Summary of Study ST003712
This data is available at the NIH Common Fund's National Metabolomics Data Repository (NMDR) website, the Metabolomics Workbench, https://www.metabolomicsworkbench.org, where it has been assigned Project ID PR002305. The data can be accessed directly via it's Project DOI: 10.21228/M8PG2P This work is supported by NIH grant, U2C- DK119886.
See: https://www.metabolomicsworkbench.org/about/howtocite.php
This study contains a large results data set and is not available in the mwTab file. It is only available for download via FTP as data file(s) here.
| Study ID | ST003712 |
| Study Title | ndufs2-/- mitochondrial Leigh syndrome zebrafish model has shortened lifespan, morphologic anomalies, and altered one-carbon metabolism |
| Study Summary | Mitochondrial complex I (CI) deficiency is a common biochemical pathophysiology underlying Leigh syndrome spectrum (LSS), which manifests with progressive multi-system dysfunction, lactic acidemia, and early mortality. To facilitate future high-throughput screening of therapeutic candidates for CI deficient LSS, we used CRISPR/Cas9 to generate an ndufs2-/- 16 bp deletion zebrafish strain. ndufs2-/- larvae exhibit markedly reduced survival, severe neuromuscular dysfunction including impaired swimming capacity, multiple morphologic malformations, reduced growth, hepatomegaly, uninflated swim bladder, yolk retention, small intestines, and small eyes and pupils with abnormal retinal ganglion cells. Transcriptome profiling of ndufs2-/- larvae revealed dysregulation of the electron transport chain, the TCA cycle, fatty acid beta-oxidation, and one-carbon metabolism. Similar transcriptomic profiles were observed in ndufs2-/- missense mutant C. elegans (gas-1(fc21)) and two human CI-disease fibroblast cell lines stressed in galactose media. ndufs2-/- zebrafish had 80% reduced CI enzyme activity, and unbiased metabolomic profiling showed increased lactate, TCA cycle intermediates, and acyl-carnitine species. One-carbon metabolism associated pathway alterations appear to contribute to cellular pathophysiology, where folic acid treatment rescued the growth defect and hepatomegaly in ndufs2-/- larvae. This ndufs2-/- zebrafish knockout model exhibits severe CI deficiency and recapitulates central LSS phenotypes enabling future studies of CI disease mechanisms and therapeutic candidates. |
| Institute | Children's Hospital of Philadelphia |
| Last Name | Falk |
| First Name | Marni |
| Address | 3615 Civic Center Blvd, Philadelphia PA, 19104 |
| falkm@chop.edu | |
| Phone | 267-426-4961 |
| Submit Date | 2025-02-05 |
| Raw Data Available | Yes |
| Raw Data File Type(s) | mzML, raw(Thermo) |
| Analysis Type Detail | LC-MS |
| Release Date | 2025-08-01 |
| Release Version | 1 |
Select appropriate tab below to view additional metadata details:
Project:
| Project ID: | PR002305 |
| Project DOI: | doi: 10.21228/M8PG2P |
| Project Title: | ndufs2-/- mitochondrial Leigh syndrome zebrafish model has shortened lifespan, morphologic anomalies, and altered one-carbon metabolism |
| Project Summary: | Targeted metabolomics (LC-MS/MS) on zebrafish larvae samples from a novel human Primary Mitochondrial Disease (PMD) Complex I (CI) deficiency model generated by introducing a deletion to the NDUFS2 gene using CRISPR/Cas9 technology. Healthy zebrafish larvae were to test for differences in metabolite concentrations between healthy and diseased larvae. |
| Institute: | Children's Hospital of Philadelphia |
| Laboratory: | Mitochondrial Medicine Research Group |
| Last Name: | Falk |
| First Name: | Marni |
| Address: | 3615 Civic Center Blvd, Philadephia PA, 19104 |
| Email: | falkm@chop.edu |
| Phone: | 267-426-4961 |
Subject:
| Subject ID: | SU003844 |
| Subject Type: | Fish |
| Subject Species: | Danio rerio |
| Taxonomy ID: | 7955 |
Factors:
Subject type: Fish; Subject species: Danio rerio (Factor headings shown in green)
| mb_sample_id | local_sample_id | Genotype |
|---|---|---|
| SA406608 | N1 | Mutant (ndufs2-/-) |
| SA406609 | N2 | Mutant (ndufs2-/-) |
| SA406610 | N3 | Mutant (ndufs2-/-) |
| SA406611 | A1 | Wild-type |
| SA406612 | A2 | Wild-type |
| SA406613 | A3 | Wild-type |
| Showing results 1 to 6 of 6 |
Collection:
| Collection ID: | CO003837 |
| Collection Summary: | Twenty zebrafish larvae at 7 days post fertilization (dpf) were pooled and flash frozen per sample, with three samples per condition. 300 µL of ice cold 80:20 methanol:water was added to the frozen fish, then tissue was homogenized with a motorized pestle. |
| Sample Type: | Fish larvae |
Treatment:
| Treatment ID: | TR003853 |
| Treatment Summary: | No additional treatment. This is a genotype study. The cri4-ndufs2-/- mutant line was generated at the Children’s Hospital of Philadelphia (CHOP) Zebrafish Aquatics Core using CRISPR/Cas9 gene editing. A sgRNA targeting ndufs2 (gttcttccagatacgattatTGG) was injected into AB WT embryos at the 1-cell stage and animals were grown to adulthood to generate the F0 generation. Offspring of the F0 generation were confirmed by polymerase chain reaction (PCR) and Sanger sequencing. A 16 bp deletion was confirmed (AATCGTATCTGGAAGA) in exon 8 of ndufs2 resulting in a frameshift mutation starting at amino acid 286 and a premature stop codon at amino acid 313. As this truncation eliminates the 4Fe-4S binding site (AAs 326-347), it was anticipated to disrupt the activity of CI. |
Sample Preparation:
| Sampleprep ID: | SP003850 |
| Sampleprep Summary: | The sample underwent three freeze-thaw cycles between liquid nitrogen and 37oC. The sample was vortexed for 1 min and centrifuged at 20,160×g for 15 min at 4oC. Protein quantification was performed on the supernatant using the Pierce BCA kit. A volume equivalent to 10 µg of protein was transferred to a new tube and dried in a SpeedVac. The sample was resuspended in 100 µL 80:20 acetonitrile:water. The sample was again vortexed for 1 min and centrifuged at 20,160×g for 15 min at 4oC. The supernatant was transferred to an LC/MS vial. |
Chromatography:
| Chromatography ID: | CH004624 |
| Instrument Name: | Shimadzu Nexera X2 |
| Column Name: | SeQuant ZIC-pHILIC (150 x 2.1mm,5um) |
| Column Temperature: | 25 |
| Flow Gradient: | 0 to 15 min linear ramp from 90% B to 30% B; 15 to 18 min isocratic flow of 30% B; 18 to 19 min linear ramp from 30% B to 90% B; 19 to 27 min column regeneration with isocratic flow of 90% B |
| Flow Rate: | 0.2mL/min |
| Solvent A: | 100% water; 10mM ammonium acetate |
| Solvent B: | 100% acetonitrile |
| Chromatography Type: | HILIC |
Analysis:
| Analysis ID: | AN006092 |
| Laboratory Name: | Metabolomics Core Facility at UT Southwestern Medical Center |
| Analysis Type: | MS |
| Chromatography ID: | CH004624 |
| Num Factors: | 2 |
| Num Metabolites: | 193 |
| Units: | Normalized peak area |
| Analysis ID: | AN006093 |
| Laboratory Name: | Metabolomics Core Facility at UT Southwestern Medical Center |
| Analysis Type: | MS |
| Chromatography ID: | CH004624 |
| Num Factors: | 2 |
| Num Metabolites: | 196 |
| Units: | Normalized peak area |
