Summary of study ST001859

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 PR001172. The data can be accessed directly via it's Project DOI: 10.21228/M86T2S This work is supported by NIH grant, U2C- DK119886.

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Study IDST001859
Study TitledTor affects the fat body lipidome via Nep1r1, Ctdnep1 and Lipin
Study SummaryQuantitative MS analysis was performed on ten 4 day-old Drosophila larval fat bodies homogenized in 50µl D-PBS (Dulbecco’s Phosphate Buffered Saline without Mg2+ and Ca2+) by Lipotype using previously described methods (Grillet et al, 2016).
Institute
VIB KULeuven
DepartmentDept. of Neurosciences, KU Leuven, 3000 Leuven, Belgium
Last NameJacquemyn
First NameJulie
AddressON 4, 6e verd Campus Gasthuisberg, Herestraat 49, bus 602, Leuven, NA, 3000, Belgium
Emailjulie.jacquemyn@kuleuven.be
Phone0032479570951
Submit Date2021-06-22
Analysis Type DetailLC-MS
Release Date2021-07-18
Release Version1
Julie Jacquemyn Julie Jacquemyn
https://dx.doi.org/10.21228/M86T2S
ftp://www.metabolomicsworkbench.org/Studies/ application/zip

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Project:

Project ID:PR001172
Project DOI:doi: 10.21228/M86T2S
Project Title:Torsin and NEP1R1-CTDNEP1 affect interphase NPC insertion by lipid-dependent and -independent mechanisms
Project Type:MS analysis performed by Lipotype on Drosophila larval fat body tissue
Project Summary:The interphase nuclear envelope is extensively remodeled during nuclear pore complex insertion. The process is relatively poorly understood, including why it requires the Torsin ATPases that also regulate NE-localized lipid metabolism. Here we show that fly dTorsin affects lipid metabolism through the NEP1R1-CTDNEP1 phosphatase/ Lipin phosphatidic acid phosphatase pathway. At a cellular level, fly and mouse Torsins removed NEP1R1-CTDNEP1 from the NE to, in turn, exclude Lipin from the nucleus. NEP1R1-CTDNEP1 downregulation also restored nuclear pore membrane fusion in post-mitotic dTorsinKO fat body cells. However, Lipin downregulation was ineffective and membrane fusion defects did not correlate with lipidomic abnormalities. Further testing confirmed that membrane fusion continued in cells with excess Lipin function. It also led to the surprising finding that excess PA metabolism inhibited recruitment of the inner ring complex Nup35 subunit, resulting in elongated channel-like structures in place of mature nuclear pores. We conclude that the NEP1R1-CTDNEP1 phosphatase affects interphase NPC biogenesis by lipid-dependent and lipid-independent mechanisms and this explains some of the pleiotropic effects of Torsins.
Institute:VIB KULeuven
Department:Dept. of Neurosciences, KU Leuven, 3000 Leuven, Belgium
Last Name:Jacquemyn
First Name:Julie
Address:ON 4, 6e verd Campus Gasthuisberg, Herestraat 49, bus 602, 3000 Leuven, Belgium
Email:julie.jacquemyn@kuleuven.be
Phone:032479570951

Subject:

Subject ID:SU001936
Subject Type:Invertebrate
Subject Species:Drosophila melanogaster
Taxonomy ID:7227
Age Or Age Range:Developmentally-staged-larvae were collected (randomized) 4 days after placing male and female flies in fresh vials for a 12-hour period.
Gender:Male and female

Factors:

Subject type: Invertebrate; Subject species: Drosophila melanogaster (Factor headings shown in green)

mb_sample_id local_sample_id Genotype
SA174315s3-3dTorKO Ctdnep1
SA174316s3-1dTorKO Ctdnep1
SA174317s3-2dTorKO Ctdnep1
SA174318s4-3dTorKO Lipin
SA174319s4-1dTorKO Lipin
SA174320s4-2dTorKO Lipin
SA174321s1-2dTorKO Luciferase
SA174322s1-1dTorKO Luciferase
SA174323s1-3dTorKO Luciferase
SA174324s2-2dTorKO Nep1r1
SA174325s2-3dTorKO Nep1r1
SA174326s2-1dTorKO Nep1r1
SA174327s9-2w- Ctdnep1
SA174328s9-3w- Ctdnep1
SA174329s9-1w- Ctdnep1
SA174330s8-3w- Lipin
SA174331s8-2w- Lipin
SA174332s8-1w- Lipin
SA174333s5-2w- Luciferase
SA174334s5-1w- Luciferase
SA174335s5-3w- Luciferase
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Collection:

Collection ID:CO001929
Collection Summary:Fat body tissues of the indicated genotypes were collected and stored at -80°C until processed for MS analysis. Quantitative MS analysis was performed on ten 4 day-old Drosophila larval fat bodies homogenized in 50µl D-PBS (Dulbecco’s Phosphate Buffered Saline without Mg2+ and Ca2+) by Lipotype using previously described methods (Grillet et al, 2016).
Sample Type:Insect tissue

Treatment:

Treatment ID:TR001948
Treatment Summary:In this study we did not apply any additional treatment(s) to the samples.

Sample Preparation:

Sampleprep ID:SP001942
Sampleprep Summary:Lipids from fat body tissues of indicated genotypes were extracted using chloroform and methanol (1). Samples were spiked with lipid class-specific internal standards prior to extraction. After drying and resuspending in MS acquisition mixture, lipid extracts were subjected to mass spectrometric analysis. Sample preparations were done by Lipotype. (1) Sampaio, J. L. et al. Membrane lipidome of an epithelial cell line. Proceedings of the National Academy of Sciences 108, 1903–1907 (2011).

Combined analysis:

Analysis ID AN003013 AN003014
Analysis type MS MS
Chromatography type None (Direct infusion) None (Direct infusion)
Chromatography system none none
Column none none
MS Type ESI ESI
MS instrument type Orbitrap Orbitrap
MS instrument name Thermo Q Exactive Orbitrap Thermo Q Exactive Orbitrap
Ion Mode POSITIVE NEGATIVE
Units pmol pmol

Chromatography:

Chromatography ID:CH002233
Chromatography Summary:NA
Instrument Name:none
Column Name:none
Chromatography Type:None (Direct infusion)

MS:

MS ID:MS002802
Analysis ID:AN003013
Instrument Name:Thermo Q Exactive Orbitrap
Instrument Type:Orbitrap
MS Type:ESI
MS Comments:Spectra acquisition Mass spectra were acquired on a hybrid quadrupole/Orbitrap mass spectrometer equipped with an automated nano-flow electrospray ion source in both positive and negative ion mode. Data processing and normalization Lipid identification using LipotypeXplorer (2) was performed on unprocessed (*.raw format) mass spectra. For MS-only mode, lipid identification was based on the molecular masses of the intact molecules. MSMS mode included the collision induced fragmentation of lipid molecules and lipid identification was based on both the intact masses and the masses of the fragments. Prior to normalization and further statistical analysis lipid identifications were filtered according to mass accuracy, occupation threshold, noise and background. Lists of identified lipids and their intensities were stored in a database optimized for the particular structure inherent to lipidomic datasets. Intensity of lipid class-specific internal standards was used for lipid quantification. (2) Herzog, R. et al. A novel informatics concept for high-throughput shotgun lipidomics based on the molecular fragmentation query language. Genome Biol. 12, R8 (2011).
Ion Mode:POSITIVE
  
MS ID:MS002803
Analysis ID:AN003014
Instrument Name:Thermo Q Exactive Orbitrap
Instrument Type:Orbitrap
MS Type:ESI
MS Comments:Spectra acquisition Mass spectra were acquired on a hybrid quadrupole/Orbitrap mass spectrometer equipped with an automated nano-flow electrospray ion source in both positive and negative ion mode. Data processing and normalization Lipid identification using LipotypeXplorer (2) was performed on unprocessed (*.raw format) mass spectra. For MS-only mode, lipid identification was based on the molecular masses of the intact molecules. MSMS mode included the collision induced fragmentation of lipid molecules and lipid identification was based on both the intact masses and the masses of the fragments. Prior to normalization and further statistical analysis lipid identifications were filtered according to mass accuracy, occupation threshold, noise and background. Lists of identified lipids and their intensities were stored in a database optimized for the particular structure inherent to lipidomic datasets. Intensity of lipid class-specific internal standards was used for lipid quantification. (2) Herzog, R. et al. A novel informatics concept for high-throughput shotgun lipidomics based on the molecular fragmentation query language. Genome Biol. 12, R8 (2011).
Ion Mode:NEGATIVE
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