Glial ferritin maintains neural stem cells via transporting iron required for self-renewal in Drosophila
- School of Life Science and Technology, Department of Neurosurgery, Zhongda Hospital, The Key Laboratory of Developmental Genes and Human Disease, Ministry of Education, Southeast University, China
- Co-innovation Center of Neuroregeneration, Nantong University, China
Figures
Figure 1 with 3 supplements
Ferritin in the glia is required for neuroblast (NB) maintenance and proliferation.
(A) NB number and proliferation rate after Fer1HCH or Fer2LCH RNAi in glia. (B) Quantification of NB number in central brain (CB) (A). (C) Quantification of NB proliferation rate in CB (A). (D and E)…
Figure 1—figure supplement 1
Glial ferritin knockdown leads to low neuroblast (NB) proliferation and number.
(A) Quantification of NB number in ventral nerve cord (VNC). (B) Quantification of NB proliferation rate in VNC. (C) 5-ethynyl-2’-deoxyuridine (EdU) incorporation in central nervous system (CNS) …
Figure 1—figure supplement 2
Ferritin knockdown in different glial subpopulations did not induce any discernible phenotype.
(A) Knocking down ferritin genes in subperineurial glia, perineurial glia, ensheathing glia, and astrocyte-like glia, respectively.
Figure 1—figure supplement 3
Verify the phenotype induced by glial ferritin knockdown using different manipulations.
(A) Neuroblast (NB) number and proliferation in other RNAi lines targeting on ferritin. (B) Rescue of NB number in ventral nerve cord (VNC) when simultaneously overexpressed RNAi-resistant Fer1HCH …
Figure 2 with 1 supplement
Ferritin is produced mainly in glial cells in the Drosophila central nervous system (CNS) and secreted into neuroblasts (NBs) through a vesicle-dependent pathway.
(A) The pattern of DsRed Stinger driven by Fer1HCH/Fer2LCH-GAL4 in CNS. (A’) The magnification of (A). (B) The distribution of ferritin labeled by Fer1HCHG188 in CNS of control and glial ferritin …
Figure 2—figure supplement 1
Fer2LCH-GAL4 was used to validate the pattern of Fer1HCH/Fer2LCH-GAL4.
(A) Most DsRed driven by Fer2LCH-GAL4 was colocalized with glial marker Repo.
Figure 3 with 3 supplements
Glial ferritin defects lead to iron deficiency in neuroblasts (NBs).
(A) The conditions of brain development in Drosophila with feeding iron chelator bathophenanthrolinedisulfonic acid disodium (BPS) or supplementary iron ferric ammonium citrate (FAC). (B) Iron …
Figure 3—figure supplement 1
Brain defects induced by glial ferritin knockdown were caused by iron deficiency.
(A) Test the feeding behavior by adding Brilliant Blue to normal food or bathophenanthrolinedisulfonic acid disodium (BPS) food. (B) Iron deficiency by blocking iron absorption in gut also induced …
Figure 3—figure supplement 2
Reactive oxygen species (ROS) accumulates in central nervous system (CNS) after glial ferritin knockdown, but inhibiting ROS cannot restore neuroblast (NB) number and proliferation.
(A) ROS level marked by Gstd-GFP after knocking down ferritin in glia. (B) Quantification of ROS level in (A). (C) Overexpress antioxidant sod1 or cat in glial cells. (D) Overexpress antioxidant …
Figure 3—figure supplement 3
Glial ferritin knockdown does not induce apoptosis in the central nervous system (CNS).
(A) Apoptotic signal labeled by Caspase-3 of glial ferritin knockdown. (B) Quantification of Caspase-3 negative neuroblast (NB) ratio. (C) TUNEL staining of Fer1HCH RNAi driven by repo-GAL4. (D) …
Figure 4 with 1 supplement
Glial ferritin defects result in impaired iron-sulfur (Fe-S) cluster activity and ATP production.
(A) The determination of cytosolic aconitase activity. (B) Knockdown of mitochondrial iron-sulfur protein Nfs1 and cytosolic iron-sulfur protein assembly (CIA) complex in neuroblasts (NBs). (C) …
Figure 4—figure supplement 1
Validation of RNA-seq data by qRT-PCR using sorted neuroblast (NB) lineages and Ndi1 overexpression.
(A) qRT-PCR analysis of ND-15 and CG15715 to validate the enrichment result. (B) Ndi1 overexpression could restore ATP production. (A and B, n=3; Statistical results were presented as means ± SD, p …
Figure 5 with 3 supplements
Glial ferritin defects lead to the premature differentiation of neuroblasts (NBs).
(A) Temporal window of glial ferritin maintaining NBs. The temperature shift of flies with glial ferritin knockdown from the permissive temperature (18℃) to the restrictive temperature (29℃). (B) …
Figure 5—figure supplement 1
Neuroblast (NB) loss upon glial ferritin knockdown is not due to NB origin and apoptosis.
(A) Temporal window of glial ferritin maintaining NBs. The temperature shift of flies with glial ferritin knockdown from the restrictive temperature (29℃) to the permissive temperature (18℃). (B) …
Figure 5—figure supplement 2
Ndi1 overexpression can restore Pros localization.
(A) Pros staining of Ndi1 overexpression upon glial ferritin knockdown. (B) Quantification of the ratio of Pros+ Dpn+ NBs to Dpn+ NBs in (A). (B, n=6, 9; Statistical results were presented as means …
Figure 5—figure supplement 3
Glial ferritin defects do not affect neuroblast (NB) reactivation and asymmetric division.
(A) aPKC localization in NB after knocking down Fer2LCH. (B) NB size upon glial ferritin knockdown was not changed. (C) Activation of Insulin signaling could not restore NB number and proliferation. …
Figure 6 with 1 supplement
Ferritin functions as a potential target for tumor suppression.
(A) Knockdown of ferritin in glia could inhibit the tumor induced by numb RNAi. (B) Quantification of larval brain size in (A). (C) Iron chelator bathophenanthrolinedisulfonic acid disodium (BPS) …
Figure 6—figure supplement 1
Iron chelator inhibits tumor progression in Drosophila and mice.
(A) Knockdown of ferritin in glia could inhibit the tumor induced by brat RNAi. (B) Quantification of larval brain size in (A). (C) Iron chelator bathophenanthrolinedisulfonic acid disodium (BPS) …
Figure 7
The level of ferritin is controlled by neuroblast (NB) proliferation.
(A) Fer1HCH subunit level in Drosophila central nervous system (CNS) after manipulating proliferation. (B) Fer2LCH subunit level in Drosophila CNS after manipulating proliferation. (C) …
Author response image 1
The schematic diagram of relationship between energy and NB function in different groups.
“T” represents total energy for NB maintenance and proliferation. “N” represents the energy for NB maintenance. “P” represents the energy for NB proliferation. T=N+P
Author response image 2
The bar graph of survival time of mice treated with DFP.
(The unpaired two-sided Student’s t test was employed to assess statistical significance. Statistical results were presented as means ± SD. n=7,6; *: p<0.05)
Tables
Table 1
The phenotype of ferritin knockdown in different glial subpopulations.
BDSC: Bloomington Drosophila Stock Center; DGRC: Drosophila Genetic Resource Center; +++: strong phenotype; ++: weak phenotype; -: no phenotype.
| GAL4 driver | Stock center | Stock number | Expression pattern | Fer1HCH RNAi | Fer2LCH RNAi |
|---|---|---|---|---|---|
| repo-GAL4 | BDSC | 7415 | all glia | +++ | +++ |
| nrv2-GAL4 | BDSC | 6799 | cortex glia | + | + |
| mdr65-GAL4 | BDSC | 50472 | subperineurial glia | - | - |
| moody-GAL4 | BDSC | 90883 | subperineurial glia | - | - |
| NP6293-GAL4 | DGRC | 105188 | perineurial glia | - | - |
| alrm-GAL4 | BDSC | 67032 | astrocyte-like glia | - | - |
| NP6520-GAL4 | DGRC | 105240 | ensheathing glia | - | - |
Additional files
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Supplementary file 1
- https://cdn.elifesciences.org/articles/93604/elife-93604-supp1-v1.xlsx
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Supplementary file 2
The phenotype of iron-related genes driven by repo-GAL4 or Insc-GAL4.
- https://cdn.elifesciences.org/articles/93604/elife-93604-supp2-v1.xlsx
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MDAR checklist
- https://cdn.elifesciences.org/articles/93604/elife-93604-mdarchecklist1-v1.pdf
