Nuclear phosphoproteome analysis of 3T3-L1 preadipocyte differentiation reveals system-wide phosphorylation of transcriptional regulators

Atefeh Rabiee, Veit Schwämmle, Simone Sidoli, Jie Dai, Adelina Rogowska-Wrzesinska, Susanne Mandrup, Ole N. Jensen

Research output: Contribution to journalArticle

4 Citations (Scopus)

Abstract

Adipocytes (fat cells) are important endocrine and metabolic cells critical for systemic insulin sensitivity. Both adipose excess and insufficiency are associated with adverse metabolic function. Adipogenesis is the process whereby preadipocyte precursor cells differentiate into lipid-laden mature adipocytes. This process is driven by a network of transcriptional regulators (TRs). We hypothesized that protein PTMs, in particular phosphorylation, play a major role in activating and propagating signals within TR networks upon induction of adipogenesis by extracellular stimulus. We applied MS-based quantitative proteomics and phosphoproteomics to monitor the alteration of nuclear proteins during the early stages (4 h) of preadipocyte differentiation. We identified a total of 4072 proteins including 2434 phosphorylated proteins, a majority of which were assigned as regulators of gene expression. Our results demonstrate that adipogenic stimuli increase the nuclear abundance and/or the phosphorylation levels of proteins involved in gene expression, cell organization, and oxidation–reduction pathways. Furthermore, proteins acting as negative modulators involved in negative regulation of gene expression, insulin stimulated glucose uptake, and cytoskeletal organization showed a decrease in their nuclear abundance and/or phosphorylation levels during the first 4 h of adipogenesis. Among 288 identified TRs, 49 were regulated within 4 h of adipogenic stimulation including several known and many novel potential adipogenic regulators. We created a kinase–substrate database for 3T3-L1 preadipocytes by investigating the relationship between protein kinases and protein phosphorylation sites identified in our dataset. A majority of the putative protein kinases belong to the cyclin-dependent kinase family and the mitogen-activated protein kinase family including P38 and c-Jun N-terminal kinases, suggesting that these kinases act as orchestrators of early adipogenesis.

Original languageEnglish (US)
Article number1600248
JournalProteomics
Volume17
Issue number6
DOIs
StatePublished - Mar 1 2017
Externally publishedYes

Fingerprint

Phosphorylation
Adipogenesis
Adipocytes
Gene expression
Proteins
Gene Regulatory Networks
Protein Kinases
Pulse time modulation
Insulin
Gene Expression
Endocrine Cells
Cyclin-Dependent Kinases
JNK Mitogen-Activated Protein Kinases
Gene Expression Regulation
Regulator Genes
Nuclear Proteins
Mitogen-Activated Protein Kinases
Proteomics
Modulators
Insulin Resistance

Keywords

  • Adipogenesis
  • Cell biology
  • Kinase networks
  • Nuclear proteome
  • Phosphopeptide enrichment
  • Quantitative phosphoproteomics
  • Transcriptional regulators

ASJC Scopus subject areas

  • Biochemistry
  • Molecular Biology

Cite this

Nuclear phosphoproteome analysis of 3T3-L1 preadipocyte differentiation reveals system-wide phosphorylation of transcriptional regulators. / Rabiee, Atefeh; Schwämmle, Veit; Sidoli, Simone; Dai, Jie; Rogowska-Wrzesinska, Adelina; Mandrup, Susanne; Jensen, Ole N.

In: Proteomics, Vol. 17, No. 6, 1600248, 01.03.2017.

Research output: Contribution to journalArticle

Rabiee, Atefeh ; Schwämmle, Veit ; Sidoli, Simone ; Dai, Jie ; Rogowska-Wrzesinska, Adelina ; Mandrup, Susanne ; Jensen, Ole N. / Nuclear phosphoproteome analysis of 3T3-L1 preadipocyte differentiation reveals system-wide phosphorylation of transcriptional regulators. In: Proteomics. 2017 ; Vol. 17, No. 6.
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AU - Rabiee, Atefeh

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AU - Sidoli, Simone

AU - Dai, Jie

AU - Rogowska-Wrzesinska, Adelina

AU - Mandrup, Susanne

AU - Jensen, Ole N.

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AB - Adipocytes (fat cells) are important endocrine and metabolic cells critical for systemic insulin sensitivity. Both adipose excess and insufficiency are associated with adverse metabolic function. Adipogenesis is the process whereby preadipocyte precursor cells differentiate into lipid-laden mature adipocytes. This process is driven by a network of transcriptional regulators (TRs). We hypothesized that protein PTMs, in particular phosphorylation, play a major role in activating and propagating signals within TR networks upon induction of adipogenesis by extracellular stimulus. We applied MS-based quantitative proteomics and phosphoproteomics to monitor the alteration of nuclear proteins during the early stages (4 h) of preadipocyte differentiation. We identified a total of 4072 proteins including 2434 phosphorylated proteins, a majority of which were assigned as regulators of gene expression. Our results demonstrate that adipogenic stimuli increase the nuclear abundance and/or the phosphorylation levels of proteins involved in gene expression, cell organization, and oxidation–reduction pathways. Furthermore, proteins acting as negative modulators involved in negative regulation of gene expression, insulin stimulated glucose uptake, and cytoskeletal organization showed a decrease in their nuclear abundance and/or phosphorylation levels during the first 4 h of adipogenesis. Among 288 identified TRs, 49 were regulated within 4 h of adipogenic stimulation including several known and many novel potential adipogenic regulators. We created a kinase–substrate database for 3T3-L1 preadipocytes by investigating the relationship between protein kinases and protein phosphorylation sites identified in our dataset. A majority of the putative protein kinases belong to the cyclin-dependent kinase family and the mitogen-activated protein kinase family including P38 and c-Jun N-terminal kinases, suggesting that these kinases act as orchestrators of early adipogenesis.

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KW - Phosphopeptide enrichment

KW - Quantitative phosphoproteomics

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