mTORC1 alters the expression of glycolytic genes by regulating KPNA2 abundances

J Proteomics. 2016 Mar 16:136:13-24. doi: 10.1016/j.jprot.2016.01.021. Epub 2016 Feb 2.

Abstract

Mammalian target of rapamycin complex 1 (mTORC1) plays important roles in regulating cell growth and proliferation, and the aberrant activation of mTORC1 has been observed in many human diseases. However, the proteins regulated by mTORC1 activation and their roles in mTORC1 downstream functions are still poorly understood. Using proteomic analysis, we found that proteins regulated by mTORC1 in MEFs could be categorized into eight functional groups including protein nuclear import and glycolysis. The positive regulation of Karyopherin subunit alpha-2 (KPNA2), an importer protein involved in protein nuclear import, by mTORC1 was verified in several other mouse and human cell lines. The regulation occurred at the transcriptional level, rather than at the level of S6K1- and 4E-BP1-dependent protein synthesis. KPNA2 knockdown partially blocked upregulation of glycolytic genes by mTORC1 activation, indicating that mTORC1 activation enhanced expression of glycolytic genes by increasing KPNA2 abundances. Furthermore, KPNA2 knockdown had no effects on the expression and subcellular localization of HIF1α, a transcription factor involved in regulating glycolytic genes downstream of mTORC1. In conclusion, our results proved that KPNA2 regulated the expression of glycolytic genes downstream of mTORC1 in a HIF1α-independent manner.

Significance: Identifying mTORC1-regulated proteins through proteomic method is a feasible way to study the downstream functions of mTORC1. In this study, we identified many mTORC1-regulated proteins using proteomic analysis by overlapping two different high vs low/no mTORC1 activity comparisons, TSC2(-/-) vs WT MEFs and TSC2(-/-) with/without rapamycin treatment. We found the abundances of many enzymes in glycolysis pathway and several proteins involved in protein nuclear import were positively regulated by mTORC1. More importantly, we first discovered that mTORC1 positively regulated the importer protein KPNA2, which participated in glycolysis regulation downstream of mTORC1 in a HIF1α-independent manner, indicating that mTORC1 regulates glycolysis through multiple ways.

Keywords: Glycolysis; HIF1α; KPNA2; Proteomics; Rapamycin; mTORC1.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Animals
  • Cell Line, Transformed
  • Eukaryotic Initiation Factors / biosynthesis
  • Eukaryotic Initiation Factors / genetics
  • Gene Expression Regulation / physiology*
  • Glycolysis / physiology*
  • Humans
  • Hypoxia-Inducible Factor 1, alpha Subunit / genetics
  • Hypoxia-Inducible Factor 1, alpha Subunit / metabolism*
  • Mechanistic Target of Rapamycin Complex 1
  • Mice
  • Mice, Knockout
  • Multiprotein Complexes / genetics
  • Multiprotein Complexes / metabolism*
  • Nuclear Proteins / genetics
  • Nuclear Proteins / metabolism*
  • Ribosomal Protein S6 Kinases, 90-kDa / biosynthesis
  • Ribosomal Protein S6 Kinases, 90-kDa / genetics
  • TOR Serine-Threonine Kinases / genetics
  • TOR Serine-Threonine Kinases / metabolism*
  • alpha Karyopherins

Substances

  • Eif4ebp2 protein, mouse
  • Eukaryotic Initiation Factors
  • Hif1a protein, mouse
  • Hypoxia-Inducible Factor 1, alpha Subunit
  • Kpna2 protein, mouse
  • Multiprotein Complexes
  • Nuclear Proteins
  • alpha Karyopherins
  • Mechanistic Target of Rapamycin Complex 1
  • Ribosomal Protein S6 Kinases, 90-kDa
  • Rps6ka1 protein, mouse
  • TOR Serine-Threonine Kinases