:AB_2338013		IF (1:500)
Chemical compound, drugDAPI (4′,6-diamidino-2-phenylindole, dihydrochloride)InvitrogenCat#: D1306IF (1:2000)
Chemical compound, drugDSS (dextran sulfate sodium)Sigma-AldrichCat#: 428675% solution
Chemical compound, drugBleomycin sulfate from Streptomyces verticillusSigma-AldrichCat#: B841625 μg/ml
Chemical compound, drugSucroseFisher BioreagentCat#: BP220-2125% solution

Additional files

MDAR checklist
https://cdn.elifesciences.org/articles/104723/elife-104723-mdarchecklist1-v1.docx

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