Summary of Study ST002206

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 PR001409. The data can be accessed directly via it's Project DOI: 10.21228/M8KT4Z 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	ST002206
Study Title	Lipolysis-derived Lipids Determine Autophagy Initiation during Fasting
Study Summary	For survival, autophagy is a crucial intracellular self-degradation process to provide energy sources, helping adapt to nutrient deprivation. Although nutrient availability is a key determinant of autophagy initiation, it remains elusive underlying mechanism(s) of perceiving nutritional scarcity by which cells timely turn on autophagy as the last self-destructive process for energy supply. Here, we showed that PKA-dependent lipolysis can block the initiation of futile autophagy during short-term nutritional deprivation by repressing AMPK. Using Raman microscopy imaging and metabolomics, we found that autophagy occurred by reduction in available free fatty acids (FFAs) for energy sources. By modulating genes involved in lipolysis and fatty acid oxidation, we found that the use of lipolysis-derived FFAs precedes autophagy initiation. The dysregulated autophagy suppression during short-term fasting decreased motility and lifespan extension of worms. Taken together, these data suggest that PKA is a pivotal factor to orchestrate sophisticated catabolic pathways, preferring the use of PKA-mediated lipolytic products to repress futile autophagic degradation during short-term fasting through AMPK inhibition.
Institute	Seoul National University
Last Name	Ji
First Name	Yul
Address	San 56-1, Sillim-Dong, Kwanak-Gu, Seoul, Seoul, 08826, Korea, South
Email	withyul@snu.ac.kr
Phone	01025076912
Submit Date	2022-04-27
Analysis Type Detail	LC-MS
Release Date	2022-07-22
Release Version	1

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Sample Preparation:

Sampleprep ID:	SP002298
Sampleprep Summary:	1. Sample Preparation for CE-TOFMS analysis The samples were mixed with 50% acetonitrile in water (v/v) containing internal standards (20 μM) as shown in Table 1 and homogenized by a homogenizer (1,500 rpm, 120 sec × 2 times), then, the same amount of 50% acetonitrile in water (v/v) were added.. The supernatant (400 μL) was then filtrated through 5-kDa cut-off filter(ULTRAFREE-MC-PLHCC, Human Metabolome Technologies, Yamagata, Japan) to remove macromolecules. The filtrate was centrifugally concentrated and resuspended in 25 μL of ultrapure water immediately before the measurement. 2. Sample Preparation for LC-TOFMS analysis The samples were mixed with 500 μL of 1% formic acid in acetonitrile (v/v) containing internal standards (10 μM), homogenized by a homogenizer (1,500 rpm, 120 sec × 2 times). The mixture was yet again homogenized after adding 167 μL of Milli-Q water and then centrifuged (2,300 x g, 4℃, 5min). After the supernatant was collected, 500 μL of 1% formic acid in acetonitrile (v/v) and 167 μL of MilliQ-water were added to the precipitation. The homogenization and centrifugation was performed as described previously, and the supernatant was mixed with previously collected one. The mixed supernatant was filtrated through 3-kDa cut-off filter (NANOCEP 3K OMEGA, PALL Corporation, Michigan, USA) to remove proteins and far filtrated through column (Hybrid SPE phospholipid 55261-U, Supelco, Bellefonte,PA, USA) to remove phospholipids. The filtrate was desiccated and resuspended in 200 μL of 50% isopropanol in Milli-Q water (v/v) immediately before the measurement.

Sampleprep ID:

SP002298

Sampleprep Summary:

1. Sample Preparation for CE-TOFMS analysis The samples were mixed with 50% acetonitrile in water (v/v) containing internal standards (20 μM) as shown in Table 1 and homogenized by a homogenizer (1,500 rpm, 120 sec × 2 times), then, the same amount of 50% acetonitrile in water (v/v) were added.. The supernatant (400 μL) was then filtrated through 5-kDa cut-off filter(ULTRAFREE-MC-PLHCC, Human Metabolome Technologies, Yamagata, Japan) to remove macromolecules. The filtrate was centrifugally concentrated and resuspended in 25 μL of ultrapure water immediately before the measurement. 2. Sample Preparation for LC-TOFMS analysis The samples were mixed with 500 μL of 1% formic acid in acetonitrile (v/v) containing internal standards (10 μM), homogenized by a homogenizer (1,500 rpm, 120 sec × 2 times). The mixture was yet again homogenized after adding 167 μL of Milli-Q water and then centrifuged (2,300 x g, 4℃, 5min). After the supernatant was collected, 500 μL of 1% formic acid in acetonitrile (v/v) and 167 μL of MilliQ-water were added to the precipitation. The homogenization and centrifugation was performed as described previously, and the supernatant was mixed with previously collected one. The mixed supernatant was filtrated through 3-kDa cut-off filter (NANOCEP 3K OMEGA, PALL Corporation, Michigan, USA) to remove proteins and far filtrated through column (Hybrid SPE phospholipid 55261-U, Supelco, Bellefonte,PA, USA) to remove phospholipids. The filtrate was desiccated and resuspended in 200 μL of 50% isopropanol in Milli-Q water (v/v) immediately before the measurement.