Introduction
It has been known for decades that
C18-C21 steroids are ligands to nuclear
receptors and since the 1990’s that oxysterols are ligands to
“orphan”, now “adopted”, nuclear receptors (Evans & Mangelsdorf,
2014). Importantly, nuclear receptors are expressed in brain where
oxysterols are also abundant (Warner & Gustafsson, 2015). These
receptors work via regulating gene and hence protein expression.N -methyl-D-aspartate receptors (NMDARs) are expressed in nerve
cells and work on a much shorter time scale. They are ligand-gated ion
channels activated by the neurotransmitter glutamate, critical to the
regulation of excitatory synaptic function. NMDARs are modulated by
excitatory neurosteroids and by the neuro-oxysterol
24S-hydroxycholesterol (24S-HC, Figure 1, see Supporting Information
Table S1 for systematic and common names and their abbreviations) (Paul
et al., 2013).
For simplicity, in this article we use the term oxysterol to cover
oxidised forms of cholesterol and its precursors (Javitt, 2008;
Schroepfer, 2000), and use the term sterol to specifically embrace
cholesterol and its cyclic precursors. By extension neuro-oxysterols and
neuro-sterols are the respective terms to define oxysterols and sterols
found in the central nervous system (CNS). The sterol definition is at
variance with the formal definition of sterols by Lipid Maps to includeall molecules based on the cyclopentanoperhydrophenanthrene
skeleton and ring-opened versions thereof (Fahy et al., 2005).
G protein-coupled receptors (GPCR) are cell membrane receptors that are
activated by molecules outside the cell and activate signal transduction
pathways within the cell. Cell membranes are rich in cholesterol and
membrane cholesterol plays an important role in GPCR structure and
function (Sengupta & Chattopadhyay, 2015). Several cholesterol
interaction sites have been identified in GPCRs whose occupancy may
modulate GPCR activity. Smoothened (SMO), a member of the Frizzled-class
of GPCRs and a critical component of the hedgehog (Hh) signalling
pathway, has a cholesterol binding site within its extracellular
cysteine rich domain (CRD), mutational modification of which impairs the
ability of SMO to transmit Hh signals (Byrne et al., 2016). The Hh
pathway is critical for tissue patterning during development and
abnormal function is associated with birth defects and cancer (McCabe &
Leahy, 2015). Defective Hh signalling is implicated in Smith-Lemli-Opitz
syndrome (SLOS) which presents, not only with dysmorphology, but also
learning and behavioural problems, highlighting the importance of Hh
signalling in brain (Cooper et al., 2003; Kelley et al., 1996). Numerous
oxysterols have also been found to activate the Hh signalling pathway by
binding to the CRD of SMO (Nedelcu, Liu, Xu, Jao & Salic, 2013; Qi,
Liu, Thompson, McDonald, Zhang & Li, 2019; Raleigh et al., 2018), many
of these are found in brain and are hence neuro-oxysterols (Yutuc et
al., 2020). GPCR183, also known as Epstein Barr Virus Induced GPCR or
EBI2, is involved in the trafficking of immune cells towards their EBI2
ligands i.e. oxysterols with a dihydroxycholesterol (diHC) structure,
where one additional hydroxy group is at C-7α and the second on the
side-chain (Hannedouche et al., 2011; Liu et al., 2011). As T-cell
trafficking plays a major role in the neurodegenerative autoimmune
disease multiple sclerosis and in its animal model, experimental
autoimmune encephalitis (EAE), GPCR183 and neuro-oxysterols have been
studied in this regard (Duc, Vigne & Pot, 2019).
7α,25-Dihydroxycholesterol (7α,25-diHC) and
7α,(25R)26-dihydroxycholesterol (7α,(25R)26-diHC, also known as
7α,27-dihdroxycholesterol, note, if stereochemistry at C25 is not
indicated it is assumed to be 25R) are both GPCR183 agonists present in
brain (Griffiths et al., 2019a) and 7α,25-diHC has been found to be
increased in spinal cord during EAE development (Wanke et al., 2017).
All cells in vertebrates express the enzymatic machinery to synthesise
cholesterol. The cholesterol biosynthesis pathway, also known as the
mevalonate pathway, (see
http://www.lipidmaps.org/pathways/pathway_lipids_list.php for
exact details) is regulated by the master transcription factor SREBP-2
(sterol regulatory element-binding protein 2) which also regulates the
expression of the low density lipoprotein receptor (LDLR) (Horton,
Goldstein & Brown, 2002). When cholesterol levels are low, SREBP-2 is
transported by SCAP (SREBP cleavage activating protein) from the
endoplasmic reticulum to the Golgi where it is processed to its active
form which translocates to the nucleus and activates target gene
transcription, hence up-regulating cholesterol biosynthesis and import
via LDLRs and restoring cholesterol levels in the cell (Goldstein,
DeBose-Boyd & Brown, 2006). When cholesterol levels are elevated,
cholesterol in the endoplasmic reticulum binds to SCAP which becomes
tethered to the resident endoplasmic reticulum protein INSIG (insulin
induced gene) and prevents transport of SREBP-2 to the Golgi for
activation (Goldstein, DeBose-Boyd & Brown, 2006; Sun, Seemann,
Goldstein & Brown, 2007). In this way cholesterol regulates its own
biosynthesis and import via the LDLR. Oxysterols also inhibit
cholesterol biosynthesis, but in this case, by binding to INSIG and
tethering the SCAP - SREBP-2 complex in the Golgi (Radhakrishnan, Ikeda,
Kwon, Brown & Goldstein, 2007). Although the turnover of cholesterol in
the adult brain is slow (0.4% per day in mouse, 0.03% in human) the
rate of synthesis is much higher during development (Dietschy & Turley,
2004). In the foetal mouse, the blood-brain barrier (BBB) is formed at
E10 - E11 and after E11 - 12 the brain is the source of essentially all
new sterols (Tint, Yu, Shang, Xu & Patel, 2006). Importantly,
cholesterol 24-hydroxylase (cytochrome P450 46A1, CYP46A1) expression is
low until E18, conserving cholesterol and restricting the biosynthesis
of the neuro-oxysterol 24S-HC (Tint, Yu, Shang, Xu & Patel, 2006).
CYP46A1 is the dominant cholesterol hydroxylase in brain and
hydroxylation of cholesterol to 24S-HC is responsible for 50 - 60% of
all cholesterol metabolism in adult brain (Russell, Halford, Ramirez,
Shah & Kotti, 2009).
Other receptors for oxysterols include cytoplasmic oxysterol-binding
protein (OSBP) and a family of proteins showing sequence homology to
OSBP called OSBP-related (ORP) or OSBP-like (OSBPL) (Olkkonen &
Hynynen, 2009). OSBP regulates lipid transport between the endoplasmic
reticulum and Golgi and also can act as a sterol-dependent scaffold for
protein phosphatases that dephosphorylate ERK (Olkkonen & Hynynen,
2009).