Diverse physiological and therapeutic insults that increase the amount of unfolded or misfolded proteins in the endoplasmic reticulum (ER) induce the unfolded protein response, an evolutionarily conserved protective mechanism that manages ER stress. Glucose-regulated protein 78/immunoglobulin heavy-chain binding protein (GRP78/BiP) is an ER-resident protein that plays a central role in the ER stress response and is the only known substrate of the proteolytic A subunit (SubA) of a novel bacterial AB(5) toxin. Here, we report that an engineered fusion protein, epidermal growth factor (EGF)-SubA, combining EGF and SubA, is highly toxic to growing and confluent epidermal growth factor receptor-expressing cancer cells, and its cytotoxicity is mediated by a remarkably rapid cleavage of GRP78/BiP. Systemic delivery of EGF-SubA results in a significant inhibition of human breast and prostate tumor xenografts in mouse models. Furthermore, EGF-SubA dramatically increases the sensitivity of cancer cells to the ER stress-inducing drug thapsigargin, and vice versa, demonstrating the first example of mechanism-based synergism in the action of a cytotoxin and an ER-targeting drug.
The function of Ric-3, which is required for nicotinic acetylcholine receptor (nAChR) expression in C. elegans, is unclear. Here we found that Ric-3 can promote or inhibit cell-surface delivery of ␣-bungarotoxin-binding nAChRs (BgtRs) composed of ␣7 subunits. At low levels, Ric-3 promoted BgtR assembly, endoplasmic reticulum (ER) release, and cell-surface delivery without trafficking from the ER. At high Ric-3 levels, Ric-3 suppressed BgtR surface delivery, but not its assembly, and BgtRs were retained in the ER or in Ric-3-containing aggregates. In PC12 cells, native BgtRs trafficked to the cell surface from the ER where low levels of endogenous Ric-3 were observed. In cultured neurons, native Ric-3 levels were higher than in PC12 cells, and Ric-3 and ␣7 subunits were found in somata and dendrites, but not axons, of inhibitory interneurons. Ric-3 trafficked with ␣7 subunits in rapidly moving vesicles to dendrites, where it was restricted to the ER subcompartment. We conclude that Ric-3 has two potential functions. At low levels, Ric-3 interactions are short-lived and promote BgtR assembly and ER release. At higher levels, Ric-3 interactions are longer-lived and mediate ER retention. In neurons, Ric-3 ER retention appears to promote transport within the dendritic ER subcompartment, thereby restricting ␣7 trafficking to dendrites and preventing axonal transport.
Crystal structures of racemic and
homochiral forms of 2-phenylbutyramide
(1) and 3-methyl-3-phenylpyrrolidine-2,5-dione (2) were investigated in detail by a single crystal X-ray diffraction
study. Absolute configurations of the homochiral forms of 1 and 2, obtained by chromatographic separation of racemates,
were determined. It was revealed that racemate and homochiral forms
of 1 are very similar in terms of supramolecular organization
(H-bonded ribbons) in crystal, infrared (IR) spectral characteristics,
and melting points. The presence of two different molecular conformations
in homochiral forms of 1 allowed mimicking of crystal
packing of the H-bonded ribbons in racemate 1. Two polymorph
modifications (monoclinic and orthorhombic) comprising very similar
H-bonded zigzag-like chains were found for the homochiral
forms of compound 2 that were significantly different
in terms of crystal structure, IR spectra, and melting points from
the racemic form of 2. Unlike compound 1, homochiral forms of compound 2 have a higher density
than the corresponding racemate which contradicts the Wallach rule
and indicates that, in this case, homochiral forms are more stable
than racemate forms.
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