Phosphorylation of sterol regulatory element binding protein-1a by protein kinase A (PKA) regulates transcriptional activity

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Abstract

The counter-regulatory hormone glucagon inhibits lipogenesis via downregulation of sterol regulatory element binding protein 1 (SREBP-1). The effect of glucagon is mediated via protein kinase A (PKA). To determine if SREBP-1 is a direct phosphorylation target of PKA, we conducted mass spectrometry analysis of recombinant n-terminal SREBP-1a following PKA treatment in vitro. This analysis identified serines 331/332 as bona-fide phosphorylation targets of PKA. To determine the functional consequences of phosphorylation at these sites, we constructed mammalian expression vector for both nSREBP-1a and 1c isoforms in which the candidate PKA phosphorylation sites were mutated to active phosphomimetic or nonphosphorylatable amino acids. The transcriptional activity of SREBP was reduced by the phosphomimetic mutation of S332 of nSREBP-1a and the corresponding serine (S308) of nSREBP-1c. This site is a strong candidate for mediating the negative regulatory effect of glucagon on SREBP-1 and lipogenesis.

Original languageEnglish (US)
Pages (from-to)449-454
Number of pages6
JournalBiochemical and Biophysical Research Communications
Volume449
Issue number4
DOIs
StatePublished - Jan 1 2014

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Sterol Regulatory Element Binding Protein 1
Phosphorylation
Cyclic AMP-Dependent Protein Kinases
Glucagon
Lipogenesis
Serine
Mass spectrometry
Mass Spectrometry
Protein Isoforms
Down-Regulation
Hormones
Amino Acids
Mutation

All Science Journal Classification (ASJC) codes

  • Biophysics
  • Biochemistry
  • Molecular Biology
  • Cell Biology

Cite this

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title = "Phosphorylation of sterol regulatory element binding protein-1a by protein kinase A (PKA) regulates transcriptional activity",
abstract = "The counter-regulatory hormone glucagon inhibits lipogenesis via downregulation of sterol regulatory element binding protein 1 (SREBP-1). The effect of glucagon is mediated via protein kinase A (PKA). To determine if SREBP-1 is a direct phosphorylation target of PKA, we conducted mass spectrometry analysis of recombinant n-terminal SREBP-1a following PKA treatment in vitro. This analysis identified serines 331/332 as bona-fide phosphorylation targets of PKA. To determine the functional consequences of phosphorylation at these sites, we constructed mammalian expression vector for both nSREBP-1a and 1c isoforms in which the candidate PKA phosphorylation sites were mutated to active phosphomimetic or nonphosphorylatable amino acids. The transcriptional activity of SREBP was reduced by the phosphomimetic mutation of S332 of nSREBP-1a and the corresponding serine (S308) of nSREBP-1c. This site is a strong candidate for mediating the negative regulatory effect of glucagon on SREBP-1 and lipogenesis.",
author = "Qingming Dong and Francesco Giorgianni and Xiong Deng and Sarka Beranova and Dave Bridges and Edwards Park and Rajendra Raghow and Marshall Elam",
year = "2014",
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T1 - Phosphorylation of sterol regulatory element binding protein-1a by protein kinase A (PKA) regulates transcriptional activity

AU - Dong, Qingming

AU - Giorgianni, Francesco

AU - Deng, Xiong

AU - Beranova, Sarka

AU - Bridges, Dave

AU - Park, Edwards

AU - Raghow, Rajendra

AU - Elam, Marshall

PY - 2014/1/1

Y1 - 2014/1/1

N2 - The counter-regulatory hormone glucagon inhibits lipogenesis via downregulation of sterol regulatory element binding protein 1 (SREBP-1). The effect of glucagon is mediated via protein kinase A (PKA). To determine if SREBP-1 is a direct phosphorylation target of PKA, we conducted mass spectrometry analysis of recombinant n-terminal SREBP-1a following PKA treatment in vitro. This analysis identified serines 331/332 as bona-fide phosphorylation targets of PKA. To determine the functional consequences of phosphorylation at these sites, we constructed mammalian expression vector for both nSREBP-1a and 1c isoforms in which the candidate PKA phosphorylation sites were mutated to active phosphomimetic or nonphosphorylatable amino acids. The transcriptional activity of SREBP was reduced by the phosphomimetic mutation of S332 of nSREBP-1a and the corresponding serine (S308) of nSREBP-1c. This site is a strong candidate for mediating the negative regulatory effect of glucagon on SREBP-1 and lipogenesis.

AB - The counter-regulatory hormone glucagon inhibits lipogenesis via downregulation of sterol regulatory element binding protein 1 (SREBP-1). The effect of glucagon is mediated via protein kinase A (PKA). To determine if SREBP-1 is a direct phosphorylation target of PKA, we conducted mass spectrometry analysis of recombinant n-terminal SREBP-1a following PKA treatment in vitro. This analysis identified serines 331/332 as bona-fide phosphorylation targets of PKA. To determine the functional consequences of phosphorylation at these sites, we constructed mammalian expression vector for both nSREBP-1a and 1c isoforms in which the candidate PKA phosphorylation sites were mutated to active phosphomimetic or nonphosphorylatable amino acids. The transcriptional activity of SREBP was reduced by the phosphomimetic mutation of S332 of nSREBP-1a and the corresponding serine (S308) of nSREBP-1c. This site is a strong candidate for mediating the negative regulatory effect of glucagon on SREBP-1 and lipogenesis.

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