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The autophagic protein LC3 translocates to the nucleus and localizes in the nucleolus associated to NUFIP1 in response to cyclic mechanical stress.

Publication ,  Journal Article
Shim, MS; Nettesheim, A; Hirt, J; Liton, PB
Published in: Autophagy
July 2020

UNLABELLED: The trabecular meshwork (TM) is a key regulatory tissue of intraocular pressure (IOP) in the anterior chamber of eye. Dysfunction of the TM causes resistance to outflow of aqueous humor, which in turn leads to elevated IOP, a main risk factor of glaucomatous neurodegeneration. Due to variations in IOP, TM cells are continuously exposed to mechanical deformations. We previously reported activation of macroautophagy/autophagy, as one of the physiological responses elicited in TM cells following mechanical strain application. By using biochemical fractionation analysis and imaging techniques, we demonstrate here for the first time the nuclear accumulation of the autophagic marker MAP1LC3/LC3 (microtubule associated protein1 light chain 3)-II, endogenous and exogenously added (AdGFP-LC3, AdtfLC3), in response to cyclic mechanical stress (CMS). Wheat germ agglutinin (WGA) and leptomycin B treatment suggest LC3 to enter the nucleus by passive diffusion, but to exit in an XPO1/CRM1 (exportin 1)-dependent manner in human TM (hTM) cells. While blockage of nuclear export leads to accumulation of LC3 with promyelocytic leukemia (PML) bodies, nuclear LC3 localizes in the nucleolus in cells under CMS. Moreover, nuclear LC3 co-immunoprecipitated with NUFIP1, a ribosome receptor for starvation-induced ribophagy. More interestingly, we further demonstrate that NUFIP1 translocates from the nucleus to LAMP2 (lysosomal associated membrane protein 2)-positive organelles in the stretched cells without triggering ribophagy, suggesting a more general role of NUFIP1 as a selective autophagy receptor for another yet-to-be-identified target in CMS and a surveillance role of nuclear LC3 against stretch-induced damage. ABBREVIATION: AdGFP: adenovirus encoding GFP; ATG: autophagy-related; BSA: bovine serum albumin; CMS: cyclic mechanical stretch; Co-IP: coimmunoprecipitation; DAPI: 4',6-diamidino-2-phenylindole; DFCs: dense fibrillar components; EM: electron microscopy; FCs: fibrillar centers; GCs: granular components; GFP: green fluorescent protein; hTM: human trabecular meshwork; HBSS: Hanks balanced salt solution; IOP: intraocular pressure; LAMP1/2: lysosomal associated membrane protein 1/2; LepB: leptomycin B; MTOR: mechanistic target of rapamacyin kinase; NES: nuclear export signals; MAP1LC3/LC3: microtubule associated protein 1 light chain 3; NLS: nuclear localization signal; NPCs: nuclear pore complexes; NUFIP1: nuclear FMR1 interacting protein 1; NS: non-stretched; PBS: phosphate-buffered saline; PE: phosphatidylethanolamine; pfu: plaque-forming units; PML: promyelocytic leukemia; RFP: red fluorescent protein; RPS15A: ribosomal protein S15a; RPL26: ribosomal protein L26; rRNA: ribosomal RNA; SIRT1: sirtuin 1; SQSTM1/p62: sequestosome 1; tfLC3: mRFP-GFP tandem fluorescent-tagged LC3; TM: trabecular meshwork; WB: western blot; WDR36: WD repeat domain 36; WGA: wheat germ agglutinin; XPO1/CRM1: exportin 1.

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Published In

Autophagy

DOI

EISSN

1554-8635

Publication Date

July 2020

Volume

16

Issue

7

Start / End Page

1248 / 1261

Location

United States

Related Subject Headings

  • Trabecular Meshwork
  • Stress, Mechanical
  • Sirtuin 1
  • Receptors, Cytoplasmic and Nuclear
  • RNA-Binding Proteins
  • Protein Transport
  • Protein Binding
  • Nuclear Proteins
  • NIH 3T3 Cells
  • Models, Biological
 

Citation

APA
Chicago
ICMJE
MLA
NLM
Shim, M. S., Nettesheim, A., Hirt, J., & Liton, P. B. (2020). The autophagic protein LC3 translocates to the nucleus and localizes in the nucleolus associated to NUFIP1 in response to cyclic mechanical stress. Autophagy, 16(7), 1248–1261. https://doi.org/10.1080/15548627.2019.1662584
Shim, Myoung Sup, April Nettesheim, Joshua Hirt, and Paloma B. Liton. “The autophagic protein LC3 translocates to the nucleus and localizes in the nucleolus associated to NUFIP1 in response to cyclic mechanical stress.Autophagy 16, no. 7 (July 2020): 1248–61. https://doi.org/10.1080/15548627.2019.1662584.
Shim, Myoung Sup, et al. “The autophagic protein LC3 translocates to the nucleus and localizes in the nucleolus associated to NUFIP1 in response to cyclic mechanical stress.Autophagy, vol. 16, no. 7, July 2020, pp. 1248–61. Pubmed, doi:10.1080/15548627.2019.1662584.

Published In

Autophagy

DOI

EISSN

1554-8635

Publication Date

July 2020

Volume

16

Issue

7

Start / End Page

1248 / 1261

Location

United States

Related Subject Headings

  • Trabecular Meshwork
  • Stress, Mechanical
  • Sirtuin 1
  • Receptors, Cytoplasmic and Nuclear
  • RNA-Binding Proteins
  • Protein Transport
  • Protein Binding
  • Nuclear Proteins
  • NIH 3T3 Cells
  • Models, Biological