Heterogeneous levels of delta-like 4 within a multinucleated niche cell maintains muscle stem cell diversity
- The Eli and Edythe Broad Center for Regenerative Medicine and Stem Cell Research, Department of Orthopedic Surgery, University of California San Francisco, United States
- Department of Biomedical Sciences, University of North Dakota School of Medicine and Health Sciences, United States
- Departments of Surgery and Orofacial Sciences, Program in Craniofacial Biology, University of California San Francisco, United States
Figures
Figure 1 with 1 supplement
Adult quiescent muscle stem cells (QSCs) exist in diverse cell states.
(A) Representative images of a high and low Pax7+ stem cell (SC) on a freshly isolated muscle fiber that has corresponding high and low levels of Ddx6 protein. (B) A density map of Pax7 intensity in …
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Figure 1—source data 1
Adult QSCs exist in diverse cell states.
- https://cdn.elifesciences.org/articles/68180/elife-68180-fig1-data1-v1.xlsx
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Figure 1—source data 2
Adult QSCs exist in diverse cell states.
- https://cdn.elifesciences.org/articles/68180/elife-68180-fig1-data2-v1.xlsx
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Figure 1—source data 3
Adult QSCs exist in diverse cell states.
- https://cdn.elifesciences.org/articles/68180/elife-68180-fig1-data3-v1.xlsx
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Figure 1—source data 4
Adult QSCs exist in diverse cell states.
- https://cdn.elifesciences.org/articles/68180/elife-68180-fig1-data4-v1.xlsx
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Figure 1—source data 5
Adult QSCs exist in diverse cell states.
- https://cdn.elifesciences.org/articles/68180/elife-68180-fig1-data5-v1.xlsx
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Figure 1—source data 6
Adult QSCs exist in diverse cell states.
- https://cdn.elifesciences.org/articles/68180/elife-68180-fig1-data6-v1.xlsx
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Figure 1—source data 7
Adult QSCs exist in diverse cell states.
- https://cdn.elifesciences.org/articles/68180/elife-68180-fig1-data7-v1.xlsx
Figure 1—figure supplement 1
Adult quiescent muscle stem cells (QSCs) exist in a continuum of cell states that give rise to a continuum of fates when activated.
(A, B) Fluorescence-assisted cell sorting (FACS) strategy of obtaining GFP+ stem cells (SCs) from a Pax7-nGFP transgenic reporter line (in A) and isolation of green fluorescent protein (GFP) low, …
Figure 2 with 2 supplements
Heterogeneous expression of Dll4 on the muscle fiber correlates with stem cell (SC) diversity.
(A) Representative images of regions of muscle fiber expressing heterogeneous levels of Dll4 protein, which corresponds to the amount of Dll4 captured by the adjacent SC and the expression of Pax7. …
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Figure 2—source data 1
Heterogeneous expression of Dll4 on the muscle fiber correlates with SC diversity.
- https://cdn.elifesciences.org/articles/68180/elife-68180-fig2-data1-v1.xlsx
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Figure 2—source data 2
Heterogeneous expression of Dll4 on the muscle fiber correlates with SC diversity.
- https://cdn.elifesciences.org/articles/68180/elife-68180-fig2-data2-v1.xlsx
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Figure 2—source data 3
Heterogeneous expression of Dll4 on the muscle fiber correlates with SC diversity.
- https://cdn.elifesciences.org/articles/68180/elife-68180-fig2-data3-v1.xlsx
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Figure 2—source data 4
Heterogeneous expression of Dll4 on the muscle fiber correlates with SC diversity.
- https://cdn.elifesciences.org/articles/68180/elife-68180-fig2-data4-v1.xlsx
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Figure 2—source data 5
Heterogeneous expression of Dll4 on the muscle fiber correlates with SC diversity.
- https://cdn.elifesciences.org/articles/68180/elife-68180-fig2-data5-v1.xlsx
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Figure 2—source data 6
Heterogeneous expression of Dll4 on the muscle fiber correlates with SC diversity.
- https://cdn.elifesciences.org/articles/68180/elife-68180-fig2-data6-v1.xlsx
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Figure 2—source data 7
Heterogeneous expression of Dll4 on the muscle fiber correlates with SC diversity.
- https://cdn.elifesciences.org/articles/68180/elife-68180-fig2-data7-v1.xlsx
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Figure 2—source data 8
Heterogeneous expression of Dll4 on the muscle fiber correlates with SC diversity.
- https://cdn.elifesciences.org/articles/68180/elife-68180-fig2-data8-v1.xlsx
Figure 2—figure supplement 1
Dll4 transcripts are highly expressed in adult muscle fibers.
(A) Heatmap of Notch ligands from microarray of muscle fibers isolated from postnatal day 3 (p3), day 7 (p7), and adult Extensor Digitorum Longus (EDL) muscle (n = 2), The expression values are log2 …
Figure 2—figure supplement 2
Spatial distribution of Dll4 along muscle fibers does not map to known anatomical locations.
(A) Low magnification Spinning Disk images of WT fibers showing heterogeneous Dll4 expression and the adjacent stem cell (SC) expressing variable Pax7 expression. (B–U) Each graph represents the …
Figure 3 with 2 supplements
Muscle fiber-derived Dll4 maintains a continuum of diverse states in the quiescent muscle stem cell (QSC) pool.
(A) Schematic representation of the experimental design. (B–F) Representative images of Dll4 and Pax7 expression in Control and MF-Dll4fl/fl fibers and stem cells (SCs) (in B), quantification of …
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Figure 3—source data 1
Muscle fiber derived Dll4 maintains a continuum of diverse states in the QSC pool.
- https://cdn.elifesciences.org/articles/68180/elife-68180-fig3-data1-v1.xlsx
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Figure 3—source data 2
Muscle fiber derived Dll4 maintains a continuum of diverse states in the QSC pool.
- https://cdn.elifesciences.org/articles/68180/elife-68180-fig3-data2-v1.xlsx
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Figure 3—source data 3
Muscle fiber derived Dll4 maintains a continuum of diverse states in the QSC pool.
- https://cdn.elifesciences.org/articles/68180/elife-68180-fig3-data3-v1.xlsx
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Figure 3—source data 4
Muscle fiber derived Dll4 maintains a continuum of diverse states in the QSC pool.
- https://cdn.elifesciences.org/articles/68180/elife-68180-fig3-data4-v1.xlsx
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Figure 3—source data 5
Muscle fiber derived Dll4 maintains a continuum of diverse states in the QSC pool.
- https://cdn.elifesciences.org/articles/68180/elife-68180-fig3-data5-v1.xlsx
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Figure 3—source data 6
Muscle fiber derived Dll4 maintains a continuum of diverse states in the QSC pool.
- https://cdn.elifesciences.org/articles/68180/elife-68180-fig3-data6-v1.xlsx
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Figure 3—source data 7
Muscle fiber derived Dll4 maintains a continuum of diverse states in the QSC pool.
- https://cdn.elifesciences.org/articles/68180/elife-68180-fig3-data7-v1.xlsx
Figure 3—figure supplement 1
Deletion of Dll4 in the niche causes a reduction in the number of stem cell (SC).
(A) Representative images of Pax7+ SCs in tissue cross sections of Control and MF-Dll4fl/fl tibialis anterior (TA). (B) Quantification of the number of Pax7+ SCs in Control and MF-Dll4fl/fl fibers (n…
Figure 3—figure supplement 2
Dll4 does not have a cell autonomous role in quiescent muscle stem cells (QSCs).
(A) Schematic representation of the experimental design. (B) Quantification of the number of Pax7+ stem cells (SCs) in tibialis anterior (TA) muscle cross-sections of Control and SC-Dll4fl/fl fibers …
Figure 4
Reduction of Dll4 in the niche causes a shift in the continuum toward lower Pax7 levels.
(A–C) Quantification of the number of Pax7+ stem cells (SCs) (in A), Dll4 intensity in muscle fibers (in B), Pax7 intensity in SCs (in C) of Control mice compared to MF-Dll4fl/fl mice that were …
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Figure 4—source data 1
Reduction of Dll4 in the niche causes a shift in the continuum towards lower Pax7 levels.
- https://cdn.elifesciences.org/articles/68180/elife-68180-fig4-data1-v1.xlsx
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Figure 4—source data 2
Reduction of Dll4 in the niche causes a shift in the continuum towards lower Pax7 levels.
- https://cdn.elifesciences.org/articles/68180/elife-68180-fig4-data2-v1.xlsx
Figure 5 with 2 supplements
Niche-derived Dll4 constrains the proliferative and commitment potential of the quiescent muscle stem cell (QSC) pool.
(A) Schematic representation of the experimental design. (B) qRT-PCR for Pax7, MyoD, and Myogenin transcripts in freshly isolated stem cells (SCs) from Control and MF-Dll4fl/fl fibers (n = 2). (C) …
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Figure 5—source data 1
Niche derived Dll4 constrains the proliferative and commitment potential of the QSC pool.
- https://cdn.elifesciences.org/articles/68180/elife-68180-fig5-data1-v1.xlsx
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Figure 5—source data 2
Niche derived Dll4 constrains the proliferative and commitment potential of the QSC pool.
- https://cdn.elifesciences.org/articles/68180/elife-68180-fig5-data2-v1.xlsx
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Figure 5—source data 3
Niche derived Dll4 constrains the proliferative and commitment potential of the QSC pool.
- https://cdn.elifesciences.org/articles/68180/elife-68180-fig5-data3-v1.xlsx
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Figure 5—source data 4
Niche derived Dll4 constrains the proliferative and commitment potential of the QSC pool.
- https://cdn.elifesciences.org/articles/68180/elife-68180-fig5-data4-v1.xlsx
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Figure 5—source data 5
Niche derived Dll4 constrains the proliferative and commitment potential of the QSC pool.
- https://cdn.elifesciences.org/articles/68180/elife-68180-fig5-data5-v1.xlsx
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Figure 5—source data 6
Niche derived Dll4 constrains the proliferative and commitment potential of the QSC pool.
- https://cdn.elifesciences.org/articles/68180/elife-68180-fig5-data6-v1.xlsx
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Figure 5—source data 7
Niche derived Dll4 constrains the proliferative and commitment potential of the QSC pool.
- https://cdn.elifesciences.org/articles/68180/elife-68180-fig5-data7-v1.xlsx
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Figure 5—source data 8
Niche derived Dll4 constrains the proliferative and commitment potential of the QSC pool.
- https://cdn.elifesciences.org/articles/68180/elife-68180-fig5-data8-v1.xlsx
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Figure 5—source data 9
Niche derived Dll4 constrains the proliferative and commitment potential of the QSC pool.
- https://cdn.elifesciences.org/articles/68180/elife-68180-fig5-data9-v1.xlsx
Figure 5—figure supplement 1
Niche depletion of Dll4 causes stem cells (SCs) to proliferate faster in the presence of mitogen.
(A) Number of Pax7+ SCs at t0, t30, and t48 in cultured Control and MF-Dll4fl/fl fibers, depicted as a dot plot (each dot represents a single fiber) (n = 3 mice). (B) Control and Dll4 deleted fibers …
Figure 5—figure supplement 2
Impaired regenerative potential of stem cells (SCs) on Dll4-depleted niche.
(A) Schematic representation of the experimental design. (B) Mean cross-sectional area of Control and MF-Dll4fl/fl tibialis anterior (TA) fibers, injured and regenerated for 40 days (n = 3). Error …
Figure 6
Niche-derived Mib1 directs spatial patterning and activation of Dll4.
(A) Representative images of WT fibers showing regions of high and low Mib1 protein expression and the adjacent Pax7+ stem cell (SC). (B) A bivariate plot showing Mib1 intensity in the fiber and …
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Figure 6—source data 1
Niche-derived Mib1 directs spatial patterning and activation of Dll4.
- https://cdn.elifesciences.org/articles/68180/elife-68180-fig6-data1-v1.xlsx
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Figure 6—source data 2
Niche-derived Mib1 directs spatial patterning and activation of Dll4.
- https://cdn.elifesciences.org/articles/68180/elife-68180-fig6-data2-v1.xlsx
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Figure 6—source data 3
Niche-derived Mib1 directs spatial patterning and activation of Dll4.
- https://cdn.elifesciences.org/articles/68180/elife-68180-fig6-data3-v1.xlsx
Figure 7
Mib1 in the muscle fiber maintains quiescent muscle stem cells (QSCs) by inhibiting differentiation.
(A) Schematic representation of the experimental design. (B) qRT-PCR for mib1 transcripts in fibers of Control and MF-Mib1fl/fl normalized to GAPDH (n = 3). (C) Number of Pax7+ stem cells (SCs) in …
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Figure 7—source data 1
Niche-derived Mib1 directs spatial patterning and activation of Dll4.
- https://cdn.elifesciences.org/articles/68180/elife-68180-fig7-data1-v1.xlsx
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Figure 7—source data 2
Niche-derived Mib1 directs spatial patterning and activation of Dll4.
- https://cdn.elifesciences.org/articles/68180/elife-68180-fig7-data2-v1.xlsx
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Figure 7—source data 3
Niche-derived Mib1 directs spatial patterning and activation of Dll4.
- https://cdn.elifesciences.org/articles/68180/elife-68180-fig7-data3-v1.xlsx
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Figure 7—source data 4
Niche-derived Mib1 directs spatial patterning and activation of Dll4.
- https://cdn.elifesciences.org/articles/68180/elife-68180-fig7-data4-v1.xlsx
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Figure 7—source data 5
Niche-derived Mib1 directs spatial patterning and activation of Dll4.
- https://cdn.elifesciences.org/articles/68180/elife-68180-fig7-data5-v1.xlsx
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Figure 7—source data 6
Niche-derived Mib1 directs spatial patterning and activation of Dll4.
- https://cdn.elifesciences.org/articles/68180/elife-68180-fig7-data6-v1.xlsx
Figure 8
Model depicting heterogeneous levels of Dll4/Mib1 in the niche regulates a continuum of metastable stem cell (SC) states.
(A) Top: During tissue homeostasis, Notch ligand Dll4 is present in a heterogeneous manner along the length of the muscle fibers that maintains the size of the SC pool and a continuum of quiescent …
Additional files
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Supplementary file 1
Microarray expression of Notch ligands in p3, p7, and adult muscle fibers (log2 values).
This table includes the expression (as log2 values) of Notch ligands in postnatal day 3, postnatal day 7, and adult. Related to Figure 2—figure supplement 1.
- https://cdn.elifesciences.org/articles/68180/elife-68180-supp1-v1.docx
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Supplementary file 2
Primers used for quantitative RT-PCR.
This table includes the forward and reverse primer sequences of genes amplified by qRT-PCR. Related to Figure 2—figure supplement 1, Figures 5 and 7.
- https://cdn.elifesciences.org/articles/68180/elife-68180-supp2-v1.docx
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Transparent reporting form
- https://cdn.elifesciences.org/articles/68180/elife-68180-transrepform1-v1.pdf
