Cell "nuclear antenna" cilia-serotonergic axonic synapses drive nuclear signals to alter chromatin accessibility

Written by | Qi

Primary cilia is a microtubule-based membrane-binding compartment that extends a few microns from the matrix into the extracellular space [1].

Fibroidal defects are genetic disorders caused by mutant proteins related to ciliary structure and function, including embryonic and perinatal death to visceral inversion, polydacty, renal cyst formation, obesity, and neurological deficits, almost all of which can be attributed to abnormal
embryonic development.
Although cilia are lost in most terminally differentiated adult skeletal muscles and myocardium, they are still present in most mature neurons and glia of the brain[2], but little
is known about the function of these primary cilia.
It is important to note that the primary cilia in the adult brain are rich in certain G protein-coupled receptors (GPCRs) for neurotransmitters, including dopamine (DA), serotonin, and somatostatin [3
].
It has been shown that the removal of cilia-localized somatostatin receptor 3 (SSTR3) can lead to cognitive impairment of new object recognition without seriously affecting brain development [4
].

To gain insight into the potential function of neuronal primary cilia in the adult brain, David E.
Clapham's team from the Janelia Research Park in the United States published an article titled A serotonergic axon-cilium synapse drives nuclear signaling to alter chromatin in the journal Cell on September 1, 2022 In accessibility's article, they demonstrate the synaptic structure between the brainstem serotonergic axons and the primary cilia of hippocampal CA1 pyramidal neurons, which are rich in cilia-limiting serotonin receptor 5-HTR6, which activates the non-classical G_αq/11-RhoA pathway
that modulates nuclear actin and increases histone acetylation and chromatin accessibility.
As a signaling device close to the nucleus, axis-ciliary synaptic short neurotransmission can alter the epigenetic state
of postsynaptic neurons.


Focused ion beam scanning electron microscopy (FIB-SEM) can be used to reconstruct the microenvironment
of primary cilia of CA1 neurons 。 The researchers tracked two CA1 cilia, with a variable number of microtubule bimodals along their length, and two cilia met on axons containing synaptic vesicles and mitochondria (as shown in Figure 1A below), so are the cilia of vertebral neurons forming specialized contact with the axons, and are these specific sites of nerve transmission? So they collected FIB-SEM datasets of 8 mature mouse hippocampuses, most of which had sites in contact with axons (Figure 1
).
Since these contact-forming axons originated from neurons outside the dataset, the researchers first identified these axons
.
5-HTR6 as a 5-serotonergic GPCR, mainly located in neuronal primary cilia, the authors used endogenous Htr6-EGFP knock-in mouse lines to characterize the location of 5-HTR6 in CA1 pyramidal neurons, labeling pyramidal neuronal cilia with ADCY3 antibody showed that 94% of CA1 pyramidal neuron cilia are 5-HTR6+, while labeling axons with anti-serotonin transporter (SERT) antibodies, 35% of cilia are immediately adjacent to axons, And all axon sites juxtaposed with cilia are synaptogen-positive, indicating that these sites are serotonin release sites
.

Figure 1.
FIB-SEM reveals axon-ciliary synapses
.

Ultrastructural analysis provides anatomical evidence of axon-ciliary synapses, so will activation of serotonergic axons release serotonin onto cilia? To this end, the authors designed a cilia-targeted serotonin sensor based on a GPCR activation (GRAB) strategy and used a 5-HTR6 receptor as a scaffold (GRAB-HTR6-PM
).
Subsequently, they expressed the sensor and ChrimsonR (photochannel protein) in hippocampus and serotonergic neurons, respectively, and after giving 1 Hz photostimulation, the release of serotonin into cilia in contact with serotonergic axonal synapses could be detected
.
Next, the authors want to know about the "follow-up events"
after the release.
Based on previous work, the authors speculate that serotonin may activate the G_αq/11-Trio-RhoA pathway
.
The results showed that 10 nM serotonin stimulation can increase RhoA activity in neuronal cilia in 5-15 minutes, and if 5-HTR6 blockers or G_αq/11 inhibitors are added, this activation effect can be largely eliminated, indicating that serotonin stimulation leads to_{Gαq/11-dependent} RhoA activation
in cilia.

Figure 2.
Serotonin-stimulating ciliary HTR6 activates RhoA in cilia
.

RhoA activation can phosphorylate adductor proteins and increase its affinity for F-actin by Rho-associated kinase, while changes in nuclear actin alter the chromatin landscape [5] directly bind to and regulate the activity of the acetyl transferase KAT14, which significantly acetylates H4K5, so the authors hypothesize that stimulation of the 5-HTR6 receptor may modulate H4K5ac
in the hippocampus.
By injecting serotonin agonists, the proportion of H4K5ac increased significantly, which would not be caused if Htr6 KO mice were also administered
.
Histone acetylation is associated with increased access to chromatin, ATAC-see is a technique that uses highly active transposase mutants with fluorescently labeled oligonucleotides to label accessible chromatin in monolayer cells [6], and the injection of serotonin agonists can significantly increase the ATAC-see marker in fixed goose mouse brain sections, that is, the accessibility of chromatin increases, and if the activity of ciliary RhoA is inhibited, accessibility
is reduced.
Together, these data suggest that ciliary 5-HTR6 signaling controls chromatin remodeling via
the G_αq/11-Trio-RhoA pathway.

In summary, the main findings of this work are that axons release neurotransmitters onto axon-ciliary synapses and induce restrictive signals that differ from plasma membrane signals to be transmitted to the nucleus, while raising the possibility that primary cilia act as an epigenetic regulator to stabilize transcriptional programming in response to environmental cues
.
In this "cilia as nuclear antenna" model, cilia provide a protected compartment for shorter, more direct encoding receptor binding to regulate nuclear transcription
.

Original link:

https://doi.
org/10.
1016/j.
cell.
2022.
07.
026

Plate Maker: Eleven

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