Raptor, a positive regulatory subunit of mTOR complex 1, is a novel phosphoprotein of the rDNA transcription machinery in nucleoli and chromosomal nucleolus organizer regions (NORs)

Cell Cycle. 2011 Sep 15;10(18):3140-52. doi: 10.4161/cc.10.18.17376. Epub 2011 Sep 15.

Abstract

Raptor is the key scaffolding protein that recruits mTOR substrates to rapamycin-sensitive mTOR complex 1 (mTORC1), a molecular integrator of mitogenic and nutrient/energy environmental inputs into protein translation and cell growth. Although Raptor phosphorylation on various sites is pivotal in the regulation of mTORC1 activity, it remains to be elucidated whether site-specific phosphorylation differentially distributes Raptor to unique subcellular compartments. When exploring the spatiotemporal cell cycle dynamics of six different phospho (P)-Raptor isoforms (Thr ( 706) , Ser ( 722) , Ser ( 863) , Ser ( 792) and Ser ( 877) ), a number of remarkable events differentially defined a topological resetting of P-RaptorThr706 on interphasic and mitotic chromosomes. In interphase nuclei, P-Raptor (Thr706) co-localized with fibrillarin, a component of the nucleolar small nuclear ribonucleoprotein particle, as well as with RNA polymerase I, the enzyme that transcribes nucleolar rRNA. Upon Actinomycin D-induced nucleolar segregation and disaggregation, P-RaptorThr706 was excluded from the nucleolus to accumulate at discrete nucleoplasmic bodies. During mitosis, CDK1 inhibition-induced premature assembly of nucleoli relocated fibrillarin to the surrounding regions of chromosomal-associated P-Raptor (Thr706) , suggesting that a subpopulation of mitotic P-Raptor (Thr706) remained targeted at chromosomal loops of rDNA or nuclear organizer regions (NORs). At the end of mitosis and cytokinesis, when reassembly of incipient nucleoli begins upon NORs activation of rDNA transcription, fibrillarin spatially reorganized with P-Raptor (Thr706) to give rise to daughter nucleoli. Treatment with IGF1 exclusively hyperactivated nuclear P-Raptor (Ser706) and concomitantly promoted Ser ( 2481) autophosphorylation of mTOR, which monitors mTORC1-associated catalytic activity. Nucleolar- and NOR-associated P-Raptor (Ser706) may physically link mTORC1 signaling to ever-growing nucleolus plurifunctionality including ribosome biogenesis, cell stress sensor and cell cycle/aging control.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Adaptor Proteins, Signal Transducing / genetics
  • Adaptor Proteins, Signal Transducing / metabolism*
  • Benzimidazoles / pharmacology
  • Cell Cycle Proteins / antagonists & inhibitors
  • Cell Cycle Proteins / metabolism
  • Cell Nucleolus / drug effects
  • Cell Nucleolus / genetics
  • Cell Nucleolus / metabolism
  • Centrosome / metabolism
  • Chromosomal Proteins, Non-Histone / genetics
  • Chromosomal Proteins, Non-Histone / metabolism
  • Chromosomes, Human / genetics
  • Chromosomes, Human / metabolism
  • Cytokinesis
  • DNA, Ribosomal / genetics
  • DNA, Ribosomal / metabolism
  • Dactinomycin / pharmacology
  • Female
  • Fluorescent Antibody Technique
  • HeLa Cells
  • Humans
  • Interphase
  • Mechanistic Target of Rapamycin Complex 1
  • Microscopy, Confocal
  • Mitosis
  • Multiprotein Complexes
  • Nucleolus Organizer Region / genetics
  • Nucleolus Organizer Region / metabolism*
  • Phosphorylation
  • Protein Isoforms / genetics
  • Protein Isoforms / metabolism
  • Protein-Serine-Threonine Kinases / antagonists & inhibitors
  • Protein-Serine-Threonine Kinases / metabolism
  • Proteins / genetics
  • Proteins / metabolism*
  • Proto-Oncogene Proteins / antagonists & inhibitors
  • Proto-Oncogene Proteins / metabolism
  • Regulatory-Associated Protein of mTOR
  • TOR Serine-Threonine Kinases / genetics
  • TOR Serine-Threonine Kinases / metabolism
  • Thiophenes / pharmacology
  • Transcription, Genetic
  • Transcriptional Activation

Substances

  • 5-(5,6-dimethoxy-1H-benzimidazol-1-yl)-3-((2-(trifluoromethyl)benzyl)oxy)thiophene-2-carboxamide
  • Adaptor Proteins, Signal Transducing
  • Benzimidazoles
  • Cell Cycle Proteins
  • Chromosomal Proteins, Non-Histone
  • DNA, Ribosomal
  • Multiprotein Complexes
  • Protein Isoforms
  • Proteins
  • Proto-Oncogene Proteins
  • RPTOR protein, human
  • Regulatory-Associated Protein of mTOR
  • Thiophenes
  • fibrillarin
  • Dactinomycin
  • MTOR protein, human
  • TOR Serine-Threonine Kinases
  • Mechanistic Target of Rapamycin Complex 1
  • Protein-Serine-Threonine Kinases
  • polo-like kinase 1