Konno, T. | Search bioRxiv

bioRxiv2021

Endoplasmic Reticulum morphological regulation by RTN4/NOGO modulates neuronal regeneration by slowing luminal transport

Cell and tissue functions rely on an elaborate intracellular transport system responsible for distributing bioactive molecules with high spatiotemporal accuracy. The tubular network of the Endoplasmic Reticulum (ER) constitutes a system for the delivery of luminal solutes it stores, including Ca2+, across the cell periphery. The physical nature and factors underlying the ERs functioning as a fluidics system are unclear. Using an improved ER transport visualisation methodology combined with optogenetic Ca2+ dynamics imaging, we observed that ER luminal transport is modulated by natural ER tubule narrowing and dilation, directly proportional to the amount of an ER membrane morphogen, Reticulon 4 (RTN4). Consequently, the ER morphoregulatory effect of RTN4 defines ERs capacity for peripheral Ca2+ delivery and thus controls axonogenesis. Excess RTN4 limited ER luminal transport, Ca2+ release and iPSC-derived cortical neurons axonal extension, while RTN4 elimination reversed the effects. SummaryIntracellular transport through the lumen of the ER network is modulated through narrowing/dilation of ER tubules by a membrane morphogen - RTN4, a process controlling axonogenesis by limiting the delivery of ER-stored Ca2+.

cell biology↗

bioRxiv2021

Stress-induced protein disaggregation in the Endoplasmic Reticulum catalysed by BiP

Protein synthesis is supported by cellular machineries that ensure polypeptides fold to their native three-dimensional conformation with high fidelity whilst eliminating misfolded, aggregation-prone species. While protein aggregates can contribute to pathologies exemplified by Alzheimers and Parkinsons diseases, their abundance is normally minimised by molecular chaperones such as HSP70/90, which promote native folding and drive the recycling of aberrantly folded species. Cytoplasmic chaperones (e.g. HSP70/40) can resolve insoluble protein aggregates should the preventive mechanisms falter. However, it is unknown whether an analogous disaggregation system is needed and exists in the Endoplasmic Reticulum (ER), where [~]30% of the proteome is synthesised. Here we show that the ER of a variety of mammalian cell types, including neurons, is endowed with the capability to resolve protein aggregates that accumulate upon expression of metastable proteins. Utilising a purpose-developed protein aggregation probing system with a sub-organellar resolution, we observed steady-state aggregate accumulation in the ER. Strikingly, pharmacological induction of ER stress did not augment aggregates but rather stimulated their clearance within hours. We found that this disaggregation activity was catalysed by the stress-responsive ER molecular chaperone - BiP. Its elimination abolished the disaggregation activity in cells. Further, we reconstructed the disaggregation-reaction in-vitro by a minimal system of ATP-fuelled BiP and its J-protein cofactor. These data reveal a hitherto unknown, non-redundant function of the ER stress response. Thus, our findings may facilitate the identification of aggregation-antagonising strategies and rationalising the age-dependent protein misfolding pathology. SummaryER stress induction activates a protein disaggregation machinery, powered by BiP - an abundant ER chaperone, revealed by a FLIM-based protein aggregation monitoring in live cells.

molecular biology↗