MicroRNAs Associated with IgLON Cell Adhesion Molecule Expression.

The IgLON family of cell adhesion molecules consists of five members (LSAMP, OPCML, neurotrimin, NEGR1, and IgLON5) discovered as supporters of neuronal development, axon growth and guidance, and synapse formation and maintenance. Tumour suppression properties have recently been emerging based on antiproliferative effects through the modulation of oncogenic pathways. Available evidence endorses a role for non-coding RNAs or microRNAs as relevant controllers of IgLON molecule expression that can impact their critical physiological and pathological roles. Current findings support a function for long non-coding RNAs and microRNAs in the modulation of LSAMP expression in cell senescence, cancer biogenesis, addiction, and pulmonary hypertension. For OPCML, data point to a role for several microRNAs in the control of tumorigenesis. MicroRNAs were detected in neurotrimin-mediated functions in cancer biogenesis and in Schwann cell responses to peripheral nerve injury. For NEGR1, studies have mainly investigated microRNA involvement in neuronal responses to ischaemic injury, although data also exist about tumorigenesis and endothelial cell dysfunction. For IgLON5, information is only available about microRNA involved in myocardial infarction. In conclusion, despite much information being still missing and further research needed, the emerging picture favours a model in which non-coding RNAs exert a crucial role in modulating IgLON expression, ultimately affecting their important physiological functions.

Alternative Promoter Use Governs the Expression of IgLON Cell Adhesion Molecules in Histogenetic Fields of the Embryonic Mouse Brain.

The members of the IgLON superfamily of cell adhesion molecules facilitate fundamental cellular communication during brain development, maintain functional brain circuitry, and are associated with several neuropsychiatric disorders such as depression, autism, schizophrenia, and intellectual disabilities. Usage of alternative promoter-specific 1a and 1b mRNA isoforms in Lsamp, Opcml, Ntm, and the single promoter of Negr1 in the mouse and human brain has been previously described. To determine the precise spatiotemporal expression dynamics of Lsamp, Opcml, Ntm isoforms, and Negr1, in the developing brain, we generated isoform-specific RNA probes and carried out in situ hybridization in the developing (embryonic, E10.5, E11.5, 13.5, 17; postnatal, P0) and adult mouse brains. We show that promoter-specific expression of IgLONs is established early during pallial development (at E10.5), where it remains throughout its differentiation through adulthood. In the diencephalon, midbrain, and hindbrain, strong expression patterns are initiated a few days later and begin fading after birth, being only faintly expressed during adulthood. Thus, the expression of specific IgLONs in the developing brain may provide the means for regionally specific functionality as well as for specific regional vulnerabilities. The current study will therefore improve the understanding of how IgLON genes are implicated in the development of neuropsychiatric disorders.

A long non-coding RNA, BC048612 and a microRNA, miR-203 coordinate the gene expression of neuronal growth regulator 1 (NEGR1) adhesion protein.

The regulatory roles for non-coding RNAs, the long non-coding RNAs and microRNAs, are emerging as crucial determinants of central nervous system development and function. Neuronal growth regulator 1 (NEGR1) is a cell adhesion molecule that has been shown to play an important role in neurite outgrowth during neuronal development. Precise expression of the Negr1 gene is crucial for proper brain development and is dysregulated during brain injury. Hence, we attempted to elucidate the non-coding RNAs that control Negr1 gene expression. A long non-coding RNA, BC048612, transcribed from the bidirectional GC-rich Negr1 gene promoter was found to influence Negr1 mRNA expression. In vitro knockdown of the long non-coding RNA resulted in significant down-regulation of Negr1 mRNA expression, NEGR1 protein levels and neurite length whereas over-expression enhanced Negr1 mRNA expression, NEGR1 protein levels and increased neurite length. Meanwhile, another non-coding RNA, microRNA-203, was found to target the 3' untranslated region of the Negr1 mRNA. Inhibition of microRNA-203 led to increased expression of Negr1 mRNA, elevated NEGR1 protein levels and increased neurite length. Conversely, microRNA-203 over-expression decreased the level of Negr1 mRNA, NEGR1 protein and neurite length. Neither microRNA-203 nor the long non-coding RNA, BC048612 could influence each other's expression. Hence, the long non-coding RNA, BC048612, and microRNA-203 were determined to be positive and negative regulators of Negr1 gene expression respectively. These processes have a direct effect on NEGR1 protein levels and neurite length, thus highlighting the importance of the regulatory non-coding RNAs in modulating Negr1 gene expression for precise neuronal development.

Expression studies of six human obesity-related genes in seven tissues from divergent pig breeds.

Obesity has reached epidemic proportions globally and has become the cause of several major health risks worldwide. Presently, more than 100 loci have been related to obesity and metabolic traits in humans by genome-wide association studies. The complex genetic architecture behind obesity has triggered a need for the development of better animal models than rodents. The pig has emerged as a very promising biomedical model to study human obesity traits. In this study, we have characterized the expression patterns of six obesity-related genes, leptin (LEP), leptin receptor (LEPR), melanocortin 4 receptor (MC4R), fat mass and obesity associated (FTO), neuronal growth regulator 1 (NEGR)1 and adiponectin (ADIPOQ), in seven obesity-relevant tissues (liver; muscle; pancreas; hypothalamus; and retroperitoneal, subcutaneous and mesenteric adipose tissues) in two pig breeds (production pigs and Göttingen minipigs) that deviate phenotypically and genetically from each other with respect to obesity traits. We observe significant differential expression for LEP, LEPR and ADIPOQ in muscle and in all three adipose tissues. Interestingly, in pancreas, LEP expression is only detected in the fat minipigs. FTO shows significant differential expression in all tissues analyzed, and NEGR1 shows significant differential expression in muscle, pancreas, hypothalamus and subcutaneous adipose tissue. The MC4R transcript can be detected only in hypothalamus. In general, the expression profiles of the investigated genes are in accordance with those observed in human studies. Our study shows that both the differences between the investigated breeds and the phenotypic state with respect to obesity/leanness play a large role for differential expression of the obesity-related genes.

The IgLON family of cell adhesion molecules expressed in developing neural circuits ensure the proper functioning of the sensory system in mice.

Deletions and malfunctions of the IgLON family of cell adhesion molecules are associated with anatomical, behavioral, and metabolic manifestations of neuropsychiatric disorders. We have previously shown that IgLON genes are expressed in sensory nuclei/pathways and that IgLON proteins modulate sensory processing. Here, we examined the expression of IgLON alternative promoter-specific isoforms during embryonic development and studied the sensory consequences of the anatomical changes when one of the IgLON genes, Negr1, is knocked out. At the embryonal age of E12.5 and E13.5, various IgLONs were distributed differentially and dynamically in the developing sensory areas within the central and peripheral nervous system, as well as in limbs and mammary glands. Sensory tests showed that Negr1 deficiency causes differences in vestibular function and temperature sensitivity in the knockout mice. Sex-specific differences were noted across olfaction, vestibular functioning, temperature regulation, and mechanical sensitivity. Our findings highlight the involvement of IgLON molecules during sensory circuit formation and suggest Negr1's critical role in somatosensory processing.

The Risk Genes for Neuropsychiatric Disorders negr1 and opcml Are Expressed throughout Zebrafish Brain Development.

The immunoglobulin LAMP/OBCAM/NTM (IgLON) family of cell adhesion molecules comprises five members known for their involvement in establishing neural circuit connectivity, fine-tuning, and maintenance. Mutations in IgLON genes result in alterations in these processes and can lead to neuropsychiatric disorders. The two IgLON family members NEGR1 and OPCML share common links with several of them, such as schizophrenia, autism, and major depressive disorder. However, the onset and the underlying molecular mechanisms have remained largely unresolved, hampering progress in developing therapies. NEGR1 and OPCML are evolutionarily conserved in teleosts like the zebrafish (Danio rerio), which is excellently suited for disease modelling and large-scale screening for disease-ameliorating compounds. To explore the potential applicability of zebrafish for extending our knowledge on NEGR1- and OPCML-linked disorders and to develop new therapeutic strategies, we investigated the spatio-temporal expression of the two genes during early stages of development. negr1 and opcml are expressed maternally and subsequently in partially distinct domains of conserved brain regions. Other areas of expression in zebrafish have not been reported in mammals to date. Our results indicate that NEGR1 and OPCML may play roles in neural circuit development and function at stages earlier than previously anticipated. A detailed functional analysis of the two genes based on our findings could contribute to understanding the mechanistic basis of related psychiatric disorders.

Negr1-Derived Peptides Trigger ALK Degradation and Halt Neuroblastoma Progression In Vitro and In Vivo.

Neuroblastoma is among the most common childhood cancers. Neuroblastoma in advanced stages is one of the most intractable pediatric cancers, notwithstanding the recent therapeutic advances. ALK mutations are among the leading cause of hereditary neuroblastoma and account for more than 14% of the somatically acquired alterations. ALK kinase activity is currently one of the main targets for pharmacological strategies. However, evidence from ALK fusion-positive lung cancer studies has shown that resistance to ALK inhibition arises during the therapy, causing a relapse within several years. IgLONs are membrane-bound proteins involved in cell-to-cell adhesion. The expression of the IgLON family results altered in different cancers. We found that the IgLON member Negr1 is downregulated in neuroblastoma. The ectopic overexpression of Negr1 impairs neuroblastoma growth in vitro and in vivo. Negr1 exists as a GPI-anchored membrane-bound protein and as a soluble protein released upon metalloprotease cleavage. We generated and characterized a panel of Negr1-derived peptides. The treatment with Negr1 protein and derived peptides induce ALK downregulation and halt neuroblastoma progression in vitro and in vivo.

The Role of IgLON Cell Adhesion Molecules in Neurodegenerative Diseases.

In the brain, cell adhesion molecules (CAMs) are critical for neurite outgrowth, axonal fasciculation, neuronal survival and migration, and synapse formation and maintenance. Among CAMs, the IgLON family comprises five members: Opioid Binding Protein/Cell Adhesion Molecule Like (OPCML or OBCAM), Limbic System Associated Membrane Protein (LSAMP), neurotrimin (NTM), Neuronal Growth Regulator 1 (NEGR1), and IgLON5. IgLONs exhibit three N-terminal C2 immunoglobulin domains; several glycosylation sites; and a glycosylphosphatidylinositol anchoring to the membrane. Interactions as homo- or heterodimers in cis and in trans, as well as binding to other molecules, appear critical for their functions. Shedding by metalloproteases generates soluble factors interacting with cellular receptors and activating signal transduction. The aim of this review was to analyse the available data implicating a role for IgLONs in neuropsychiatric disorders. Starting from the identification of a pathological role for antibodies against IgLON5 in an autoimmune neurodegenerative disease with a poorly understood mechanism of action, accumulating evidence links IgLONs to neuropsychiatric disorders, albeit with still undefined mechanisms which will require future thorough investigations.

Exercise training remodels inguinal white adipose tissue through adaptations in innervation, vascularization, and the extracellular matrix.

Inguinal white adipose tissue (iWAT) is essential for the beneficial effects of exercise training on metabolic health. The underlying mechanisms for these effects are not fully understood, and here, we test the hypothesis that exercise training results in a more favorable iWAT structural phenotype. Using biochemical, imaging, and multi-omics analyses, we find that 11 days of wheel running in male mice causes profound iWAT remodeling including decreased extracellular matrix (ECM) deposition and increased vascularization and innervation. We identify adipose stem cells as one of the main contributors to training-induced ECM remodeling, show that the PRDM16 transcriptional complex is necessary for iWAT remodeling and beiging, and discover neuronal growth regulator 1 (NEGR1) as a link between PRDM16 and neuritogenesis. Moreover, we find that training causes a shift from hypertrophic to insulin-sensitive adipocyte subpopulations. Exercise training leads to remarkable adaptations to iWAT structure and cell-type composition that can confer beneficial changes in tissue metabolism.

Neuronal growth regulator 1 may modulate interleukin-6 signaling in adipocytes.

Interleukin-6 (IL-6) is a pleiotropic cytokine that plays both anti- and pro-inflammatory roles. Due to the restricted expression of membrane IL-6 receptor (IL-6R), most pro-inflammatory functions of IL-6 are attributed to its association with soluble IL-6R (sIL-6R). Neuronal growth regulator 1 (NEGR1) is a brain-enriched membrane protein that has recently been recognized as a risk factor for many human diseases including obesity, depression, and autism. In the present study, we report that the expression levels of IL-6 and IL-6R, as well as the phosphorylation of signal transducer and activator of transcription (STAT) 3, were significantly elevated in white adipose tissues of Negr1 knockout mice. Elevated levels of circulating IL-6 and sIL-6R have also been observed in Negr1 -/- mice. Furthermore, NEGR1 interacted with IL-6R, which was supported by subcellular fractionation and an in situ proximity ligation assay. Importantly, NEGR1 expression attenuated the phosphorylation of STAT3 by sIL-6R, suggesting that NEGR1 negatively regulates IL-6 trans-signaling. Taken together, we propose that NEGR1 may play a regulatory role in IL-6 signaling by interacting with IL-6R, which may contribute to a molecular link underlying obesity, inflammation, and the depression cycle.

Depression-Associated Negr1 Gene-Deficiency Induces Alterations in the Monoaminergic Neurotransmission Enhancing Time-Dependent Sensitization to Amphetamine in Male Mice.

In GWAS studies, the neural adhesion molecule encoding the neuronal growth regulator 1 (NEGR1) gene has been consistently linked with both depression and obesity. Although the linkage between NEGR1 and depression is the strongest, evidence also suggests the involvement of NEGR1 in a wide spectrum of psychiatric conditions. Here we show the expression of NEGR1 both in tyrosine- and tryptophan hydroxylase-positive cells. Negr1-/- mice show a time-dependent increase in behavioral sensitization to amphetamine associated with increased dopamine release in both the dorsal and ventral striatum. Upregulation of transcripts encoding dopamine and serotonin transporters and higher levels of several monoamines and their metabolites was evident in distinct brain areas of Negr1-/- mice. Chronic (23 days) escitalopram-induced reduction of serotonin and dopamine turnover is enhanced in Negr1-/- mice, and escitalopram rescued reduced weight of hippocampi in Negr1-/- mice. The current study is the first to show alterations in the brain monoaminergic systems in Negr1-deficient mice, suggesting that monoaminergic neural circuits contribute to both depressive and obesity-related phenotypes linked to the human NEGR1 gene.

CircDLGAP4 overexpression relieves oxygen-glucose deprivation-induced neuronal injury by elevating NEGR1 through sponging miR-503-3p.

Circular RNAs (circRNAs) have been reported to play vital regulatory roles in human diseases. However, the functions of circRNAs in ischemic stroke (IS) are limited. In this study, we aimed to explore the functions and mechanisms of circRNA DLG associated protein 4 (circDLGAP4) in IS development. Oxygen-glucose deprivation (OGD)-treated HCN-2 cells were used to mimic IS environment in vitro. Quantitative real-time polymerase chain reaction (qRT-PCR) assay was used to detect the levels of circDLGAP4, microRNA-503-3p (miR-503-3p) and neuronal growth regulator 1 (NEGR1) mRNA. RNase R assay was conducted to analyze the stability of circDLGAP4. Cell Counting Kit-8 (CCK-8) assay and flow cytometry analysis were adopted for cell viability and death, respectively. Western blot assay was performed for protein levels. Enzyme-linked immunosorbent assay (ELISA) kits were used to examine the concentrations of inflammatory cytokines. Dual-luciferase reporter assay, RNA immunoprecipitation (RIP) assay and RNA pull-down assay were employed to analyze the relationships among circDLGAP4, miR-503-3p and NEGR1. CircDLGAP4 level was declined in HCN-2 cells after OGD treatment. CircDLGAP4 overexpression promoted cell viability and suppressed cell death and inflammatory cytokine concentrations in OGD-treated HCN-2 cells. CircDLGAP4 acted as the sponge for miR-503-3p and the impacts of circDLGAP4 overexpression on cell viability, death and inflammation in OGD-treated HCN-2 cells were reversed by miR-503-3p elevation. Furthermore, NEGR1 was the target gene of miR-503-3p. MiR-503-3p inhibition ameliorated OGD-induced HCN-2 cell impairments, but NEGR1 knockdown abolished the effects. CircDLGAP4 alleviated OGD-induced HCN-2 cell damage by regulating miR-503-3p/NEGR1 axis.

Genetic variants in obesity-related genes and the risk of osteoporotic fracture. The Hortega Follow-up Study.

BACKGROUND: Osteoporosis and obesity are major public health problems that are closely correlated, as they share various features, including a genetic predisposition. A genetic correlation between obesity and osteoporosis due to the biological common pathways of bone and fat metabolism, which implies pleiotropic genes regulating has been described. The objective of our study was to analyse whether polymorphisms in obesity-related genes modify the risk of osteoporotic bone fracture. METHODS: We studied 575 subjects from the Hortega Study. The subjects were followed-up for 12-14 years. 202 subjects were overweight, 143 obese and 221 had bone fractures. The distribution of 39 genetic variants in 22 obesity-related genes were studied. RESULTS: The results showed a relationship between polymorphisms in the FTO and NEGR1 genes and the susceptibility to osteoporotic fracture. The variant genotype of the rs2568958 NEGR1 polymorphism and the rs6499649, rs3751812, and rs8044769 genetic variants in FTO were associated with susceptibility to bone fracture. In the best of our knowledge, this is the first time that these variants in NEGR1 and FTO genes have been associated with the susceptibility to osteoporotic bone fracture, supporting the hypothesis that the NEGR1 and FTO genes might be candidates for osteoporosis and bone fracture. CONCLUSIONS: In conclusion, this study associates obesity-related polymorphisms in the NEGR1 and FTO genes with osteoporotic bone fracture, reinforcing the hypothesis that obesity and bone metabolism are closely correlated genetically.

High-Fat Diet Induces Pre-Diabetes and Distinct Sex-Specific Metabolic Alterations in Negr1-Deficient Mice.

In the large GWAS studies, NEGR1 gene has been one of the most significant gene loci for body mass phenotype. The purpose of the current study was to clarify the role of NEGR1 in the maintenance of systemic metabolism, including glucose homeostasis, by using both male and female Negr1-/- mice receiving a standard or high fat diet (HFD). We found that 6 weeks of HFD leads to higher levels of blood glucose in Negr1-/- mice. In the glucose tolerance test, HFD induced phenotype difference only in male mice; Negr1-/- male mice displayed altered glucose tolerance, accompanied with upregulation of circulatory branched-chain amino acids (BCAA). The general metabolomic profile indicates that Negr1-/- mice are biased towards glyconeogenesis, fatty acid synthesis, and higher protein catabolism, all of which are amplified by HFD. Negr1 deficiency appears to induce alterations in the efficiency of energy storage; reduced food intake could be an attempt to compensate for the metabolic challenge present in the Negr1-/- males, particularly during the HFD exposure. Our results suggest that the presence of functional Negr1 allows male mice to consume more HFD and prevents the development of glucose intolerance, liver steatosis, and excessive weight gain.

1p31.1 microdeletion including only NEGR1 gene in two patients.

Neuronal growth regulator 1 (NEGR1), a member of the immunoglobulin superfamily cell adhesion molecule subgroup IgLON, has been involved in neuronal growth and connectivity. Genetic variants, in or near the NEGR1 locus, have been associated with obesity and, more recently, with learning difficulties, intellectual disability, and psychiatric disorders. Here, we described the only second report of NEGR1 gene disruption in 1p31.1 microdeletion in two patients. Patient 1 is a 14-year-old female with neurological and psychiatric features present also in her family. Patient 2 is a 5-month-old infant showing global hypotonia as unique neurological features till now. This patient also carries 7p22.1 duplication, of paternal origin, that could be responsible for some malformations present in the child. We hypothesize a role of NEGR1 in producing the phenotype of our patients and compare them with other cases previously reported in the literature and DECIPHER database to better identify a possible genotype-phenotype correlation.

Circ_HECW2 functions as a miR-30e-5p sponge to regulate LPS-induced endothelial-mesenchymal transition by mediating NEGR1 expression.

Endothelial-mesenchymal transition (EndoMT) plays a critical role in the dysfunction of the blood-brain barrier (BBB). Circular RNAs (circRNAs) function as crucial regulatory factors in EndoMT. Nevertheless, the underlying mechanisms of circRNA HECW2 (circ_HECW2, hsa_circ_0057583) in lipopolysaccharide (LPS)-induced EndoMT remain largely unclear. The levels of circ_HECW2, miR-30e-5p and neuronal growth regulator 1 (NEGR1) were detected by quantitative real-time polymerase chain reaction (qRT-PCR) or western blot. Ribonuclease (RNase) R and Actinomycin D assays were performed to validate the stability of circ_HECW2. Cell colony formation, proliferation and apoptosis were tested by a standard colony formation assay, the 3-(4,5-dimethylthiazol-2yl)-5-(3-carboxymethoxyphenyl)-2-(4-sulfophenyl)-2H-tetrazolium (MTS) assay and flow cytometry, respectively. Targeted relationships among circ_HECW2, miR-30e-5p and NEGR1 were verified by a dual-luciferase reporter assay. Our data indicated that LPS increased circ_HECW2 expression and reduced miR-30e-5p expression in human brain microvascular endothelial cells (HBMECs). Circ_HECW2 silencing promoted cell proliferation and suppressed cell apoptosis and EndoMT in LPS-treated HBMECs. Mechanistically, circ_HECW2 directly interacted with miR-30e-5p by binding to miR-30e-5p. MiR-30e-5p was a functional mediator of circ_HECW2 in regulating LPS-induced cell EndoMT. Furthermore, Circ_HECW2 regulated NEGR1 expression through functioning as a miR-30e-5p sponge. Moreover, miR-30e-5p overexpression repressed the EndoMT of LPS-treated HBMECs by targeting NEGR1. Collectively, our current study demonstrated that circ_HECW2 silencing suppressed LPS-triggered HBMEC EndoMT at least in part through the regulation of the miR-30e-5p/NEGR1 axis, illuminating a promising strategy for EndoMT inhibition.

Polymorphisms in Neuronal Growth Regulator 1 and Otoancorin Alternate the Susceptibility to Lung Cancer in Chinese Nonsmoking Females.

Cell adhesion molecules (CAMs) play crucial roles in the genesis and progress of tumor. We investigated the effects of single nucleotide polymorphisms (SNPs) of CAMs, neuronal growth regulator 1 (NEGR1), and Otoancorin (OTOA) on lung cancer susceptibility in Chinese nonsmoking females. Logistic regression and Cox regression analyses were conducted to investigate the effects of SNPs and environmental factors. For rs3102911, genotype TT carriers decreased the risk of lung cancer with an odds ratio (OR) of 0.635. AA genotypes of rs741718 increased the risk of lung cancer with an OR of 3.527. In stratified analysis, genotype AA carriers of rs741718 had a high susceptibility to lung adenocarcinoma compared with GG and AG genotypes. Analyses of association between SNPs and clinical characteristics revealed that rs3102911 as a protective factor and rs741718 as a risk factor influenced the lung cancer occurrence and progression in nonsmoking females.

Association of the NEGR1 rs2815752 with obesity and related traits in Pakistani females.

Introduction: The variant NEGR1 rs2815752 has recently been linked with obesity in Caucasians. However, a very limited number of studies have examined the association of the NEGR1 rs2815752 with overweight/obesity in non-Caucasians with no such study ever performed in Pakistani population. Therefore, the present study was undertaken to seek the association of the rs2815752 with overweight, obesity, and related traits in Pakistanis.Subjects and methods: The study involved 112 overweight/control pairs (total 224) and 194 obese/control pairs (total 388). Anthropometric parameters were measured by employing standard procedures. Metabolic parameters were determined by biochemical assays. Behavioral information was collected through a questionnaire. The rs2815752 was genotyped via TaqMan allelic discrimination assay. Regression analyses were employed to analyze the data in SPSS software.Results: The study revealed significant gender-specific association of the rs2815752 with obesity (OR 3.03; CI 1.19-7.72, p = 0.020) and some obesity-related anomalous anthropometric traits (weight, BMI, waist circumference, hip circumference, and abdominal and supra-iliac skinfold thicknesses) in females according to dominant model (h = 0.023). However, no association of the rs2815752 with obesity-related behavioral and metabolic parameters was observed.Conclusion: The NEGR1 rs2815752 may be associated with obese phenotype and some of the related anthropometric traits in Pakistani females.

From Bound Cells Comes a Sound Mind: The Role of Neuronal Growth Regulator 1 in Psychiatric Disorders.

Cell-to-cell adhesion is important for maintenance of brain structure and function. Abnormal neuronal cell adhesion and loss of its connectivity are considered a main cause of psychiatric disorders such as major depressive disorder (MDD). Various cell adhesion molecules (CAMs) are involved in neuronal cell adhesions and thereby affect brain functions such as learning and memory, cognitive functions, and psychiatric functions. Compared with other CAMs, neuronal growth regulator 1 (Negr1) has a distinct functioning mechanism in terms of its cross-talk with cytokine receptor signaling. Negr1 is a member of the immunoglobulin LON (IgLON) family of proteins and is involved in neuronal outgrowth, dendritic arborization, and synapse formation. In humans, Negr1 is a risk gene for obesity based on a genome-wide association study. More recently, accumulating evidence supports that it also plays a critical role in psychiatric disorders. In this review, we discuss the recent findings on the role of Negr1 in MDD, focusing on its regulatory mechanism. We also provide evidence of putative involvement of Negr1 in other psychiatric disorders based on the novel behavioral phenotypes of Negr1 knockout mice.

Deployment of Label-Free Quantitative Olfactory Proteomics to Detect Cerebrospinal Fluid Biomarker Candidates in Synucleinopathies.

Nowadays, diagnosis of neurodegenerative disorders is mainly based on neuroimaging and clinical symptoms, although postmortem neuropathological confirmation remains the gold standard diagnostic technique. Therefore, cerebrospinal fluid (CSF) proteome is considered a valuable molecular repository for diagnosing and targeting the neurodegenerative process. It is well known that olfactory dysfunction is among the earliest features of synucleinopathies such as Parkinson's disease (PD). Consequently, we consider that the application of tissue proteomics in primary olfactory structures is an ideal approach to explore early pathophysiological changes, detecting olfactory proteins that might be tested in CSF as potential biomarkers. Data mining of mass spectrometry-generated datasets has revealed that 30% of the olfactory bulb (OB) proteome is also localized in CSF. In this chapter, we describe a method that utilizes label-free quantitative proteomics and computational analysis to characterize human OB proteomes and potential cerebrospinal fluid (CSF) biomarkers associated with neurodegenerative syndromes. For that, we applied peptide fractionation methods, followed by tandem mass spectrometry (nanoLC-MS/MS), in silico analysis, and semi-quantitative orthogonal techniques in OB derived from PD subjects. After obtaining the differential OB proteome across Lewy-type alpha-synucleinopathy (LTS) stages and further validating the method, this workflow was applied to probe changes in NEGR1 (neuronal growth regulator 1) and GNPDA2 (glucosamine-6-phosphate deaminase 2) protein levels in CSF derived from parkinsonian subjects with respect to controls, observing an inverse correlation between both proteins and α-synuclein, the principal component analysis of Lewy pathology.