- Intermediate filaments associate with aggresome-like structures in proteostressed C. elegans neurons and influence large vesicle extrusions as exophers

IFD proteins can be extruded in exophers a IFD-1-positive puncta extruded in an exopher in a strain that expresses bzIs166[Pmec-4mCherry] for touch neuron/exopher visualization. Shown is an adult touch neuron from a strain expressing bzIs166[Pmec-4mCherry]; bzSi3[Pmec-7GFP::IFD-1]. Soma outline is in white, nucleus is outlined with a white-dashed line. One GFP::IFD-1 focus remains in the soma, and one is ejected in the exopher. Scale bar = 2 um, representative of N > 10 GFP::IFD-1+ exophers. b We examined a strain expressing bzIs166[Pmec-4mCherry]; bzSi3[Pmec-7GFP::IFD-1] on Ad3 and determined how often GFP::IFD-1 was extruded in exophers. In total, we observed 61 ALM Ad3 exopher events, with 11 having GFP::IFD-1 included in the exopher, 3 trials, error bar is SEM. c We looked at neurons that ejected a GFP:IFD-1. Of 9 neurons in which GFP::IFD-1 was expelled in the exopher, 4 neurons ejected the only – and entire - GFP::IFD-1 organelle visibly present. Graphed as GFP::IFD-1 diameter in arbitrary units. d Shown is an adult touch neuron from a strain expressing bzSi39[Pmec7 HttQ74::mNG]; bzSi34[Pmec-7mScarlet::IFD-1] on Ad1 after a 6 hr fast. Soma outline is in white, nucleus is outlined with a white dashed line; arrowhead points to the concentrated HttQ74 punctum that colocalizes to the IFD-1 domain. Scale bar = 2 µm, representative of N > 10 exophers. e We examined a strain expressing bzSi39[Pmec7HttQ74::mNG]; bzSi34[Pmec-7mScarlet::IFD-1] on Ad1 after a 6 hr fast to increase exopher yields, and asked how often mScarlet::IFD-1 was extruded in exophers. We observed 30 ALM exopher events, with 14 having mSc::IFD-1 included in the exopher. Next we asked how often the 14 mScarlet::IFD-1-positive exopher events also included at least one concentrated HttQ74 puncta overlapping with mSc::IFD-1. In 11/14 ALM IFD-1-positive exophers, there was at least one mNG::HttQ74 punctum colocalized with mSc::IFD-1. f We examined the same exopher events as described in (e) above. We determined how often HttQ74 aggregates were extruded in exophers. Out of N = 30 ALM exopher events, 22 exopher events had at least one HttQ74 aggregate included in the exopher. We asked how often mNG::HttQ74 positive exopher events (N = 22) included species where HttQ74 is a cargo of the mSc-aggresome-like organelle to find 50% (11/22) of ALM HttQ74-positive exophers had at least one incidence of HttQ74 including mSc::IFD-1.