Stuck in a lockdown: Dreams, bad dreams, nightmares, and their relationship to stress, depression and anxiety during the COVID-19 pandemic.

BACKGROUND: An upsurge in dream and nightmare frequency has been noted since the beginning of the COVID-19 pandemic and research shows increases in levels of stress, depression and anxiety during this time. Growing evidence suggests that dream content has a bi-directional relationship with psychopathology, and that dreams react to new, personally significant and emotional experiences. The first lockdown experience was an acute event, characterized by a combination of several unprecedent factors (new pandemic, threat of disease, global uncertainty, the experience of social isolation and exposure to stressful information) that resulted in a large-scale disruption of life routines. This study aimed at investigating changes in dream, bad dream and nightmare recall; most prevalent dream themes; and the relationship between dreams, bad dreams, nightmares and symptoms of stress, depression and anxiety during the first COVID-19 lockdown (April-May 2020) through a national online survey. METHODS: 968 participants completed an online survey. Dream themes were measured using the Typical Dreams Questionnaire; stress levels were measured by the Cohen's Perceived Stress Scale; symptoms of anxiety were assessed by Generalized Anxiety Disorder (GAD-7) scale; and symptoms of depression were assessed using the Quick Inventory of Depressive Symptomatology. RESULTS: 34% (328) of participants reported increased dream recall during the lockdown. The most common dream themes were centered around the topics of 1) inefficacy (e.g., trying again and again, arriving late), 2) human threat (e.g., being chased, attacked); 3) death; and 4) pandemic imagery (e.g., being separated from loved ones, being sick). Dream, bad dream and nightmare frequency was highest in individuals with moderate to severe stress levels. Frequency of bad dreams, nightmares, and dreams about the pandemic, inefficacy, and death were associated with higher levels of stress, as well as with greater symptoms of depression and anxiety. CONCLUSIONS: Results support theories of dream formation, environmental susceptibility and stress reactivity. Dream content during the lockdown broadly reflected existential concerns and was associated with increased symptoms of mental health indices.

Dynamics of Auxilin 1 and GAK in clathrin-mediated traffic.

Clathrin-coated vesicles lose their clathrin lattice within seconds of pinching off, through the action of the Hsc70 "uncoating ATPase." The J- and PTEN-like domain-containing proteins, auxilin 1 (Aux1) and auxilin 2 (GAK), recruit Hsc70. The PTEN-like domain has no phosphatase activity, but it can recognize phosphatidylinositol phosphate head groups. Aux1 and GAK appear on coated vesicles in successive transient bursts, immediately after dynamin-mediated membrane scission has released the vesicle from the plasma membrane. These bursts contain a very small number of auxilins, and even four to six molecules are sufficient to mediate uncoating. In contrast, we could not detect auxilins in abortive pits or at any time during coated pit assembly. We previously showed that clathrin-coated vesicles have a dynamic phosphoinositide landscape, and we have proposed that lipid head group recognition might determine the timing of Aux1 and GAK appearance. The differential recruitment of Aux1 and GAK correlates with temporal variations in phosphoinositide composition, consistent with a lipid-switch timing mechanism.

L1 endocytosis is controlled by a phosphorylation-dephosphorylation cycle stimulated by outside-in signaling by L1.

Dynamic regulation of the cell surface expression of adhesion molecules is an important mechanism for controlling neuronal growth cone motility and guidance. Clathrin-mediated vesicular internalization of L1 via the tyrosine-based endocytosis motif YRSL regulates adhesion and signaling by this Ig superfamily molecule. Here, we present evidence that tyrosine-1176 (Y1176) of the YRSL motif is phosphorylated in vivo. The nonreceptor tyrosine kinase (p60src) is implicated in L1-mediated neurite outgrowth, and we find that p60src phosphorylates Y1176 in vitro. Phosphorylation of Y1176 prevents L1 binding to AP-2, an adaptor required for clathrin-mediated internalization of L1. mAb 74-5H7 recognizes the sequence immediately NH2-terminal to the tyrosine-based motif and binds L1 only when Y1176 is dephosphorylated. 74-5H7 identifies a subset of L1 present at points of cell-cell contact and in vesicle-like structures that colocalize with an endocytosis marker. L1-L1 binding or L1 cross-linking induces a rapid increase in 74-5H7 immunoreactivity. Our data suggest a model in which homophilic binding or L1 cross-linking triggers transient dephosphorylation of the YRSL motif that makes L1 available for endocytosis. Thus, the regulation of L1 endocytosis through dephosphorylation of Y1176 is a critical regulatory point of L1-mediated adhesion and signaling.

Reconstitution of Clathrin Coat Disassembly for Fluorescence Microscopy and Single-Molecule Analysis.

The disassembly of the clathrin lattice surrounding coated vesicles is the obligatory last step in their life cycle. It is mediated by the coordinated recruitment of auxilin and Hsc70, an ATP-driven molecular clamp. Here, we describe the preparation of reagents and the single-particle fluorescence microscopy imaging assay in which we visualize directly the Hsc70-driven uncoating of synthetic clathrin coats or clathrin-coated vesicles.

Key interactions for clathrin coat stability.

Clathrin-coated vesicles are major carriers of vesicular traffic in eukaryotic cells. This endocytic pathway relies on cycles of clathrin coat assembly and Hsc70-mediated disassembly. Here we identify histidine residues as major determinants of lattice assembly and stability. They are located at the invariant interface between the proximal and distal segments of clathrin heavy chains, in triskelions centered on two adjacent vertices of the coated-vesicle lattice. Mutation of these histidine residues to glutamine alters the pH dependence of coat stability. We then describe single-particle fluorescence imaging experiments in which we follow the effect of these histidine mutations on susceptibility to Hsc70-dependent uncoating. Coats destabilized by these mutations require fewer Hsc70 molecules to initiate disassembly, as predicted by a model in which Hsc70 traps conformational distortions during the auxilin- and Hsc70:ATP-mediated uncoating reaction.

A motif in the clathrin heavy chain required for the Hsc70/auxilin uncoating reaction.

The 70-kDa heat-shock cognate protein (Hsc70) chaperone is an ATP-dependent "disassembly enzyme" for many subcellular structures, including clathrin-coated vesicles where it functions as an uncoating ATPase. Hsc70, and its cochaperone auxilin together catalyze coat disassembly. Like other members of the Hsp70 chaperone family, it is thought that ATP-bound Hsc70 recognizes the clathrin triskelion through an unfolded exposed hydrophobic segment. The best candidate is the unstructured C terminus (residues 1631-1675) of the heavy chain at the foot of the tripod below the hub, containing the sequence motif QLMLT, closely related to the sequence bound preferentially by the substrate groove of Hsc70 (Fotin et al., 2004b). To test this hypothesis, we generated in insect cells recombinant mammalian triskelions that in vitro form clathrin cages and clathrin/AP-2 coats exactly like those assembled from native clathrin. We show that coats assembled from recombinant clathrin are good substrates for ATP- and auxilin-dependent, Hsc70-catalyzed uncoating. Finally, we show that this uncoating reaction proceeds normally when the coats contain recombinant heavy chains truncated C-terminal to the QLMLT motif, but very inefficiently when the motif is absent. Thus, the QLMLT motif is required for Hsc-70-facilitated uncoating, consistent with the proposal that this sequence is a specific target of the chaperone.

The mu2 subunit of the clathrin adaptor AP-2 binds to FDNPVY and YppØ sorting signals at distinct sites.

The endocytic sorting signal on the low-density lipoprotein receptor for clathrin-mediated internalization is the sequence FDNPVY in the receptor's cytosolic tail. We have used a combination of surface plasmon resonance and crosslinking with a photoactivated peptide probe to demonstrate the interaction between FDNPVY-containing peptides and the mu2 chain of purified AP-2 clathrin adaptors (the complexes responsible for plasma membrane sorting). We show that recognition of the FDNPVY signal is mediated by a binding site in the mu2-subunit that is distinct from the site for the more general YppØ sorting signal, another tyrosine-based sequence also recognized by mu2-adaptin. These results suggest the possibility that low-density lipoprotein receptor uptake may be modulated specifically and independently of other proteins in the clathrin pathway.