Dysregulated astrocyte cholesterol synthesis in Huntington's disease: A potential intersection with other cellular dysfunctions

Abstract

Astrocytes are key elements for synapse development and function. Several astrocytic dysfunctions contribute to the pathophysiology of various neurodegenerative disorders, including Huntington's disease (HD), an autosomal-dominant neurodegenerative disorder that is characterized by motor and cognitive defects with behavioral/psychiatric disturbances. One dysfunction in HD related to astrocytes is reduced cholesterol synthesis, leading to a decreased availability of local cholesterol for synaptic activity. This review describes the specific role of astrocytes in the brain local cholesterol synthesis and presents evidence supporting a defective astrocyte-neuron cholesterol crosstalk in HD, by focusing on SREBP-2, the transcription factor that regulates the majority of genes involved in the cholesterol biosynthetic pathway. The emerging coordination of SREBP-2 with other physiological processes, such as energy metabolism, autophagy, and Sonic Hedgehog signaling, is also discussed. Finally, this review intends to stimulate future research directions to explore whether the impairment of astrocytic SREBP-2-mediated cholesterol synthesis in HD associates with other cellular dysfunctions in the disease.

Keywords

cholesterol astrocyte synapse SREBP-2 energy metabolism autophagy Sonic Hedgehog

Introduction

Neuroglial cells were identified in 1846 by Rudolf Virchow, who described these cells as a homogenous population that generally supports neuronal function.¹ Later, in 1893, Michael von Lenhossék coined the term “astrocyte” to describe the star-shaped cells found in the central nervous system (CNS).²

Thanks to advances in knowledge over the past few decades, astrocytes are now defined as highly specialized and sophisticated cells with diverse functions, including ion homeostasis, synapse formation, elimination, and neurotransmitter release.³ Astrocytes also supply metabolic and neurotrophic support and regulate blood flow and blood-brain barrier (BBB) permeability.⁴ Astrocytes differ amongst different brain regions, and morphologically and molecularly different astrocytes may coexist with specialized roles in synaptic activity as key elements of the tripartite synapse within the same brain region.⁵ The intricate arborization and anatomical specialization reflect the functional complexity of these cells. In the mouse striatum, for instance, each astrocyte, within its territory, has ∼11 MSNs and about 51,000 excitatory synapses.⁶ Human astrocytes are larger than mouse astrocytes, and each human astrocyte surrounds ∼ 2 million excitatory synapses within its territory,^7,8 suggesting a more sophisticated role of human astrocytes in brain complexity.

Astrocytes are widely implicated in several neurodegenerative diseases, including Huntington's disease (HD), an autosomal-dominant neurodegenerative disorder that is characterized by motor and cognitive defects with behavioral/psychiatric disturbances appearing during midlife, and caused by a CAG-repeat expansion in the gene encoding the huntingtin protein (HTT).⁹ An exhaustive description of the astrocytic dysfunctions associated with HD pathophysiology can be found in a recent excellent review.¹⁰ A non-cell autonomous mechanism occurring in the astrocytes and contributing to HD phenotypes is related to the reduced capability to produce cholesterol within the brain in the presence of mutant HTT (mHTT). Extensive studies carried out in several HD mouse models over the past 20 years highlight the relevance of this dysfunction in the disease.^11–16

Cholesterol in healthy and Huntington's disease brain

In mammals, ∼25% of total body cholesterol resides in the brain.¹⁷ Most of the brain cholesterol (70–80%) is found in myelin and produced during post-natal myelination. Even though the adult brain only produces a small amount of cholesterol, it is crucial for brain function since peripheral cholesterol cannot pass across the BBB.¹⁸

Cholesterol synthesis is a complex and energy-intensive process. Across the tissues, the biosynthetic pathway starts with the conversion of acetyl-CoA to 3-hydroxy-3-methylglutaryl-CoA (HMG-CoA), which is then converted to mevalonate (Figure 1). By means of enzymatic processes, mevalonate is converted into 3-isopentenyl pyrophosphate, farnesyl pyrophosphate, squalene and lanosterol. After lanosterol, two parallel and interlinked processes, named Kandutsch-Russell (KR) and Bloch pathways respectively, can be used to produce cholesterol (Figure 1). Studies carried out in in vitro cell cultures suggest that neurons preferentially use the KR pathway, whereas astrocytes the Bloch pathway.¹⁹ However, experimental proof of the cell-specific use of these pathways in neurons and astrocytes in vivo is still lacking.

Figure 1.

Cholesterol biosynthesis pathway.

Astrocytes are considered the main neural cells producing cholesterol, which is then transported as apolipoprotein E (ApoE)-containing lipoprotein particles to neurons to regulate neuronal function and synaptic activity²⁰ (Figure 2). At the synapse, the astrocyte-derived cholesterol plays different roles supporting the structure and function of synaptic vesicles and lipid rafts; in particular, at the pre-synaptic level, cholesterol regulates the optimal release of neurotransmitters, while, in post-synaptic terminals is important for receptor-mediated intracellular signaling (Figure 2). While astrocytes represent the main source of cholesterol, neurons are mainly responsible for its elimination from the brain.²¹ Indeed, the neuronal Cytochrome P450 Family 46 Subfamily A Member 1 (CYP46A1) hydroxylates cholesterol at carbon-24 by to produce the hydrophilic catabolite 24(S)-hydroxycholesterol (24S-OHC), which crosses the BBB and is subsequently metabolized in the liver.^22,23 Notably, 24S-OHC is also a potent allosteric NMDA modulator in mouse hippocampus slices.²⁴ The synthesis of new cholesterol (in astrocytes) and the secretion of 24S-OHC (in neurons) are finely regulated and closely coupled as suggested by the evidence that in mice the genetic deletion of cyp46a1 leads to reduced catabolism and, in turn, cholesterol synthesis within the brain.²⁵

Figure 2.

Astrocyte-derived cholesterol at healthy and HD tripartite synapse.

Although the molecular mechanisms regulating this crosstalk have not been fully elucidated, the disruption of this balance contributes – in common or disease-specific ways - to the development or progression of several neurological diseases, including not only HD but also Alzheimer's disease, Parkinson's disease, and neurodevelopmental disorders.²⁶ Cholesterol is therefore frequently implicated in brain disorders, but how it is determinant is still often unclear for some of these diseases.

Several studies in HD rodent models demonstrate that brain cholesterol biosynthesis is decreased,^12–16 and the striatum is the first region to be affected.¹⁶ This dysfunction occurs before the onset of motor symptoms,^12–14 and is CAG repeat length-dependent.¹⁶ The early feature of this dysfunction was first demonstrated by earlier gene expression data in inducible striatal-like HD cells and in postmortem caudate of grade-I HD patients where a reduced expression of key cholesterol biosynthesis genes was demonstrated.^11,12 Further validation was obtained by measuring the in vivo rate of cholesterol synthesis with deuterium-labeled water in different brain regions of the heterozygous knock-in zQ175 mouse model during disease progression.¹⁶ In this mouse model, which recapitulates genetically and phenotypically the human disease, both cholesterol precursors and the rate of cholesterol synthesis are significantly reduced in the striatum at 5 weeks of age, several weeks before any other disease changes.¹⁶ Notably, several pre-clinical trials demonstrated the efficacy of different cholesterol-increasing strategies in the brain to improve cognitive performance and other disease-associated phenotypes in HD mouse,^27–31 emphasizing the extensive rationale and evidence in support of brain cholesterol therapies for HD.^32,33

Beyond the defective cholesterol biosynthesis, which occurs mainly in astrocytes,^29,34 neuronal cholesterol catabolism is also affected.^15,35–37 Indeed, pre-clinical studies based on gene therapy approaches targeting either cholesterol biosynthesis in astrocytes²⁹ or cholesterol catabolism in neurons^35,36 have proven equally effective at recovering many disease-associated phenotypes in HD mouse models. Moreover, a clinical trial is ongoing to test the safety of a gene therapy targeting brain cholesterol metabolism (AB-1001) in HD patients.³⁸

Further evidence of impaired cholesterol metabolism/homeostasis in HD comes from measuring the plasmatic level of 24S-OHC in HD patients. As 24S-OHC in human plasma is almost entirely of brain origin,²² its plasma levels may reflect cholesterol synthesis/catabolism in the brain. Decreased levels of plasma 24S-OHC have been described in HD patients in several cross-sectional studies.^39–42 A longitudinal study is ongoing to test the validity of 24S-OHC as a candidate biomarker in HD patients⁴³ that can be used in future clinical trials focused on cholesterol-based strategies.^32,33 Of note, noninvasive technologies testing a CYP46A1-targeted positron emission tomography (PET) tracer have been recently developed to assess brain cholesterol homeostasis in the brain of living animal models and patients,⁴⁴ allowing the acceleration of the clinical translation of cholesterol-based strategies in HD.

Astrocyte-neuron cholesterol crosstalk

According to the Tripartite Synapse concept, astrocytes are integral elements in the synaptic function,⁴⁵ and astrocyte-derived cholesterol is an active player in this activity.²⁰

Cholesterol biosynthesis is supposed to be a common pathway among the different types of astrocytes.⁴⁶ A different demand for cholesterol could make specific brain regions and circuits more susceptible to disrupted astrocyte-neuron cholesterol coordination. For example, the striatum, which collects and integrates information from several brain areas and funnels them forward to reach the basal ganglia output structures,⁴⁷ may require robust cholesterol supply. Unfortunately, at present, it is still not possible to quantify in vivo the demand for cholesterol at the single-cell level, and the hypothesis that the striatum is more dependent on cholesterol than other brain regions is still unproven. However, in support of this hypothesis, cholesterol content is higher in the striatum compared to the cortex in different mammalian brains at physiological conditions.^{13,14,16,48–50} Interestingly, two different populations of astrocytes that selectively interact with MSN belonging to the main striatal circuits – the direct and indirect pathways – have been identified,⁵¹ suggesting a high specific functional interaction between astrocytes and synapses in this region.⁵² Whether the different striatal circuits require varying amounts of astrocyte-derived cholesterol, and whether this is critical for the function and dysfunction of the striatum, is still unknown but it is certainly worth investigating.

The hub role of SREBP-2 in the cholesterol biosynthesis pathway

The promoter regions of almost all genes involved in cholesterol biosynthesis contain a DNA element called the sterol regulatory element (SRE), which is recognized by the master regulator Sterol regulatory-element binding protein 2 (SREBP-2), a basic-helix-loop-helix-leucine zipper protein, firstly identified in 1993 by Goldstein & Brown.⁵³

Three closely related isoforms of SREBPs (SREBP-1a, SREBP-1c, and SREBP-2) exist in mammals, and SREBP-2 selectively regulates the genes encoding enzymes involved in cholesterol biosynthesis. SREBP-2 is indispensable for embryonic development, and SREBP-2-deficient mice die in utero with limb bud malformations,⁵⁴ probably because of impaired Hedgehog signaling.

SREBPs are synthesized as inactive membrane proteins inserted into the endoplasmic reticulum (ER). The activation of SREBPs requires their transportation to the Golgi apparatus by the SREBP Cleavage-Activating Protein (SCAP), an ER sterol-sensing protein. In this compartment, SREBPs are subjected to two-step proteolytic processing by two membrane-bound transcription factor Site-1 Proteases (S1P and S2P) to obtain an N-terminal fragment that enters the nucleus, binds to SRE sequences, and stimulates the transcription of target genes. The specificity of the SREBP isoforms for different target genes is partly explained by unique binding affinity to cis-elements, primarily SREs, of the cholesterol biosynthesis genes as well as to SREs and enhancer boxes (E-boxes) in lipogenic genes, together with cofactors such as SP1 and NFY.⁵⁵ SREBP-2 can be subjected to several post-translational modifications, such as acetylation,⁵⁶ phosphorylation, and sumoylation,^57,58 which further regulate its transcriptional activity.

The critical role of astrocytic SREBP-2 for neuronal activity and cognitive processes has been explored through conditional knock-out mice targeting SREBP-2, SCAP, or key cholesterol biosynthesis genes. SREBP-2 deletion in GFAP-expressing astrocytes in mice causes a loss of cholesterol production, substantially reduces brain size, and causes abnormal performance in learning and memory tasks.⁵⁹ Deletion of SCAP - upstream of SREBPs - in astrocytes results in a more severe phenotype with progressive motor defects with immature synapses, reduced number of synaptic vesicles, and impaired synaptic function.^60,61 Similarly, conditional knock-out mice of the neuron-specific LDL Receptor-related Protein 1 (LRP1), essential for glial cholesterol uptake by neurons, show neurodegeneration, motor impairment, and synaptic defects.⁶² To further support the role of astrocyte-derived cholesterol biosynthesis, knockout of the fdft1 gene, encoding the squalene synthase—the enzyme that catalyzes a key step in cholesterol biosynthesis—in adult mouse neurons does not affect neuronal survival and function in mice.⁶³

Reduced cholesterol biosynthesis in HD: focus on SREBP-2 in astrocytes

In vitro and in vivo studies demonstrated that the nuclear level of the active form of SREBP-2 is reduced in HD cells,^12,28,29,34 leading to a decreased transcription of cholesterol biosynthesis genes,^12,29,64,65 likely due to the sequestration in the cytoplasm of the SREBP-2/importin b complex required for nuclear import.⁶⁶ This dysfunction has been also described in HD astrocytes, and it is detrimental to several cholesterol-dependent functions in neurons at pre-synaptic and post-synaptic terminals (Figure 2).³⁴ Primary astrocytes from HD mice and neural-stem-derived astrocytes expressing mHTT show reduced cholesterol biosynthesis and reduced secretion of ApoE-bound cholesterol in the medium in a CAG-dependent manner.^15,34 Conditioned medium collected from HD astrocytes and lipoprotein-depleted conditioned medium from healthy astrocytes are not able to promote neurite outgrowth and do not support synaptic activity in HD neurons. This synaptic impairment does not occur when conditioned medium from wt astrocytes or cholesterol supplementation are used.³⁴

The top pathways altered in HD astrocytes are related to cholesterol biosynthesis and degradation.^67,68 However, the suppression of genes involved in cholesterol biosynthesis is more evident in mouse and human glial cells expressing the exon-1 of mHTT rather than full-length mHTT.⁶⁸ Further studies are therefore needed to decipher how the exon-1 and the full-length mHTT impact on this astrocytic metabolic process. Furthermore, a recent bulk RNA sequencing in IPSC-derived HD astrocytes with a large range of polyQ lengths (at an early differentiation time point) revealed that cholesterol secretion is significantly increased in 45Q–58Q astrocytes, unaffected in 69–81Q astrocytes and down-regulated in 125Q and 180Q astrocytes.⁶⁹ These findings suggest that human astrocytes bearing a low pathological range of polyQ (45Q–58Q) may undergo compensatory mechanisms as an early homeostatic response. As the disease burden accumulates, this response may weak, as occurs in 125Q-180Q astrocytes.

Overall, in-vitro and in-vivo findings indicate that the astrocytic cholesterol dysregulation is relevant in HD and therefore it should be considered as potential therapeutic target. In co-culture in-vitro systems, genetic perturbation of SREBP-2 in wt astrocytes has detrimental effects on HD neurons, while the over-expression of the N-terminal fragment of SREBP-2 in HD astrocytes reverses neurite outgrowth and synaptic defects in HD neurons.³⁴ In vivo, the forced expression of the N-terminal of human SREBP-2 (hSREBP-2) in striatal astrocytes of the R6/2 HD mouse model - by using a recombinant adeno-associated virus 2/5 (AAV2/5) - significantly enhances endogenous cholesterol biosynthesis in the striatum and normalizes the synaptic transmission of both inhibitory and excitatory synapses, increases the number of striatal MSNs expressing dopamine-D2 receptor, counteracts mHTT aggregation, and rescues motor defects and cognitive decline.²⁹ All these findings highlight that striatal injection of AAV2/5-hSREBP-2 modifies core features of the disease, at least in this mouse model characterized by an early disease onset and aggressive phenotype.

Further studies using IPSC-derived human astrocytes, in parallel with the advancement of astrocytic differentiation protocols to mimic the complexity of human astrocytes in vitro, will help to better identify the substantial role of astrocytic cholesterol/SREBP-2 activity in HD pathogenesis.

SREBP-2: a potential node of convergence with other cellular processes in health and HD?

Over the last decades, new regulatory mechanisms and unexpected transcriptional targets have expanded the roles of SREBPs as metabolic integrators in cellular homeostasis beyond lipid/cholesterol synthesis, at least in peripheral tissues.^70–72 However, because of the complexity and cell heterogeneity of cholesterol and its synthesis in the brain, whether and how these novel roles are present in various neural cells remain an open challenge. Moreover, SREBPs frequently cooperate with additional transcription factors, providing a mechanism for tissue-specific, SREBP-dependent gene expression that could be exclusive to the brain.

The critical role of cholesterol at the synapse and how defective astrocyte-derived cholesterol/SREBP-2 activity in HD negatively affects synapses in vivo have been pointed out in a recent comprehensive review.³² The next paragraph provides examples of integrated and bidirectional signals mediated by SREBP-2 in physiology, and how a defective astrocyte SREBP-2 activity in HD could contribute to other metabolic and cellular processes altered in the disease (Figure 3).

Figure 3.

The convergence roles of SREBP-2 with other cellular processes.

SREBP-2, mitochondria, and energy metabolism

The brain is one of the most metabolically active organs in the body but lacks fuel stores, requiring a continuous supply of energy substrates to maintain a proper balance between excitatory and inhibitory activity.⁷³ Through cellular metabolic pathways, astrocytes play a critical role in preserving brain energy balance and supporting neuronal function. Notably, striatal MSNs have high energy requirements due to their dendritic complexity and high synaptic activity in intricate circuits,⁷⁴ which may make them more vulnerable to mitochondrial/energy damage. Nevertheless, the molecular mechanisms underlying these defects are not fully elucidated and the relevance of these abnormalities in early HD remains elusive.

Glucose is the primary energy substrate for the brain, providing the necessary energy for neural activity, synaptic transmission, and cellular maintenance. Glucose is transported into the brain through the BBB via specialized transporters, e.g., Glucose Transporter Type 1 (GLUT1). Glucose metabolism and cholesterol synthesis are interconnected. Physiologically, glucose metabolism through glycolysis produces pyruvate, which can be reduced to produce lactate or used to generate acetyl-CoA, a key substrate for cholesterol synthesis. Abnormalities in these pathways influence acetyl-CoA availability, which may affect the balance between energy metabolism and lipid/cholesterol synthesis. The impact of such defects could be more relevant in astrocytes, which express high levels of GLUT1 and produce and supply both glycolytic lactate and cholesterol to neurons for their activities,^20,75 with negative consequences on HD neurons (Figure 3). Of note, mice with srebp2 knock-out, specifically in astrocytes, have impaired brain development, behavioral defects, and impaired glucose metabolism.⁵⁹ Moreover, in PET imaging studies, progressive striatal and cortical glucose hypometabolism has been observed in HD patients,^76–78 and GLUT1 expression is reduced in cortical tissues and fibroblasts from pediatric HD patients.⁷⁹

A further connection between SREBP-2 and energy metabolism comes from a recent work reporting cholesterol synthesis depends on glutamine sensing.⁸⁰ Although the study was carried out only in vitro, it shows that glutamine is required for the transport-dependent proteolytic activation of SREBP-2, and chronic mitochondrial dysfunction dysregulates glutamine uptake and alters cholesterol levels.⁸⁰ Notably, a system-wide analysis performed in the R6/2 mouse spatial proteome revealed a defective astrocytic glutamate-GABA-glutamine cycling, causing impaired glutamine release and, consequently, GABA synthesis (Figure 3).⁸¹

Lastly, together with other lipids, cholesterol may contribute to the structure of mitochondrial membranes, thereby influencing mitochondrial efficiency, for example, via the formation of lipid rafts in mitochondrial membranes, which are necessary for the functioning of mitochondrial proteins involved in energy metabolism and ATP synthesis.^82,83 Reduced SREBP-2 activity in the HD astrocytes may contribute to the structural abnormalities in mitochondrial morphology observed in postmortem cortical tissue from HD patients,⁸⁴ or mitochondria dynamics and function.⁸⁵ At the molecular level, mHTT impairs the activity of Peroxisome Proliferator-Activated Receptor-γ Coactivator 1-alpha (PPARGC1-alpha or PGC-1-alpha), the transcriptional coactivator regulating the expression of genes involved in mitochondrial biogenesis and respiration. In line with these findings, PGC-1 knock-out mice exhibit mitochondrial defects accompanied by striatal degeneration.⁸⁶ These mice also show decreased brain cholesterol and its precursors and reduced expression of key cholesterol biosynthesis genes. In cellular models, the over-expression of PGC-1-alpha increases SREBP-2 promoter activity,⁸⁷ further suggesting a link between energy metabolism and cholesterol synthesis at the transcriptional level.

Future investigations in HD astrocytes are needed to explore the in vivo connection between dysfunctional cholesterol synthesis/SREBP-2 activity and defects in energy metabolism via multiple molecular mechanisms.

SREBP-2 and autophagy

In 2011, the research group headed by T. Osborne, through a genome-wide localization of SREBP-2 in hepatic chromatin, predicted a role of SREBP-2 in autophagy.⁸⁸ These authors demonstrated that SREBP-2 directly activates autophagy genes during cell-sterol depletion. Additionally, they also showed that SREBP-2 knockdown during nutrient depletion decreases autophagosome formation and lipid droplet association of the autophagosome targeting Microtubule-associated protein 1A/1B-light chain 3 (LC3) protein,⁸⁸ suggesting that SREBP-2 plays a role in regulating autophagic processes under certain conditions.

Recent in-vitro findings proposed that autophagy is regulated by SREBP-2/cholesterol via Mechanistic Target Of Rapamycin Complex 1 (mTORC1), a key protein in regulating autophagy.⁸⁹ In particular, mTORC1 is activated by SLC38A9, a lysosomal transmembrane protein containing a cholesterol-binding domain. When the cholesterol content in lysosomes increases, SLC38A9 binds cholesterol, leading to a conformational change of the protein and allowing the recruitment and activation of mTORC1 in the lysosome (Figure 3).⁸⁹

Other in-vitro studies showed that activated mTORC1 triggers the translocation of SREBP-2 to the Golgi where SREBP-2 is cleaved to translocate to the nucleus and where it activates expression of the genes involved in cholesterol synthesis.⁹⁰ mTORC1, through its regulation of autophagy and endosomal membrane trafficking, regulates cholesterol trafficking to lysosomes and thereby modulates ER cholesterol. This process controls SREBP-2 activation and transcriptional activity.⁹¹ All these findings argue for a bidirectional coordination between SREBP-2 and mTORC1.

Notably, mTORC1 activity is reduced in the striatum of HD mice (i.e., N171-82Q mouse model by assessed the ratio of the phosphitylated and total S6 Ribosomal Protein (pS6/S6) by western blot).⁶⁵ The over-expression of Ras Homolog Enriched in Brain protein (RHEB) in the striatum of HD mice potentiates mTORC1 activity and enhances pathways implicated in mHTT clearance and the mRNA levels of SREBP-2-dependent cholesterogenic genes.⁶⁵ Additionally, all cholesterol-based strategies tested so far in HD mouse models, aimed at supplying exogenous cholesterol to the brain or enhancing endogenous cholesterol synthesis in striatal astrocytes, successfully counteract mHTT aggregates,^27–31 further supporting a link between astrocytic SREBP-2 activity/cholesterol with clearance pathways.

Despite some indirect in-vivo data, the majority of the findings regarding the bidirectional cooperation between SREBP-2 and mTORC1 were collected from cultured cell models. It therefore remains to be determined whether the regulatory pathways between SREBP-2 and mTORC1 also occur in vivo, possibly in different brain regions, and/or cell types, and how this bidirectional interplay influences astrocytic and neuronal function in health and disease progression. Further research into the interaction between astrocytic SREBP-2 and autophagic pathways could eventually help identifying the molecular mechanisms underlying the capability of cholesterol-based strategies to counteract mHTT aggregation. This approach will also likely favor the development of new therapeutic avenues for HD and other neurodegenerative disorders where cholesterol metabolism and autophagy are both disrupted.

SREBP-2 and Sonic Hedgehog signaling

The Sonic Hedgehog (SHH) signaling is a key regulator of cell differentiation, proliferation, and tissue patterning during development.⁹² Cholesterol can covalently modify SHH (Figure 3), and this modification is required for proper localization of SHH to the cell membrane, where it can interact with its receptors Patched and Smoothened.⁹³ Indeed, human syndromes caused by defects in the final stages of cholesterol biosynthesis are characterized by a defective response to SHH signaling,⁹⁴ and holoprosencephaly, a severe brain malformation caused by abnormal SHH signaling transduction, is due to the defects of cholesterol biosynthesis.⁹⁵ Furthermore, the lack of SREBP-2 in the developing mouse embryos leads to abnormal regulation of some genes in the SHH signaling axis involved in limb bud morphogenesis,⁵⁴ indicating a functional crosstalk between SREBP-2 and SHH during brain development.

Reduced cholesterol availability in pathological conditions may alter the SHH signaling with different tissue- and cell-specific consequences. Of note, emerging studies have identified the SHH signaling pathway as an essential regulator of molecular identity and functional features of astrocytes during development; this pathway is therefore essential to correctly shape circuit assembly and proper function.^96,97

A piece of the puzzle that could link the reduced cholesterol accessibility in the HD brain with SHH is related to primary cilia - single, non-motile signaling organelles found on the surface of most mammalian cells – that are essential for SHH signal transduction. Primary cilia are critical for the synapses of newborn neurons,⁹⁸ and defects in primary cilia lead to the shortening of dendrites that fail to integrate into mature circuits;^97,98 this evidence suggests that the cilium-related SHH pathway is essential for a correct brain functional development. Of note, mice with primary ciliary-deficient astrocytes show behavioral deficits in sensorimotor function, sociability, learning, and memory.⁹⁷ Both primary cilia and SHH signaling are impaired in neurodegenerative and neurodevelopmental diseases,^99,100 and it has been reported that primary cilia formation and structure are defective in HD.^101,102 However, further studies are needed to explore whether there is a bidirectional connection between the cilium-related SHH pathway and cholesterol and if reduced availability of astrocytic cholesterol in HD influences the cilium-related SHH defects. Insights in this direction could reveal unexpected SREBP-2-dependent mechanisms that contribute to abnormal brain development in HD and increased vulnerability to degeneration later in life.

Conclusions

Cholesterol biosynthesis is an energetically expensive process that requires significant supply of acetyl-CoA, ATP, oxygen, and the reducing co-factors NADPH and NADH. Accordingly, it must be tightly regulated at multiple levels. In the brain, this regulation is mediated by a crosstalk signaling between astrocytes and neurons. Reduced SREBP-2 activity in HD astrocytes impairs this crosstalk with negative consequences for neuronal and synaptic activity.

Understanding the complexity of cholesterol biology in the mammalian brain is one of the upcoming challenges. The current models and theories about SREBP-2 regulation are mostly based on work on peripheral tissues, but additional molecular processes controlling SREBP-2 activity and cholesterol synthesis in the brain could also be at work. Expanding the knowledge of cell-specific contribution of SREBP-2 in different types of neurons, astrocytes, and other glial cells is indeed essential to better understand the association of cholesterol and the complexity of the brain including neurodegenerative disorders, such as HD. New cutting-edge technologies are urgently needed to decipher the cellular heterogeneity, diverse cellular coordination, and multiple SREBP-2-dependent molecular mechanisms of brain cholesterol in vivo.

SREBP-2 could also be a potential node of convergence amongst global biological signaling networks involved in various brain critical physiological processes. Whether and how abnormal cholesterol metabolism also influences or responds to additional signals in disease processes requires further and extensive investigations to connect the dots in the integrated pathophysiology of HD.

Footnotes

Acknowledgments

I would like to thank all the HD families because their hope fuels and inspires all researchers to bring progress and improve the lives of those who are affected by the disease. I also thank Prof. Mirko Baruscotti for his valuable advice and insightful discussions.

ORCID iD

Marta Valenza

Funding

The author(s) disclosed receipt of the following financial support for the research, authorship, and/or publication of this article: This work was supported by the Telethon Foundation (GMR22T1079) to MV, and by the Italian Ministry of Universities and Research (MIUR), Research Projects of National Interest (G53D23004420006) to MV.

Conflicting interests

The author(s) declared no potential conflicts of interest with respect to the research, authorship, and/or publication of this article.

References

Chvátal

Verkhratsky

. An early history of neuroglial research: personalities. Neuroglia 2018; 16: 245–257.

Lenhossek

. Der feinere Bau des Nervensystems im Lichte neuester Forschungen. 1895.

Haim

Rowitch

. Functional diversity of astrocytes in neural circuit regulation. Nat Rev Neurosci 2016; 18: 311–321.

Santello

Toni

Volterra

. Astrocyte function from information processing to cognition and cognitive impairment. Nat Neurosci 2019; 22: 154–166.

Khakh

Deneen

. The emerging nature of astrocyte diversity. Annu Rev Neurosci 2019; 42: 187–207.

Endo

Kasai

Soto

, et al. Molecular basis of astrocyte diversity and morphology across the CNS in health and disease. Science 2022; 378: eabn8461.

Oberheim

Goldman

Nedergaard

. Heterogeneity of astrocytic form and function. Methods Mol Biol 2012; 814: 23–45.

Oberheim

Takano

Han

, et al. Uniquely hominid features of adult human astrocytes. J Neurosci 2009; 29: 3276–3287.

Zuccato

Valenza

Cattaneo

. Molecular mechanisms and potential therapeutic targets in Huntington’s disease. Physiol Rev 2010; 90: 905–981.

10.

Khakh

Goldman

. Astrocytic contributions to Huntington’s disease pathophysiology. Ann N Y Acad Sci 2023; 1522: 42–59.

11.

Sipione

Rigamonti

Valenza

, et al. Early transcriptional profiles in huntingtin-inducible striatal cells by microarray analyses. Hum Mol Genet 2002; 11: 1953–1965.

12.

Valenza

Rigamonti

Goffredo

, et al. Dysfunction of the cholesterol biosynthetic pathway in Huntington’s disease. J Neurosci 2005; 25: 9932–9939.

13.

Valenza

Leoni

Tarditi

, et al. Progressive dysfunction of the cholesterol biosynthesis pathway in the R6/2 mouse model of Huntington’s disease. Neurobiol Dis 2007; 28: 133–142.

14.

Valenza

Carroll

Leoni

, et al. Cholesterol biosynthesis pathway is disturbed in YAC128 mice and is modulated by huntingtin mutation. Hum Mol Genet 2007; 16: 2187–2198.

15.

Valenza

Leoni

Karasinska

, et al. Cholesterol defect is marked across multiple rodent models of Huntington’s disease and is manifest in astrocytes. J Neurosci 2010; 30: 10844–10850.

16.

Shankaran

Di Paolo

Leoni

, et al. Early and brain region-specific decrease of de novo cholesterol biosynthesis in Huntington’s disease: a cross-validation study in Q175 knock-in mice. Neurobiol Dis 2017; 98: 25–38.

17.

Dietschy

Turley

. Cholesterol metabolism in the central nervous system during early development and in the mature animal. J Lipid Res 2004; 45: 1375–1397.

18.

Jurevics

Morell

. Cholesterol for synthesis of myelin is made locally, not imported into brain. J Neurochem 1995; 64: 895–901.

19.

Nieweg

Schaller

Pfrieger

. Marked differences in cholesterol synthesis between neurons and glial cells from postnatal rats. J Neurochem 2009; 109: 125–134.

20.

Mauch

Nägier

Schumacher

, et al. CNS Synaptogenesis promoted by glia-derived cholesterol. Science 2001; 294: 1354–1357.

21.

Lütjohann

Breuer

Ahlborg

, et al. Cholesterol homeostasis in human brain: evidence for an age-dependent flux of 24S-hydroxycholesterol from the brain into the circulation. Proc Natl Acad Sci U S A 1996; 93: 9787–9791.

22.

Björkhem

Lütjohann

Diczfalusy

, et al. Cholesterol homeostasis in human brain: turnover of 24S-hydroxycholesterol and evidence for a cerebral origin of most of this oxysterol in the circulation. J Lipid Res 1998; 39: 1594–1600.

23.

Paul

Doherty

Robichaud

, et al. The major brain cholesterol metabolite 24(S)-hydroxycholesterol is a potent allosteric modulator of N-Methyl-D-Aspartate receptors. J Neurosci 2013; 33: 17290–17302.

24.

Sun

Linsenbardt

Emnett

, et al. 24(S)-hydroxycholesterol As a modulator of neuronal signaling and survival. Neuroscientist 2016; 22: 132–152.

25.

Lund

Xie

Kotti

, et al. Knockout of the cholesterol 24-hydroxylase gene in mice reveals a brain-specific mechanism of cholesterol turnover. J Biol Chem 2003; 278: 22980–22986.

26.

Martín

Pfrieger

Dotti

. Cholesterol in brain disease: sometimes determinant and frequently implicated. EMBO Rep 2014; 15: 1036–1052.

27.

Valenza

Chen

Di Paolo

, et al. Cholesterol-loaded nanoparticles ameliorate synaptic and cognitive function in Huntington’s disease mice. EMBO Mol Med 2015; 7: 1547–1564.

28.

Birolini

Valenza

Di Paolo

, et al. Striatal infusion of cholesterol promotes dose-dependent behavioral benefits and exerts disease-modifying effects in Huntington’s disease mice. EMBO Mol Med 2020; 12: e12619.

29.

Birolini

Verlengia

Talpo

, et al. SREBP2 Gene therapy targeting striatal astrocytes ameliorates Huntington’s disease phenotypes. Brain 2021; 144: 3175–3190.

30.

Birolini

Valenza

Ottonelli

, et al. Insights into kinetics, release, and behavioral effects of brain-targeted hybrid nanoparticles for cholesterol delivery in Huntington’s disease. J Control Release 2021; 330: 587–598.

31.

Birolini

Valenza

Ottonelli

, et al. Chronic cholesterol administration to the brain supports complete and long-lasting cognitive and motor amelioration in Huntington’s disease. Pharmacol Res 2023; 194: 106823.

32.

Valenza

Birolini

Cattaneo

. The translational potential of cholesterol-based therapies for neurological disease. Nat Rev Neurol 2023; 19: 583–598.

33.

Cattaneo

Barker

. Brain cholesterol therapy for Huntington’s disease—does it make sense? Clin Transl Med 2024; 14: e1746.

34.

Valenza

Marullo

Di Paolo

, et al. Disruption of astrocyte-neuron cholesterol cross talk affects neuronal function in Huntington’s disease. Cell Death Differ 2015; 22: 708–722.

35.

Boussicault

Alves

Lamazière

, et al. CYP46A1, The rate-limiting enzyme for cholesterol degradation, is neuroprotective in Huntington’s disease. Brain 2016; 139: 953–970.

36.

Kacher

Lamazière

Heck

, et al. CYP46A1 Gene therapy deciphers the role of brain cholesterol metabolism in Huntington’s disease. Brain 2019; 142: 2432–2450.

37.

Kreilaus

Spiro

McLean

, et al. Evidence for altered cholesterol metabolism in Huntington’s disease post mortem brain tissue. Neuropathol Appl Neurobiol 2016; 42: 535–546.

38.

National Institutes of Health. A study to evaluate AB-1001 striatal administration in adults with early manifest Huntington's disease. ClinicalTrials.gov Identifier: NCT05541627.

39.

Leoni

Mariotti

Tabrizi

, et al. Plasma 24S-hydroxycholesterol and caudate MRI in pre-manifest and early Huntington’s disease. Brain 2008; 131: 2851–2859.

40.

Leoni

Long

Mills

, et al. Plasma 24S-hydroxycholesterol correlation with markers of Huntington disease progression. Neurobiol Dis 2013; 55: 37–43.

41.

Leoni

Mariotti

Nanetti

, et al. Whole body cholesterol metabolism is impaired in Huntington’s disease. Neurosci Lett 2011; 494: 245–249.

42.

Gray

Dai

Smith

, et al. Changes in 24(S)-hydroxycholesterol are associated with cognitive performance in early Huntington’s disease: data from the TRACK and ENROLL HD cohorts. J Huntingtons Dis 2024; 13: 449–465.

43.

National Institutes of Health. Clinical study to monitor Plasma levels of 24OHC in subject with HD (Chol-HD). ClinicalTrials.gov Identifier: NCT04257513.

44.

Haider

Zhao

Wang

, et al. Assessment of cholesterol homeostasis in the living human brain. Sci Transl Med 2022; 14: eadc9967.

45.

Perea

Navarrete

Araque

. Tripartite synapses: astrocytes process and control synaptic information. Trends Neurosci 2009; 32: 421–431.

46.

Batiuk

Martirosyan

Wahis

, et al. Identification of region-specific astrocyte subtypes at single cell resolution. Nat Commun 2020; 11: 1220.

47.

Nelson

Kreitzer

. Reassessing models of basal ganglia function and dysfunction. Annu Rev Neurosci 2014; 37: 117–135.

48.

Runquist

Parmryd

Chojnacki

, et al. Distribution of branch point prenyltransferases in regions of bovine brain. J Neurochem 1995; 65: 2299–2306.

49.

Zhang

Appelkvist

Kristensson

, et al. The lipid compositions of different regions of rat brain during development and aging. Neurobiol Aging 1996; 17: 869–875.

50.

Turley

Burns

Dietschy

. Preferential utilization of newly synthesized cholesterol for brain growth in neonatal lambs. Am J Physiol 1998; 274: E1099–E1105.

51.

Martín

Bajo-Grañeras

Moratalla

, et al. Circuit-specific signaling in astrocyte-neuron networks in basal ganglia pathways. Science 2015; 349: 730–734.

52.

Durkee

Araque

. Diversity and specificity of astrocyte–neuron communication. Neuroscience 2019; 396: 73–78.

53.

Hua

Yokoyama

, et al. SREBP-2, a second basic-helix-loop-helix-leucine zipper protein that stimulates transcription by binding to a sterol regulatory element. Proc Natl Acad Sci U S A 1993; 90: 11603–11607.

54.

Vergnes

Chin

De Vallim

, et al. SREBP-2-deficient and hypomorphic mice reveal roles for SREBP-2 in embryonic development and SREBP-1c expression. J Lipid Res 2016; 57: 410–421.

55.

Horton

Goldstein

Brown

. SREBPs: transcriptional mediators of lipid homeostasis. Cold Spring Harb Symp Quant Biol 2002; 67: 491–498.

56.

Giandomenico

Simonsson

Grünroos

, et al. Coactivator-dependent acetylation stabilizes members of the SREBP family of transcription factors. Mol Cell Biol 2003; 23: 2587–2599.

57.

Sundqvist

Bengoechea-Alonso

, et al. Control of lipid metabolism by phosphorylation-dependent degradation of the SREBP family of transcription factors by SCFFbw7. Cell Metab 2005; 1: 379–391.

58.

Hirano

Murata

Tanaka

, et al. Sterol regulatory element-binding proteins are negatively regulated through SUMO-1 modification independent of the ubiquitin/26 S proteasome pathway. J Biol Chem 2003; 278: 16809–16819.

59.

Ferris

Perry

Moreira

, et al. Loss of astrocyte cholesterol synthesis disrupts neuronal function and alters whole-body metabolism. Proc Natl Acad Sci U S A 2017; 114: 1189–1194.

60.

Camargo

Brouwers

Loos

, et al. High-fat diet ameliorates neurological deficits caused by defective astrocyte lipid metabolism. FASEB J 2012; 26: 4302–4315.

61.

Van Deijk

ALF

Camargo

Timmerman

, et al. Astrocyte lipid metabolism is critical for synapse development and function in vivo. Glia 2017; 65: 670–682.

62.

Liu

Trotter

Zhang

, et al. Neuronal LRP1 knockout in adult mice leads to impaired brain lipid metabolism and progressive, age-dependent synapse loss and neurodegeneration. J Neurosci 2010; 30: 17068–17078.

63.

Fünfschilling

Saher

Xiao

, et al. Survival of adult neurons lacking cholesterol synthesis in vivo. BMC Neurosci 2007; 8: 1.

64.

Bobrowska

Donmez

Weiss

, et al. SIRT2 Ablation has no effect on tubulin acetylation in brain, cholesterol biosynthesis or the progression of Huntington’s disease phenotypes in vivo. PLoS One 2012; 7: e34805.

65.

Lee

Tecedor

Chen

, et al. Reinstating aberrant mTORC1 activity in Huntington’s disease mice improves disease phenotypes. Neuron 2015; 85: 303–315.

66.

Di Pardo

Monyror

Morales

, et al. Mutant huntingtin interacts with the sterol regulatory element-binding proteins and impairs their nuclear import. Hum Mol Genet 2020; 29: 418–431.

67.

Diaz-Castro

Gangwani

, et al. Astrocyte molecular signatures in Huntington’s disease. Sci Transl Med 2019; 11: eaaw8546.

68.

Benraiss

Mariani

Osipovitch

, et al. Cell-intrinsic glial pathology is conserved across human and murine models of Huntington’s disease. Cell Rep 2021; 36: 109308.

69.

Lange

Gillham

Flower

, et al. Polyq length-dependent metabolic alterations and DNA damage drive human astrocyte dysfunction in Huntington’s disease. Prog Neurobiol 2023; 225: 102448.

70.

Shao

Espenshade

. Expanding roles for SREBP in metabolism. Cell Metab 2012; 16: 414–419.

71.

Jeon

Osborne

. SREBPs: metabolic integrators in physiology and metabolism. Trends Endocrinol Metab 2012; 23: 65–72.

72.

Osborne

Espenshade

. Evolutionary conservation and adaptation in the mechanism that regulates SREBP action: what a long, strange tRIP it’s been. Genes Dev 2009; 23: 2578–2591.

73.

Bonvento

Bolaños

. Astrocyte-neuron metabolic cooperation shapes brain activity. Cell Metab 2021; 33: 1546–1564.

74.

Benarroch

. Intrinsic circuits of the striatum. Neurology 2016; 86: 1531–1542.

75.

Pellerin

Magistretti

. Glutamate uptake into astrocytes stimulates aerobic glycolysis: a mechanism coupling neuronal activity to glucose utilization. Proc Natl Acad Sci U S A 1994; 91: 10625–10629.

76.

Ciarmiello

Cannella

Lastoria

, et al. Brain white-matter volume loss and glucose hypometabolism precede the clinical symptoms of Huntington’s disease. J Nucl Med 2006; 47: 215–222.

77.

Tang

Feigin

, et al. Metabolic network as a progression biomarker of premanifest Huntington’s disease. J Clin Invest 2013; 123: 4076–4088.

78.

Delva

Van Laere

Vandenberghe

. Longitudinal imaging of regional brain volumes, SV2A, and glucose metabolism in Huntington’s disease. Mov Disord 2023; 38: 1515–1526.

79.

Tramutola

Bakels

Perrone

, et al. GLUT-1 changes in paediatric Huntington disease brain cortex and fibroblasts: an observational case-control study. EBioMedicine 2023; 97: 104849.

80.

Garcia

Melchinger

Medeiros

, et al. Glutamine sensing licenses cholesterol synthesis. EMBO J 2024; 43: 5837–5856.

81.

Skotte

Andersen

Santos

, et al. Integrative characterization of the R6/2 mouse model of Huntington’s disease reveals dysfunctional astrocyte metabolism. Cell Rep 2018; 23: 2211–2224.

82.

Goicoechea

Conde de la Rosa

Torres

, et al. Mitochondrial cholesterol: metabolism and impact on redox biology and disease. Redox Biol 2023; 61: 102643.

83.

Decker

Funai

. Mitochondrial membrane lipids in the regulation of bioenergetic flux. Cell Metab 2024; 36: 1963–1978.

84.

Goebel

Heipertz

Scholz

, et al. Juvenile Huntington chorea: clinical, ultrastructural, and biochemical studies. Neurology 1978; 28: 23–31.

85.

Machiela

Rudich

Traa

, et al. Targeting mitochondrial network disorganization is protective in C. Elegans models of Huntington’s disease. Aging Dis 2021; 12: 1753–1772.

86.

Cui

Jeong

Borovecki

, et al. Transcriptional repression of PGC-1α by mutant huntingtin leads to mitochondrial dysfunction and neurodegeneration. Cell 2006; 127: 59–69.

87.

Xiang

Valenza

Cui

, et al. Peroxisome-proliferator-activated receptor gamma coactivator 1 α contributes to dysmyelination in experimental models of Huntington’s disease. J Neurosci 2011; 31: 9544–9553.

88.

Seo

Jeon

Il Chong

, et al. Genome-wide localization of SREBP-2 in hepatic chromatin predicts a role in autophagy. Cell Metab 2011; 13: 367–375.

89.

Castellano

Thelen

Moldavski

, et al. Lysosomal cholesterol activates mTORC1 via an SLC38A9-Niemann-Pick C1 signaling complex. Science 2017; 355: 1306–1311.

90.

Peterson

Sengupta

Harris

, et al. MTOR complex 1 regulates lipin 1 localization to control the SREBP pathway. Cell 2011; 146: 408–420.

91.

Eid

Dauner

Courtney

, et al. mTORC1 activates SREBP-2 by suppressing cholesterol trafficking to lysosomes in mammalian cells. Proc Natl Acad Sci U S A 2017; 114: 7999–8004.

92.

Bertrand

Dahmane

. Sonic hedgehog signaling in forebrain development and its interactions with pathways that modify its effects. Trends Cell Biol 2006; 16: 597–605.

93.

Guy

. Inhibition of sonic hedgehog autoprocessing in cultured mammalian cells by sterol deprivation. Proc Natl Acad Sci U S A 2000; 97: 7307–7312.

94.

Cooper

Wassif

Krakowiak

, et al. A defective response to Hedgehog signaling in disorders of cholesterol biosynthesis. Nat Genet 2003; 33: 508–513.

95.

Incardona

Roelink

. The role of cholesterol in Shh signaling and teratogen-induced holoprosencephaly. Cell Mol Life Sci 2000; 57: 1709–1719.

96.

Hill

Garcia

ADR

. Sonic hedgehog signaling in astrocytes. Cell Mol Life Sci 2021; 78: 1393–1403.

97.

Wang

Guo

Acharya

, et al. Primary cilia signaling in astrocytes mediates development and regional-specific functional specification. Nat Neurosci 2024; 27: 1708–1720.

98.

Kumamoto

Wang

, et al. A role for primary cilia in glutamatergic synaptic integration of adult-born neurons. Nat Neurosci 2012; 15: 399–405.

99.

Frasca

Spiombi

Palmieri

, et al. MECP2 Mutations affect ciliogenesis: a novel perspective for Rett syndrome and related disorders. EMBO Mol Med 2020; 12: e10270.

100.

Schmidt

Luecken

Trümbach

, et al. Primary cilia and SHH signaling impairments in human and mouse models of Parkinson’s disease. Nat Commun 2022; 13: 4819.

101.

Keryer

Pineda

Liot

, et al. Ciliogenesis is regulated by a huntingtin-HAP1-PCM1 pathway and is altered in Huntington disease. J Clin Invest 2011; 121: 4372–4338.

102.

Kaliszewski

Knott

Bossy-Wetzel

. Primary cilia and autophagic dysfunction in Huntington’s disease. Cell Death Differ 2015; 22: 1413–1424.