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@ARTICLE{MarinZapata:131000,
      author       = {P. A. Marin Zapata$^*$ and C. J. Beese$^*$ and A.
                      Jünger$^*$ and G. Dalmasso$^*$ and N. R. Brady$^*$ and A.
                      Hamacher-Brady$^*$},
      title        = {{T}ime course decomposition of cell heterogeneity in {TFEB}
                      signaling states reveals homeostatic mechanisms restricting
                      the magnitude and duration of {TFEB} responses to m{TOR}
                      activity modulation.},
      journal      = {BMC cancer},
      volume       = {16},
      number       = {1},
      issn         = {1471-2407},
      address      = {London},
      publisher    = {BioMed Central},
      reportid     = {DKFZ-2017-06076},
      pages        = {355},
      year         = {2016},
      abstract     = {TFEB (transcription factor EB) regulates metabolic
                      homeostasis through its activation of lysosomal biogenesis
                      following its nuclear translocation. TFEB activity is
                      inhibited by mTOR phosphorylation, which signals its
                      cytoplasmic retention. To date, the temporal relationship
                      between alterations to mTOR activity states and changes in
                      TFEB subcellular localization and concentration has not been
                      sufficiently addressed.mTOR was activated by renewed
                      addition of fully-supplemented medium, or inhibited by
                      Torin1 or nutrient deprivation. Single-cell TFEB protein
                      levels and subcellular localization in HeLa and MCF7 cells
                      were measured over a time course of 15 hours by
                      multispectral imaging cytometry. To extract single-cell
                      level information on heterogeneous TFEB activity phenotypes,
                      we developed a framework for identification of TFEB activity
                      subpopulations. Through unsupervised clustering, cells were
                      classified according to their TFEB nuclear concentration,
                      which corresponded with downstream lysosomal responses.Bulk
                      population results revealed that mTOR negatively regulates
                      TFEB protein levels, concomitantly to the regulation of TFEB
                      localization. Subpopulation analysis revealed maximal
                      sensitivity of HeLa cells to mTOR activity stimulation,
                      leading to inactivation of $100 \%$ of the cell population
                      within 0.5 hours, which contrasted with a lower sensitivity
                      in MCF7 cells. Conversely, mTOR inhibition increased the
                      fully active subpopulation only fractionally, and full
                      activation of $100 \%$ of the population required
                      co-inhibition of mTOR and the proteasome. Importantly, mTOR
                      inhibition activated TFEB for a limited duration of
                      1.5 hours, and thereafter the cell population was
                      progressively re-inactivated, with distinct kinetics for
                      Torin1 and nutrient deprivation treatments.TFEB protein
                      levels and subcellular localization are under control of a
                      short-term rheostat, which is highly responsive to negative
                      regulation by mTOR, but under conditions of mTOR inhibition,
                      restricts TFEB activation in a manner dependent on the
                      proteasome. We further identify a long-term,
                      mTOR-independent homeostatic control negatively regulating
                      TFEB upon prolonged mTOR inhibition. These findings are of
                      relevance for developing strategies to target TFEB activity
                      in disease treatment. Moreover, our quantitative approach to
                      decipher phenotype heterogeneity in imaging datasets is of
                      general interest, as shifts between subpopulations provide a
                      quantitative description of single cell behaviour,
                      indicating novel regulatory behaviors and revealing
                      differences between cell types.},
      keywords     = {Basic Helix-Loop-Helix Leucine Zipper Transcription Factors
                      (NLM Chemicals) / TFEB protein, human (NLM Chemicals) / MTOR
                      protein, human (NLM Chemicals) / TOR Serine-Threonine
                      Kinases (NLM Chemicals)},
      cin          = {B190 / A150 / B170},
      ddc          = {610},
      cid          = {I:(DE-He78)B190-20160331 / I:(DE-He78)A150-20160331 /
                      I:(DE-He78)B170-20160331},
      pnm          = {312 - Functional and structural genomics (POF3-312)},
      pid          = {G:(DE-HGF)POF3-312},
      typ          = {PUB:(DE-HGF)16},
      pubmed       = {pmid:27268034},
      pmc          = {pmc:PMC4896000},
      doi          = {10.1186/s12885-016-2388-9},
      url          = {https://inrepo02.dkfz.de/record/131000},
}