Summary of Study ST001673
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 PR001076. The data can be accessed directly via it's Project DOI: 10.21228/M8MH5J This work is supported by NIH grant, U2C- DK119886.
See: https://www.metabolomicsworkbench.org/about/howtocite.php
| Study ID | ST001673 |
| Study Title | Targeted Sphingolipid analysis of HeLa silenced for or overexpressing GOLPH3 or LCS |
| Study Summary | A group of sequentially-acting enzymes operating at the branchpoint among sphingolipid synthetic pathways binds the Golgi-localised oncoprotein GOLPH3. GOLPH3 sorts these enzymes into vesicles for intra-Golgi retro-transport, acting as a component of the cisternae inter-conversion mechanisms. Through these effects, GOLPH3 controls the sub-Golgi localisation, and the lysosomal degradation rate of specific enzymes. Here we evaluated the impact of overexpressing or silencing GOLPH3 or its client enzyme lactosylceramide synthase (LCS) on the sphingolipid composition of HeLa cells by targeted lipid analysis. |
| Institute | École polytechnique fédérale de Lausanne (EPFL) |
| Department | IBI |
| Laboratory | UPDANGELO |
| Last Name | D'Angelo |
| First Name | Giovanni |
| Address | Station 15, Lausanne, Vaud, 1015, Switzerland |
| giovanni.dangelo@epfl.ch | |
| Phone | +41 216934276 |
| Submit Date | 2021-01-29 |
| Raw Data Available | Yes |
| Raw Data File Type(s) | raw(Thermo) |
| Analysis Type Detail | LC-MS |
| Release Date | 2021-02-26 |
| Release Version | 1 |
Select appropriate tab below to view additional metadata details:
Project:
| Project ID: | PR001076 |
| Project DOI: | doi: 10.21228/M8MH5J |
| Project Title: | Targeted Sphingolipid analysis of HeLa silenced for or overexpressing GOLPH3 or LCS |
| Project Summary: | A group of sequentially-acting enzymes operating at the branchpoint among sphingolipid synthetic pathways binds the Golgi-localised oncoprotein GOLPH3. GOLPH3 sorts these enzymes into vesicles for intra-Golgi retro-transport, acting as a component of the cisternae inter-conversion mechanisms. Through these effects, GOLPH3 controls the sub-Golgi localisation, and the lysosomal degradation rate of specific enzymes. Here we evaluated the impact of overexpressing or silencing GOLPH3 or its client enzyme lactosylceramide synthase (LCS) on the sphingolipid composition of HeLa cells by targeted lipid analysis. |
| Institute: | École polytechnique fédérale de Lausanne (EPFL) |
| Department: | IBI |
| Laboratory: | UPDANGELO |
| Last Name: | D'Angelo |
| First Name: | Giovanni |
| Address: | Station 15, Lausanne, Vaud, 1015, Switzerland |
| Email: | giovanni.dangelo@epfl.ch |
| Phone: | +41 216934276 |
Subject:
| Subject ID: | SU001750 |
| Subject Type: | Cultured cells |
| Subject Species: | Homo sapiens |
| Taxonomy ID: | 9606 |
Factors:
Subject type: Cultured cells; Subject species: Homo sapiens (Factor headings shown in green)
| mb_sample_id | local_sample_id | Treatment |
|---|---|---|
| SA154464 | GOLPH3_OE_1 | GOLPH3 overexpression |
| SA154465 | GOLPH3_OE_3 | GOLPH3 overexpression |
| SA154466 | GOLPH3_OE_2 | GOLPH3 overexpression |
| SA154467 | GOLPH3_KD_3 | GOLPH3 silencing |
| SA154468 | GOLPH3_KD_1 | GOLPH3 silencing |
| SA154469 | GOLPH3_KD_2 | GOLPH3 silencing |
| SA154470 | LCS_OE_2 | LCS overexpression |
| SA154471 | LCS_OE_1 | LCS overexpression |
| SA154472 | LCS_OE_3 | LCS overexpression |
| SA154473 | LCS_KD_3 | LCS silencing |
| SA154474 | LCS_KD_2 | LCS silencing |
| SA154475 | LCS_KD_1 | LCS silencing |
| SA154476 | CTRL_2 | no treatment |
| SA154477 | CTRL_1 | no treatment |
| SA154478 | CTRL_3 | no treatment |
| Showing results 1 to 15 of 15 |
Collection:
| Collection ID: | CO001743 |
| Collection Summary: | Cells were washed in ice cold PBS and collected by scaling |
| Sample Type: | HeLa cells |
Treatment:
| Treatment ID: | TR001763 |
| Treatment Summary: | Cells were treated to overexposes or down regulate GOLPH3 or LCS |
Sample Preparation:
| Sampleprep ID: | SP001756 |
| Sampleprep Summary: | Samples were fortified with an Internal standard Mix and lipids were extracted twice with an extraction mix consisting of 85:15 Ethyl acetate 70% Isopropanol. All lipids that were used in the Internal standard mix and in the Calibration mixes were purchased from Avanti Polar Lipids Inc. After evaporating the cell extract to dryness, the samples were reconstituted in the mobile phase. |
Combined analysis:
| Analysis ID | AN002731 |
|---|---|
| Analysis type | MS |
| Chromatography type | Reversed phase |
| Chromatography system | Thermo Accela 1250 |
| Column | Thermo Accucore C18 (100 x 2.1mm,2.6um) |
| MS Type | ESI |
| MS instrument type | Triple quadrupole |
| MS instrument name | Thermo Quantum Ultra |
| Ion Mode | POSITIVE |
| Units | pmoles |
Chromatography:
| Chromatography ID: | CH002017 |
| Chromatography Summary: | Samples were analyzed with a Quantum Ultra triple quadrupole mass spectrometer connected to an Accela HPLC and Accela autosampler using a solvent gradient. Ceramides identity was achieved through MRM analysis with soft fragmentation. Quantitative analysis is based on calibration curves generated for each ceramide. J. Bielawski et al. / Methods 39 (2006) 82–91 |
| Instrument Name: | Thermo Accela 1250 |
| Column Name: | Thermo Accucore C18 (100 x 2.1mm,2.6um) |
| Chromatography Type: | Reversed phase |
MS:
| MS ID: | MS002528 |
| Analysis ID: | AN002731 |
| Instrument Name: | Thermo Quantum Ultra |
| Instrument Type: | Triple quadrupole |
| MS Type: | ESI |
| MS Comments: | Samples were analyzed with a Quantum Ultra triple quadrupole mass spectrometer connected to an Accela HPLC and Accela autosampler using a solvent gradient. Ceramides identity was achieved through MRM analysis with soft fragmentation. Quantitative analysis is based on calibration curves generated for each ceramide. J. Bielawski et al. / Methods 39 (2006) 82–91 |
| Ion Mode: | POSITIVE |
