The Spatial Scale of Synaptic Protein Allocation during Homeostatic Plasticity

Abstract

An individual neuron hosts up to 10,000 individual synapses that can be made stronger or weaker by local and cell-wide plasticity mechanisms, both of which require protein synthesis. To address over what spatial scale a neuron allocates synaptic resources, we quantified the distribution of newly synthesized proteins after global homeostatic upscaling using metabolic labeling and single-molecule localization (DNA-PAINT). Following upscaling, we observed a global increase in locally synthesized nascent protein in synapses and at dendrites, with a high degree of variability between individual synapses. We determined the smallest spatial scale over which nascent proteins were evenly distributed and found that it is best described by synaptic neighborhoods (~ 10 microns in length)-smaller than a dendritic branch and larger than an individual synapse. Protein allocation at the level of neighborhoods thus represents a solution to the problem of protein allocation within a neuron that balances local autonomy and global homeostasis.

Unbiased choice of global clustering parameters for single-molecule localization microscopy

“…In Fig. 5, we show the clustering results for single-molecule localization DNA-PAINT data of newly synthesized proteins after global homeostatic scaling in neuronal dendrites [6] (see Fig. S13 for an analogous analysis for a dataset of neuronal AMPA-receptors).…”

“…Analysis of newly synthesized proteins in neuronal dendrites in DNA-PAINT data [6]. Left: Localizations analyzed using FINDER, CAML (07VEJJ) and CAML (87B144) [31].…”

“…Classical light microscopy can only resolve structural features that are larger than the diffraction limit of light of a few hundred nanometers [3]. By overcoming the diffraction limit, super resolution microscopy has revealed long hidden mechanisms underlying intracellular transport processes [4] and the spatial organization of mRNA translation [5, 6]. The major feature of single-molecule localization microscopy (SMLM) is its ability to exploit the stochastic and sparse switching of fluorescence emission of specific labels binding to target molecules [7].…”

A Picture Worth a Thousand Molecules—Integrative Technologies for Mapping Subcellular Molecular Organization and Plasticity in Developing Circuits

“…Super-resolution microscopy is complementary in that it provides spatial location information about RNA and proteins and can also generate corroborating measurements of target abundance using molecular counting techniques (Sugiyama et al, 2005;Specht et al, 2013;Jungmann et al, 2016;Cella Zanacchi et al, 2019). In support of biochemical evidence for local protein translation in subcellular compartments, superresolution imaging experiments have confirmed the presence of stalled polysomes in neurites (Graber et al, 2017), ribosomes, and mRNAs in a majority of synapses (Younts et al, 2016;Sakers et al, 2017;Hafner et al, 2019) as well as axons, dendrites, and glial processes adjacent to synapses (Chen et al, 2016;Ouwenga et al, 2017;Sakers et al, 2017), and nascent protein production in dendrites and peripheral glial processes (Sakers et al, 2017;Sun et al, 2020;Figure 6). A central challenge is the future integration of spatial transcriptomic and proteomic imaging for high-throughput spatial mapping of mRNAs and proteins together with nanoscale resolution.…”

Pervasive compartment-specific regulation of gene expression during homeostatic synaptic scaling

“…One can speculate that it might allow neurons to maintain excitability while taking into account individual plasticity needs of synapses or dendritic domains 46 . Intriguingly, a very recent study described the local allocation of proteins in so-called "synaptic neighborhoods", about 10 micron long dendritic domains, in a related form of homeostatic plasticity, synaptic upscaling 47 . In agreement with our results, many of these allocated proteins might arise from local dendritic mRNA translation.…”