Learning and Sleep Have Divergent Effects on Cytosolic and Membrane-Associated Ribosomal mRNA Profiles in Hippocampal Neurons

Abstract

The hippocampus plays an essential role in consolidating transient experiences into long-lasting memories. Memory consolidation can be facilitated by post-learning sleep, although the underlying cellular mechanisms are undefined. Here, we addressed this question using a mouse model of hippocampally-mediated, sleep-dependent memory consolidation (contextual fear memory; CFM), which is known to be disrupted by post-learning sleep loss. We used translating ribosome affinity purification (TRAP) to quantify ribosome-associated RNAs in different subcellular compartments (cytosol and membrane) and in different hippocampal cell populations (either whole hippocampus, Camk2a+ excitatory neurons, or highly active neurons expressing phosphorylated ribosomal subunit S6 [pS6+]). Using RNA-seq, we examined how these transcript profiles change as a function of sleep vs. sleep deprivation (SD) and as a function of prior learning (contextual fear conditioning; CFC). Surprisingly, we found that while many mRNAs on cytosolic ribosomes were altered by sleep loss, almost none were altered by learning. Of the few changes in cytosolic ribosomal transcript abundance following CFC, almost all were occluded by subsequent SD. This effect was particularly pronounced in pS6+ neurons with the highest level of neuronal activity following CFC, suggesting SD-induced disruption of post-learning transcript changes in putative “engram” neurons. In striking contrast, far fewer transcripts on membranebound (MB) ribosomes were altered by SD, and many more mRNAs (and lncRNAs) were altered on MB ribosomes as a function of prior learning. For hippocampal neurons, cellular pathways most significantly affected by CFC were involved in structural remodeling. Comparisons of post-CFC transcript profiles between freely-sleeping and SD mice implicated changes in cellular metabolism in Camk2a+ neurons, and increased protein synthesis capacity in pS6+ neurons, as biological processes disrupted by post-learning sleep loss.

Sleep and Memory Consolidation Dysfunction in Psychiatric Disorders: Evidence for the Involvement of Extracellular Matrix Molecules

“…Decreased PNN composition during sleep (Pantazopoulos et al, 2020a) coincides with reduced LTP amplitude during the light phase in the hippocampus (Chaudhury et al, 2005). Further evidence for the involvement of ECM molecules including CSPGs in memory consolidation during sleep comes from a recent single cell gene expression profiling study describing altered gene expression of CSPG synthesis and degradation pathways in CamKIIα neurons when animals that slept normally were compared to sleep deprived animals following fear conditioning (Delorme J. et al, 2020).…”

Are there roles for heterogeneous ribosomes during sleep in the rodent brain?

“…Sleep deprivation primarily affected mRNA translated in the cytosol, while learning mainly altered transcripts on membrane-bound ribosomes, suggesting a first level of translational specificity within neurons. Importantly, sleep after learning showed increased translation of membrane-bound transcripts, including mRNAs of 13 RPs, such as Rps27 and Rps28, with ~50% of the RPs located at the ribosome surface (Figure 2B) (Delorme et al, 2021). These changes were specifically allocated to sleep as RP mRNA expression did not increase if sleep was prevented after learning.…”

“…(B) Heatmap (Morpheus, https://software.broadinstitute.org/morpheus) representing heterogeneous expression of selected RPs in neurons from in vivo studies in rodents or in vitro studies. The colour bar depicts log (2) fold changes (FC) of the particular conditions (indicated on the left) against control condition values (see D and (Rozenbaum et al, 2018;Lyons et al, 2020;Delorme et al, 2021;Fusco et al, 2021) for experimental details). From top to bottom: 1) 13 RP transcripts on membrane-bound ribosomes in activated (pS6+) neurons in the hippocampus following contextual fear conditioning and subsequent sleep in mice (Delorme et al, 2021).…”

“…The colour bar depicts log (2) fold changes (FC) of the particular conditions (indicated on the left) against control condition values (see D and (Rozenbaum et al, 2018;Lyons et al, 2020;Delorme et al, 2021;Fusco et al, 2021) for experimental details). From top to bottom: 1) 13 RP transcripts on membrane-bound ribosomes in activated (pS6+) neurons in the hippocampus following contextual fear conditioning and subsequent sleep in mice (Delorme et al, 2021). 2) 7 RP mRNAs affected by acute (5 h) sleep deprivation in excitatory neurons (CamKIIα+) in the mouse hippocampus (Lyons et al, 2020).…”

“…Besides global and specific control through signalling pathways and RBPs/ncRNAs, respectively, whether the formation of specialised ribosomes could also contribute to selected translation during sleep remains unclear. However, sleep-dependent translational changes were examined within neurons in the hippocampus after learning in mice (Delorme et al, 2021) (Figure 2C). Using TRAP, ribosome-associated transcripts were identified from different subcellular fractions of neurons (i.e., cytosolic vs. membrane-associated ribosomes).…”