Collection Citation: Melissa Hoyer, Cristina Capitanio, Ian R. Smith, Julia C. Paoli, Anna Bieber, Yizhi Jiang, Joao A. Paulo, Florian Wilfling, Brenda A. Schulman, Harper JW 2024. Combinatorial selective ER-phagy remodels the ER during neurogenesis. protocols.io https://dx.doi.org/10.17504/protocols.io.81wgbx13nlpk/v3Version created by Melissa Hoyer
Manuscript citation:
Hoyer MJ, Capitanio, C, Smith IR*, Paoli, JC, Bieber, A, Jiang, Y, Paulo JA, Gonzalez-Lozano, MA, Wilfling, F, Schulman, BA, and Harper JW (2023). Combinatorial selective ER-phagy remodels the ER during neurogenesis.
*, equal contribution
License: This is an open access collection distributed under the terms of the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are credited
Protocol status: Working
We use this collection and it's working
Created: February 09, 2024
Last Modified: June 01, 2024
Collection Integer ID: 94913
Keywords: ASAPCRN, ER-phagy, induced neurons, axons, whole cell proteomics, differentiation, live cell fluorescence microscopy, Keima flux, autophagy, er during neurogenesi, axonal er accumulation within synaptic bouton, er shaping protein, quantitative landscape of er proteome remodelling, tubular er membranes within autophagosome, er during cell state transition, axonal er accumulation, endoplasmic reticulum, molecular inventory of er, specific autophagic capture of er, neurogenesi, er proteome remodelling, specific subsets of er membrane, selectivity of er protein clearance, process of neurogenesi, er membrane, formed neuronal projection, er protein clearance, phagy receptor fam134b, role for autophagy, refined tubular er network, deficient neurons in vivo, deficient neuron, synaptic bouton, selective autophagy, neuronal projection, diverse proteome landscape, tubular er membrane, neuron, autophagosome, phagy receptor, phagy receptor mutant, autophagy, protein, tractable induced neuron, embedded er, neuronal projections by
Funders Acknowledgements:
ASAP
Grant ID: ASAP-000282
Abstract
The endoplasmic reticulum (ER) employs a diverse proteome landscape
to orchestrate many cellular functions, ranging from protein and lipid
synthesis to calcium ion flux and inter-organelle communication. A case
in point concerns the process of neurogenesis, where a refined tubular
ER network is assembled via ER shaping proteins into the newly formed
neuronal projections to create highly polarized dendrites and axons.
Previous studies have suggested a role for autophagy in ER remodelling,
as autophagy-deficient neurons in vivo display axonal ER accumulation
within synaptic boutons, and the membrane-embedded ER-phagy receptor
FAM134B has been genetically linked with human sensory and autonomic
neuropathy. However, our understanding of the mechanisms underlying
selective removal of the ER and the role of individual ER-phagy receptors is
limited. Here we combine a genetically tractable induced neuron (iNeuron)
system for monitoring ER remodelling during in vitro differentiation with
proteomic and computational tools to create a quantitative landscape of ER
proteome remodelling via selective autophagy. Through analysis of single
and combinatorial ER-phagy receptor mutants, we delineate the extent to
which each receptor contributes to both the magnitude and selectivity of
ER protein clearance. We define specific subsets of ER membrane or lumenal
proteins as preferred clients for distinct receptors. Using spatial sensors and
flux reporters, we demonstrate receptor-specific autophagic capture of ER
in axons, and directly visualize tubular ER membranes within
autophagosomes in neuronal projections by cryo-electron tomography. This molecular
inventory of ER proteome remodelling and versatile genetic toolkit provide
a quantitative framework for understanding the contributions of individual
ER-phagy receptors for reshaping ER during cell state transitions.
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