Stem cell topography splits growth and homeostatic functions in the fish gill
- Heidelberg University, Germany
- Royal Children’s Hospital, Australia
- Universidad Pablo de Olavide, Spain
- National Institute for Basic Biology, National Institutes of Natural Sciences, Japan
Figures
Figure 1
Growth and homeostasis in the medaka gill.
(A) Enucleated entire gills of medaka at different post-embryonic times show that organ size increases during post-embryonic growth (left). A gill contains four pairs of branchial arches (middle left…
Figure 2
Gill stem cells located at the periphery of branchial arches generate more filaments life-long.
(A) Scheme showing the expected outcome assuming a progenitor (left bottom) or a stem cell (right bottom) model. Please note that the schemes considered as ‘labelled’ any filament containing EGFP+ ce…
Figure 3 with 1 supplement
Filament growth stem cells are located at the apical tip.
(A) Scheme showing the expected outcome of IdU pulse and chase experiments depending on the location of growth stem cells. (B) IdU pulse and chase experiment shows the apical region devoted of …
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Figure 3—source data 1
Transcriptome of apical and medial domains in a gill filament.
- https://doi.org/10.7554/eLife.43747.006
Figure 3—figure supplement 1
Technical analysis of the RNA-seq data.
(A) Principal Component Analysis (PCA) plot of the three biological replicates of transcripts from the apical and the medial part of the medaka gill filaments. (B) Volcano-plot representation of …
Figure 4
Filament growth stem cells are fate restricted.
(A–B) A gill (A) and a branchial arch (B) from a double transgenic GaudíUbiq.iCre GaudíRSG fish two month after induction with TMX. (C–F) Confocal images from filaments in A, B, stained for EGFP and …
Figure 5 with 1 supplement
Branchial arch stem cells are fate restricted.
(A) Scheme showing the expected outcome assuming that br-archSCs are fate restricted (middle) or multi-potent (bottom). The recombination pattern of consecutive filaments would be identical if …
Figure 5—figure supplement 1
Early embryonic recombination and transplantations at blastula stage indicate fate-restricted stem cells in the fish gill.
(A) Consecutive filaments from a double transgenic GaudíHsp70A.Cre GaudíRSG fish 3 months after induction (heat-shock at stage 20). Filaments display recombination in specific patterns (pattern 2 in …
Figure 6 with 1 supplement
p53 Coordinates growth stem cells in a lineage-specific manner.
(A) Branchial arch of a p53-/- -to-WT chimera, where p53-/- mutant cells are labelled in green. Composite filaments display the proper length as compared to non-labelled neighbour filaments. (B) …
Figure 6—figure supplement 1
Branchial Arch of p53E241X mutant.
(A) DAPI staining on a branchial arch extracted from a p53 homozygous mutant showing the un-distinguishable organisation, length and size of filaments and lamellae than wild-type samples.
Figure 7
Homeostatic stem cells locate to the base of each lamella.
(A) DAPI image of peripheral filaments indicating the increasing number of lamellae per filament. (B) DAPI image of consecutive lamellae along a filament reveals that lamellae do not increase their …
Figure 8
The homeostatic domain sustains growth after filament ablation.
(A) Scheme of the ablation procedure. The growth domain and the upper part of the homeostatic domain are mechanically ablated. (B) DAPI image of control filaments shows an intact growth domain at …
Videos
Video 1
3D reconstruction of a pattern 3-labelled filament.
A middle section of filament in an adult GaudíUbiq.iCre GaudíRSG fish that was induced for recombination at late embryonic stages. The filament shows the lineage of a growth stem cell that labels …
Video 2
3D reconstruction of a pattern 4-labelled filament.
A middle section of filament in an adult GaudíUbiq.iCre GaudíRSG fish that was induced for recombination at late embryonic stages. The filament shows the lineage of a growth stem cell that labels …
Video 3
3D reconstruction of a pattern 2-labelled filament.
A middle section of filament in an adult GaudíUbiq.iCre GaudíRSG fish that was induced for recombination at late embryonic stages. The filament shows the lineage of a growth stem cell that labels …
Video 4
3D reconstruction of a pattern 2-labelled lamella.
A middle section of filament in an adult GaudíUbiq.iCre GaudíRSG fish that was induced for recombination at late embryonic stages. The filament shows the lineage of a homeostatic stem cell that …
Tables
Key resources table
| Reagent type (species) or resource | Designation | Source or reference | Identifiers | Additional information |
|---|---|---|---|---|
| Strain, strain background (Oryzias latipes) | Cab | wild type stock, southern population | ||
| Genetic reagent (O. latipes) | p53 E241X mutant | Taniguchi et al., 2006 | ||
| Genetic reagent (O. latipes) | GaudíUbiq.iCre | Centanin et al., 2014 | ||
| Genetic reagent (O. latipes) | GaudíHsp70.A | Centanin et al., 2014 | ||
| Genetic reagent (O. latipes) | GaudíRSG | Centanin et al., 2014 | ||
| Genetic reagent (O. latipes) | GaudiLoxPOUT | Centanin et al., 2014 | ||
| Genetic reagent (O. latipes) | GaudiBBW | Centanin et al., 2014 | ||
| Antibody | a-EGFP (Rabbit IgG polyclonal) | Invitrogen (now Thermo Fischer) | CAB4211; RRID: AB_10709851 | Dilution 1:750 |
| Antibody | a-EGFP (Chicken IgY polyclonal) | life technologies | A10262; RRID: AB_2534023 | Dilution 1:750 |
| Antibody | a-Na + K + ATP-ase (Rabbit monoclonal) | Abcam | ab76020, EP1845Y | Dilution 1:200 |
| Antibody | a-BrdU/IdU (Mouse IgG monoclonal) | Becton Dickinson | 347580 | Dilution 1:50 |
| Antibody | Alexa 488 Goat a-Rabbit | Invitrogen (now Thermo Fischer) | A-11034 | Dilution 1:500 |
| Antibody | Alexa 488 Donkey a-Chicken | Invitrogen (now Thermo Fischer) | 703-545-155 | Dilution 1:500 |
| Antibody | Alexa 647 Goat a-Rabbit | Life Technologies | A-21245 | Dilution 1:500 |
| Antibody | Alexa 647 Goat a-Rabbit | Life Technologies | A-21245 | Dilution 1:500 |
| Antibody | Cy5 Donkey a-Mouse | Jackson | 715-175-151 | Dilution 1:500 |
| Chemical compound, drug | tamoxifen | Sigma | T5648 | |
| Chemical compound, drug | tricaine | Sigma-Aldrich | A5040-25G | |
| Chemical compound, drug | BrdU | Sigma-Aldrich | B5002 | final concentration of 0,4 g/l |
| Chemical compound, drug | IdU | Sigma-Aldrich | I7125 | final concentration of 0,4 g/l |
| Chemical compound, drug | Trizol | |||
| Software, algorithm | Ensemble | Public | ||
| Other | DAPI | Roth | final concentration of 5 ug/l | |
| Software, algorithm | DAVID 6.8 | https://david.ncifcrf.gov/home.jsp |
Additional files
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Supplementary file 1
Simulation of recombined pattern in branchial arches assuming progenitor cells.
The table represents simulated values of recombination (0 = unlabelled; 1 = labelled) in five gills, according to the ‘progenitor’ model described in M and M. The labelling efficiency was calculated from real data from 5 GaudíUbiq.iCre GaudíRSG recombined gills.
- https://doi.org/10.7554/eLife.43747.018
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Supplementary file 2
Simulation of recombined pattern in branchial arches assuming stem cells.
The table represents simulated values of recombination (0 = unlabelled; 1 = labelled) in five gills, according to the ‘stem cell’ model described in M and M. The labelling efficiency was calculated from real data from 5 GaudíUbiq.iCre GaudíRSG recombined gills.
- https://doi.org/10.7554/eLife.43747.019
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Supplementary file 3
Experimental data of recombined pattern in adult branchial arches.
The table includes values for recombination (0 = unlabelled; 1 = labelled) in five gills enucleated from adult GaudíUbiq.iCre GaudíRSG fish that were induced for recombination at late embryonic stages.
- https://doi.org/10.7554/eLife.43747.020
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Supplementary file 4
Objective function representation comparing data to progenitor and stem cell models.
The table displays the values for an objective function, comparing the recombination pattern obtained in 22 GaudíUbiq.iCre GaudíRSG recombined gills with the predicted values for a stem cell or a progenitor model (See M and M). Lower values for the objective function represent a better fit between experimental and simulated data.
- https://doi.org/10.7554/eLife.43747.021
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Supplementary file 5
Enriched transcripts in the apical and medial domains of gill filaments.
Representation of transcripts obtained from the apical and the medial domain of gill filaments, ordered according to their differential expression.
- https://doi.org/10.7554/eLife.43747.022
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Supplementary file 6
Experimental data of recombined patterns in adult branchial arches with cellular resolution.
The table displays values of recombination (0 = unlabelled; 1 = Pattern 1; 2 = Pattern 2; 3 = Pattern 3; 4 = Pattern 4) in gills enucleated from adult GaudíUbiq.iCre GaudíRSG fish that were induced for recombination at late embryonic stages.
- https://doi.org/10.7554/eLife.43747.023
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Transparent reporting form
- https://doi.org/10.7554/eLife.43747.024
