Dataset Information

Proteomic profiling of HepG2 cell line after 2DE fractionation

ABSTRACT: One of the major goals of the Chromosome-centric Human Proteome Project (C-HPP) is to catalog and annotate myriads of heterogeneous proteoforms, produced by ca. 20 thousand genes. To achieve deeper and personalized dive into proteome we suggest to use customized RNA-seq library of potential proteoforms, which includes aberrant variants, specific to the certain biological sample. Multidimensional fractionation by means of two-dimensional electrophoresis and high-performance liquid chromatography, allowed us to downgrade the difficulty of biological mixture for the following shotgun mass spectrometry. To benchmark proposed pipeline we examined heterogeneity of the HepG2 hepatoblastome cell line proteome.

INSTRUMENT(S): Q Exactive

ORGANISM(S): Homo Sapiens (human)

TISSUE(S): Cell Culture

SUBMITTER: Olga Kiseleva

LAB HEAD: Ekaterina Poverennaya

PROVIDER: PXD018450 | Pride | 2020-06-01

REPOSITORIES: Pride

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Publications

Empowering Shotgun Mass Spectrometry with 2DE: A HepG2 Study.

Kiseleva Olga O Ponomarenko Elena E Poverennaya Ekaterina E

International journal of molecular sciences 20200527 11

One of the major goals of the Chromosome-Centric Human Proteome Project (C-HPP) is to catalog and annotate a myriad of heterogeneous proteoforms, produced by ca. 20 thousand genes. To achieve a detailed and personalized understanding into proteomes, we suggest using a customized RNA-seq library of potential proteoforms, which includes aberrant variants specific to certain biological samples. Two-dimensional electrophoresis coupled with high-performance liquid chromatography allowed us to downgra ...[more]

PMID: 32471280

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Project description:Lipoprotein lipase (LPL) is responsible for the intravascular catabolism of triglyceride-rich lipoproteins and plays a central role in whole-body energy balance and lipid homeostasis. As such, LPL is subject to tissue-specific regulation in different physiological conditions, but the mechanisms of this regulation remain incompletely characterized. Previous work revealed that LPL comprises a set of proteoforms with different isoelectric points, but their regulation and functional significance have not been studied thus far. Here we studied the distribution of LPL proteoforms in different rat tissues and their regulation under physiological conditions. First, analysis by two-dimensional electrophoresis and Western blot showed different patterns of LPL proteoforms (i.e., different pI or relative abundance of LPL proteoforms) in different rat tissues under basal conditions, which could be related to the tissue-specific regulation of the enzyme. Next, the comparison of LPL proteoforms from heart and brown adipose tissue between adults and 15-day-old rat pups, two conditions with minimal regulation of LPL in these tissues, yielded virtually the same tissue-specific patterns of LPL proteoforms. In contrast, the pronounced down-regulation of LPL activity observed in white adipose tissue during fasting is accompanied by a prominent reconfiguration of the LPL proteoform pattern. Furthermore, refeeding reverts this down-regulation of LPL activity and restores the pattern of LPL proteoforms in this tissue. Importantly, this reversible proteoform-specific regulation during fasting and refeeding indicates that LPL proteoforms are functionally diverse. Further investigation of potential differences in the functional properties of LPL proteoforms showed that all proteoforms exhibit lipolytic activity and have similar heparin-binding affinity, although other functional aspects remain to be investigated. Overall, this study demonstrates the ubiquity, differential distribution and specific regulation of LPL proteoforms in rat tissues and underscores the need to consider the existence of LPL proteoforms for a complete understanding of LPL regulation under physiological conditions.

			Action	DRS
	01_1.cpsx	Other
	01_1.mgf	Mgf
	01_1.raw	Raw
	01_2.cpsx	Other
	01_2.mgf	Mgf