Summary of Study ST002398

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 PR001545. The data can be accessed directly via it's Project DOI: 10.21228/M8099D 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	ST002398
Study Title	Lipidomics of Tango2 Deficient and Wildtype Zebrafish Muscle Tissue
Study Type	Untargeted Lipidomics
Study Summary	Rhabdomyolysis is a clinical emergency characterized by severe muscle damage, resulting in the release of intracellular muscle components which leads to myoglobinuria and in severe cases, acute kidney failure. Rhabdomyolysis is caused by genetic factors that are linked to increased disease susceptibility in response to extrinsic triggers. Recessive mutations in TANGO2 result in episodic rhabdomyolysis, metabolic crises, encephalopathy, and cardiac arrhythmia. The underlying mechanism contributing to disease onset in response to specific triggers remains unclear. To address these challenges, we created a zebrafish model of Tango2 deficiency. Here we demonstrate that the loss of Tango2 in zebrafish results in growth defects, early lethality, and increased susceptibility of muscle defects similar to TANGO2 patients. Detailed analyses of skeletal muscle revealed defects in the sarcoplasmic reticulum and mitochondria at the onset of disease development. The sarcoplasmic reticulum (SR) constitutes the primary lipid biosynthesis site and regulates calcium handling in skeletal muscle to control excitation-contraction coupling. Tango2 deficient SR exhibits increased sensitivity to calcium release that was partly restored by inhibition of Ryr1-mediated Ca2+ release in skeletal muscle. Using lipidomics, we identified alterations in the glycerolipid state of tango2 mutants which is critical for membrane stability and energy balance. Therefore, these studies provide insight into key disease processes in Tango2 deficiency and have increased our understanding of the impacts of specific defects on predisposition to environmental triggers in TANGO2-related disorders.
Institute	University of North Carolina at Chapel Hill
Department	Chemistry
Laboratory	MS Core Laboratory
Last Name	Wallace
First Name	Emily
Address	131 South Rd
Email	emdiane@email.unc.edu
Phone	7042453664
Submit Date	2022-12-07
Num Groups	2
Total Subjects	5
Num Males	N/A
Num Females	N/A
Study Comments	Zebrafish were all 4 weeks old when tissue was harvested, sex is determined at 4 weeks old.
Raw Data Available	Yes
Raw Data File Type(s)	mzXML
Analysis Type Detail	LC-MS
Release Date	2022-12-30
Release Version	1

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Collection:

Collection ID:	CO002480
Collection Summary:	Fish were bred and maintained using standard methods as described (Westerfield, 2000). All procedures were approved by the Brigham and Women’s Hospital Animal Care and Use Committee. tango2bwh210 and tango2bwh211 zebrafish lines were created in our laboratory by CRISPR-Cas9 approach. Zebrafish embryonic (0-2 days post fertilization) and larval stages (3-45 dpf), juvenile stage (45 dpf-3months) and adults (3 months) have been defined as described previously (Kimmel, Ballard, Kimmel, Ullmann, & Schilling, 1995). All studies presented in this work were performed on tango2bwh211 mutants obtained from heterozygous parents unless specified. Myofibers were isolated from control or tango2 larval zebrafish (45 dpf) as described previously with minor modifications (Ganassi, Zammit, & Hughes, 2021). Skinned zebrafish muscle samples were treated with collagenase for 90 minutes and triturated to release the myofibers. Myofibers were centrifuged at 1000g for 60 sec, washed and resuspended in DMEM media. Myofibers were plated on laminin coated 8 chamber permanox slides (Thermofisher Scientific) for further analysis. Fixed cells were blocked in 10% goat serum/0.3% Triton, incubated in primary antibody overnight at 4ºC, washed in PBS, incubated in secondary antibody for 1 h at room temperature (RT), washed in PBS, then mounted with Vectashield Mounting Medium (Vector Laboratories, Burlingame, CA, USA). Primary antibodies used were anti Tango2 (1:250, 27846-1-AP, Proteintech), mouse monoclonal anti sarcomeric -actinin (1:100, A7732, Millipore Sigma), mouse monoclonal anti Ryr1 (1:100, R129-100UL, Millipore Sigma), anti Tomm20 (1:100, MABT166, Millipore Sigma) and Alexa fluor 568-phalloidin (1:100, Thermo Fisher Scientific, A12380). After washing in PBS several times, samples were incubated with anti-mouse Alexa Fluor (1:100, A-11005) secondary antibody (Thermofisher Scientific). Imaging was performed using a Nikon Ti2 spinning disk confocal microscope.
Sample Type:	Muscle
Collection Method:	Muscle Dissection
Storage Conditions:	Described in summary