• Home
  • Search Results
  • Selective autophagy of cytosolic protein aggregates involves ribosome-free rough endoplasmic reticulum.

Selective autophagy of cytosolic protein aggregates involves ribosome-free rough endoplasmic reticulum.

Histochemistry and cell biology (2019-11-14)
Sujin Park, Christian Zuber, Jürgen Roth
ABSTRACT

Autophagy is a degradative cellular process that can be both non-selective and selective and begins with the formation of a unique smooth double-membrane phagophore which wraps around a portion of the cytoplasm. Excess and damaged organelles and cytoplasmic protein aggregates are degraded by selective autophagy. Previously, we reported that in fed HepG2 cells, cytoplasmic aggregates of EDEM1 and surplus fibrinogen Aα-γ assembly intermediates are targets of selective autophagy receptors and become degraded by a selective autophagy called aggrephagy. Here, we show by multiple confocal immunofluorescence and colocalization panels the codistribution of cytoplasmic protein aggregates with the selective autophagy receptors p62/SQSTM1 and NBR1 and with the phagophore marker LC3, and that phagophores induced by vinblastine treatment contain complexes of protein aggregates and selective autophagy receptors. By combined serial ultrathin section analysis and immunoelectron microscopy, we found that in fed HepG2 cells, a basically ribosome-free subdomain of rough endoplasmic reticulum (RER) cisternae forms a cradle that engulfs the cytoplasmic protein aggregates. This RER subdomain appears structurally different from omegasomes formed by the RER, which were suggested to provide a membrane platform from which the phagophore is derived in starvation-induced autophagy. Taken together, our observations provide further evidence for the importance of RER subdomains as a site and membrane source for phagophore formation and show their involvement in selective autophagy.

MATERIALS
Product Number
Brand
Product Description

Sigma-Aldrich
3,3′-Diaminobenzidine tetrahydrochloride, tablet, 10 mg substrate per tablet
Sigma-Aldrich
Saponin Quillaja sp., Sapogenin content 20-35 %
Sigma-Aldrich
Vinblastine sulfate salt, ≥97% (HPLC)
Sigma-Aldrich
Glutaraldehyde, 25% Aqueous Solution
Sigma-Aldrich
Bovine Serum Albumin, lyophilized powder, γ-irradiated, Globulin Free, BioXtra, suitable for cell culture
Sigma-Aldrich
Paraformaldehyde, for synthesis
Sigma-Aldrich
Epoxy embedding medium, for microscopy
Sigma-Aldrich
DL-Cysteine, technical grade