A hydrophobic groove in secretagogin allows for alternate interactions with SNAP-25 and syntaxin-4 in endocrine tissues.

Vesicular release of neurotransmitters and hormones relies on the dynamic assembly of the exocytosis/trans-SNARE complex through sequential interactions of synaptobrevins, syntaxins, and SNAP-25. Despite SNARE-mediated release being fundamental for intercellular communication in all excitable tissues, the role of auxiliary proteins modulating the import of reserve vesicles to the active zone, and thus, scaling repetitive exocytosis remains less explored. Secretagogin is a Ca2+-sensor protein with SNAP-25 being its only known interacting partner. SNAP-25 anchors readily releasable vesicles within the active zone, thus being instrumental for 1st phase release. However, genetic deletion of secretagogin impedes 2nd phase release instead, calling for the existence of alternative protein-protein interactions. Here, we screened the secretagogin interactome in the brain and pancreas, and found syntaxin-4 grossly overrepresented. Ca2+-loaded secretagogin interacted with syntaxin-4 at nanomolar affinity and 1:1 stoichiometry. Crystal structures of the protein complexes revealed a hydrophobic groove in secretagogin for the binding of syntaxin-4. This groove was also used to bind SNAP-25. In mixtures of equimolar recombinant proteins, SNAP-25 was sequestered by secretagogin in competition with syntaxin-4. Kd differences suggested that secretagogin could shape unidirectional vesicle movement by sequential interactions, a hypothesis supported by in vitro biological data. This mechanism could facilitate the movement of transport vesicles toward release sites, particularly in the endocrine pancreas where secretagogin, SNAP-25, and syntaxin-4 coexist in both α- and ß-cells. Thus, secretagogin could modulate the pace and fidelity of vesicular hormone release by differential protein interactions.

SCGN and STAT3 expressions are associated with the prognosis of ccRCC.

Clear cell renal cell carcinoma (ccRCC) is highly heterogeneous and accounts for about 70% of RCC. Its prognosis is worse than that of most histological types of RCC. In order to find potential biomarkers that may influence the prognosis and survival in ccRCC patients, we explored the expressions of STAT3, PDL1 and SCGN (secretagogin) in ccRCC based on the data of TCGA (n = 529), EMATAB-1980 (n = 99) and our own cohort (n = 99). Our study demonstrated that ccRCC patients with low STAT3 expression and high SCGN expression might have a better prognosis. No significant difference in the positive rate of SCGN expression was found when comparing the primary lesion with the matched metastatic liver lesions. The percentage of high SCGN expression in the primary lesion of metastatic ccRCC patients was significantly lower than that of patients with only the renal lesion. In view of the conclusion that STAT3 high expression cases are resistant to sunitinib, STAT3 immunohistochemistry results are essential for designing non-operative treatments. SCGN has the potential to become an indicator for subtype classification of ccRCC.

SCGN deficiency is a risk factor for autism spectrum disorder.

Autism spectrum disorder (ASD) affects 1-2% of all children and poses a great social and economic challenge for the globe. As a highly heterogeneous neurodevelopmental disorder, the development of its treatment is extremely challenging. Multiple pathways have been linked to the pathogenesis of ASD, including signaling involved in synaptic function, oxytocinergic activities, immune homeostasis, chromatin modifications, and mitochondrial functions. Here, we identify secretagogin (SCGN), a regulator of synaptic transmission, as a new risk gene for ASD. Two heterozygous loss-of-function mutations in SCGN are presented in ASD probands. Deletion of Scgn in zebrafish or mice leads to autism-like behaviors and impairs brain development. Mechanistically, Scgn deficiency disrupts the oxytocin signaling and abnormally activates inflammation in both animal models. Both ASD probands carrying Scgn mutations also show reduced oxytocin levels. Importantly, we demonstrate that the administration of oxytocin and anti-inflammatory drugs can attenuate ASD-associated defects caused by SCGN deficiency. Altogether, we identify a convergence between a potential autism genetic risk factor SCGN, and the pathological deregulation in oxytocinergic signaling and immune responses, providing potential treatment for ASD patients suffering from SCGN deficiency. Our study also indicates that it is critical to identify and stratify ASD patient populations based on their disease mechanisms, which could greatly enhance therapeutic success.

Immunohistochemical characterization of bipolar cells in four distantly related avian species.

Visual (and probably also magnetic) signal processing starts at the first synapse, at which photoreceptors contact different types of bipolar cells, thereby feeding information into different processing channels. In the chicken retina, 15 and 22 different bipolar cell types have been identified based on serial electron microscopy and single-cell transcriptomics, respectively. However, immunohistochemical markers for avian bipolar cells were only anecdotally described so far. Here, we systematically tested 12 antibodies for their ability to label individual bipolar cells in the bird retina and compared the eight most suitable antibodies across distantly related species, namely domestic chicken, domestic pigeon, common buzzard, and European robin, and across retinal regions. While two markers (GNB3 and EGFR) labeled specifically ON bipolar cells, most markers labeled in addition to bipolar cells also other cell types in the avian retina. Staining pattern of four markers (CD15, PKCα, PKCß, secretagogin) was species-specific. Two markers (calbindin and secretagogin) showed a different expression pattern in central and peripheral retina. For the chicken and European robin, we found slightly more ON bipolar cell somata in the inner nuclear layer than OFF bipolar cell somata. In contrast, OFF bipolar cells made more ribbon synapses than ON bipolar cells in the inner plexiform layer of these species. Finally, we also analyzed the photoreceptor connectivity of selected bipolar cell types in the European robin retina. In summary, we provide a catalog of bipolar cell markers for different bird species, which will greatly facilitate analyzing the retinal circuitry of birds on a larger scale.

Secretagogin is a Ca2+-dependent stress-responsive chaperone that may also play a role in aggregation-based proteinopathies.

Secretagogin (SCGN) is a three-domain hexa-EF-hand Ca2+-binding protein that plays a regulatory role in the release of several hormones. SCGN is expressed largely in pancreatic ß-cells, certain parts of the brain, and also in neuroendocrine tissues. The expression of SCGN is altered in several diseases, such as diabetes, cancers, and neurodegenerative disorders; however, the precise associations that closely link SCGN expression to such pathophysiologies are not known. In this work, we report that SCGN is an early responder to cellular stress, and SCGN expression is temporally upregulated by oxidative stress and heat shock. We show the overexpression of SCGN efficiently prevents cells from heat shock and oxidative damage. We further demonstrate that in the presence of Ca2+, SCGN efficiently prevents the aggregation of a broad range of model proteins in vitro. Small-angle X-ray scattering (BioSAXS) studies further reveal that Ca2+ induces the conversion of a closed compact apo-SCGN conformation into an open extended holo-SCGN conformation via multistate intermediates, consistent with the augmentation of chaperone activity of SCGN. Furthermore, isothermal titration calorimetry establishes that Ca2+ enables SCGN to bind α-synuclein and insulin, two target proteins of SCGN. Altogether, our data not only demonstrate that SCGN is a Ca2+-dependent generic molecular chaperone involved in protein homeostasis with broad substrate specificity but also elucidate the origin of its altered expression in several cancers. We describe a plausible mechanism of how perturbations in Ca2+ homeostasis and/or deregulated SCGN expression would hasten the process of protein misfolding, which is a feature of many aggregation-based proteinopathies.

Calcium-binding proteins typify the dopaminergic neuronal subtypes in the ventral tegmental area of zebra finch, Taeniopygia guttata.

Calcium-binding proteins (CBPs) regulate neuronal function in midbrain dopamine (DA)-ergic neurons in mammals by buffering and sensing the intracellular Ca2+ , and vesicular release. In birds, the equivalent set of neurons are important in song learning, directed singing, courtship, and energy balance, yet the status of CBPs in these neurons is unknown. Herein, for the first time, we probe the nature of CBPs, namely, Calbindin-, Calretinin-, Parvalbumin-, and Secretagogin-expressing DA neurons in the ventral tegmental area (VTA) and substantia nigra (SN) in the midbrain of zebra finch, Taeniopygia guttata. qRT-PCR analysis of ventral midbrain tissue fragment revealed higher Calbindin- and Calretinin-mRNA levels compared to Parvalbumin and Secretagogin. Application of immunofluorescence showed CBP-immunoreactive (-i) neurons in VTA (anterior [VTAa], mid [VTAm], caudal [VTAc]), SN (compacta [SNc], and reticulata [SNr]). Compared to VTAa, higher Calbindin- and Parvalbumin-immunoreactivity (-ir), and lower Calretinin-ir were observed in VTAm and VTAc. Secretagogin-ir was highly localized to VTAa. In SN, Calbindin- and Calretinin-ir were higher in SNc, SNr was Parvalbumin enriched, and Secretagogin-ir was not detected. Weak, moderate, and intense tyrosine hydroxylase (TH)-i VTA neurons were demarcated as subtypes 1, 2, and 3, respectively. While subtype 1 TH-i neurons were neither Calbindin- nor Calretinin-i, ∼80 and ∼65% subtype 2 and ∼30 and ∼45% subtype 3 TH-i neurons co-expressed Calbindin and Calretinin, respectively. All TH-i neuronal subtypes co-expressed Parvalbumin with reciprocal relationship with TH-ir. We suggest that the CBPs may determine VTA DA neuronal heterogeneity and differentially regulate their activity in T. guttata.

Secretagogin in the brain and pituitary of the catfish, Clarias batrachus: Molecular characterization and regulation by insulin.

Secretagogin (scgn), is a novel hexa EF-hand, phylogenetically conserved calcium-binding protein. It serves as Ca2+ sensor and participates in Ca2+ -signaling and neuroendocrine regulation in mammals. However, its relevance in the brain of non-mammalian vertebrates has largely remained unexplored. To address this issue, we studied the cDNA encoding scgn, scgn mRNA expression, and distribution of scgn-equipped elements in the brain and pituitary of a teleost, Clarias batrachus (cb). The cbscgn cDNA consists of three transcripts (T) variants: T1 (2185 bp), T2 (2151 bp) and T3 (2060 bp). While 816 bp ORF in T1 and T2 encodes highly conserved six EF-hand 272 aa protein fully capable of Ca2+ -binding, 726-bp ORF in T3 encodes 242 aa protein. The T1 showed >90% and >70% identity with scgn of catfishes, and other teleosts and mammals, respectively. The T1-mRNA was widely expressed in the brain and pituitary, while the expression of T3 was restricted to the telencephalon. Application of the anti-scgn antiserum revealed a ∼32 kDa scgn-immunoreactive (scgn-i) band (known molecular weight of scgn) in the forebrain tissue, and immunohistochemically labeled neurons in the olfactory epithelium and bulb, telencephalon, preoptic area, hypothalamus, thalamus, and hindbrain. In the pituitary, scgn-i cells were seen in the pars distalis and intermedia. Insulin is reported to regulate scgn mRNA in the mammalian hippocampus, and feeding-related neuropeptides in the telencephalon of teleost. Intracranial injection of insulin significantly increased T1-mRNA expression and scgn-immunoreactivity in the telencephalon. We suggest that scgn may be an important player in the regulation of olfactory, neuroendocrine system, and energy balance functions in C. batrachus.

Extracellular secretagogin is internalized into the cells through endocytosis.

Secretagogin (SCGN) is a calcium-sensor protein with a regulatory role in glucose metabolism and the secretion of several peptide hormones. Many, but not all, functions of SCGN can be explained by its intracellular manifestation. Despite early data on SCGN secretion, the secretory mechanism, biological fate, physiological implications and trans-cellular signalling of extracellular SCGN remain unknown. We here report that extracellular SCGN is readily internalized into the C2C12 cells in an energy-dependent manner. Using endocytosis inhibitors, we demonstrate that SCGN internalizes via clathrin-mediated endocytosis, following which, SCGN localizes largely in the cytosol. Exogenous SCGN treatment induces a global proteomic reprogramming in C2C12 cells. Gene ontology search suggests that SCGN-induced proteomic reprogramming favours protein synthesis and cellular growth. We thus validated the cell proliferative action of SCGN using C2C12, HepG2 and NIH-3T3 cell lines. Based on the data, we propose that circulatory SCGN is internalized into the target cells and modulates the expression of cell growth-related proteins. The work suggests that extracellular SCGN is a functional protein, which communicates with specific cell types and directly modulates cell proliferation.

ox-LDL induces autophagy-mediated apoptosis by suppressing secretagogin-regulated autophagic flux in pancreatic ß-cells.

Lipotoxicity has been shown to induce the loss of functional ß-cell mass and lead to type 2 diabetes, but the mechanism remains unknown. In this study, we aim to explore the role of secretagogin (SCGN) in lipotoxicity-induced ß-cell injury. Our results indicate that ox-LDL treatment leads to autophagic cell death, as evidenced by decreased cell viability, aggravated cell apoptosis, and the accumulation of the p62 protein in MIN6 cells. LysoTracker Red staining, TEM and mRFP-GFP-LC3 assays demonstrate that autophagic flux is blocked in ox-LDL-treated MIN6 cells. Intriguingly, SCGN is significantly decreased in MIN6 cells under lipotoxic conditions. Additionally, siRNA-guided SCGN knockdown blocks autophagic flux triggered by rapamycin, while SCGN restoration alleviates autophagic flux retardation and mitigates cell apoptosis. The physical interaction between SCGN and SNAP29 is validated by bioinformatics analysis, coimmunoprecipitation assay and SCGN knockdown test. Downregulation of SCGN expression reduces the interaction of these two proteins. Taken together, our results indicate that ox-LDL treatment induces apoptotic ß-cell death by blocking autophagic flux dependent on SCGN downregulation. SCGN administration prevents lipotoxic ß-cell injury and may be a potential therapeutic strategy to promote ß-cell expansion in type 2 diabetes.

Secretagogin Mediates the Regulatory Effect of Electroacupuncture on Hypothalamic-Pituitary-Adrenal Axis Dysfunction in Surgical Trauma.

Electroacupuncture (EA) improves hypothalamic-pituitary-adrenal (HPA) axis disorder by reducing corticotropin-releasing hormone (CRH) synthesis and release in the paraventricular nucleus (PVN). However, the potential mechanism underlying CRH regulation remains unclear. Secretagogin (SCGN) is closely related to stress and is involved in regulating the release of CRH. We hypothesized that SCGN in the PVN might trigger the HPA system and be involved in EA-mediated modulation of HPA dysfunction caused by surgical trauma. Serum CRH and adrenocorticotropic hormone (ACTH) and plasma corticosterone (CORT) levels at 6 h and 24 h after hepatectomy were determined by radioimmunoassay. CRH and SCGN protein levels in the PVN were detected by western blot and immunofluorescence, and CRH and SCGN mRNA levels in the PVN were determined by means of real-time polymerase chain reaction (RT-PCR) and in situ hybridization (ISH). Our studies showed that serum CRH, ACTH, and CORT levels and PVN CRH expression were significantly increased at 6 h and 24 h after hepatectomy in the hepatectomy group compared with the control group, and those in the EA+hepatectomy group were decreased compared with those in the hepatectomy group. The protein and mRNA levels of SCGN in the PVN were also increased after hepatectomy, and their expression in the EA+hepatectomy group was decreased compared with that in the hepatectomy group. When SCGN expression in the PVN was functionally knocked down by a constructed CsCI virus, we found that SCGN knockdown decreased the serum CRH, ACTH, and CORT levels in the SCGN shRNA+hepatectomy group compared with the hepatectomy group, and it also attenuated CRH expression in the PVN. In summary, our findings illustrated that EA normalized HPA axis dysfunction after surgical trauma by decreasing the transcription and synthesis of SCGN.

Secretagogin marks amygdaloid PKCδ interneurons and modulates NMDA receptor availability.

The perception of and response to danger is critical for an individual's survival and is encoded by subcortical neurocircuits. The amygdaloid complex is the primary neuronal site that initiates bodily reactions upon external threat with local-circuit interneurons scaling output to effector pathways. Here, we categorize central amygdala neurons that express secretagogin (Scgn), a Ca2+-sensor protein, as a subset of protein kinase Cδ (PKCδ)+ interneurons, likely "off cells." Chemogenetic inactivation of Scgn+/PKCδ+ cells augmented conditioned response to perceived danger in vivo. While Ca2+-sensor proteins are typically implicated in shaping neurotransmitter release presynaptically, Scgn instead localized to postsynaptic compartments. Characterizing its role in the postsynapse, we found that Scgn regulates the cell-surface availability of NMDA receptor 2B subunits (GluN2B) with its genetic deletion leading to reduced cell membrane delivery of GluN2B, at least in vitro. Conclusively, we describe a select cell population, which gates danger avoidance behavior with secretagogin being both a selective marker and regulatory protein in their excitatory postsynaptic machinery.

A novel secretagogin/ATF4 pathway is involved in oxidized LDL-induced endoplasmic reticulum stress and islet ß-cell apoptosis.

Excessive accumulation of cholesterol in ß cells initiates endoplasmic reticulum (ER) stress and associated apoptosis. We have reported that excessive uptake of cholesterol by MIN6 cells decreases the expression of secretagogin (SCGN) and then attenuates insulin secretion. Here, we aimed to determine whether cholesterol-induced SCGN decrease is involved in the modulation of ER stress and apoptosis in pancreatic ß cells. In this study, MIN6 cells were treated with oxidized low-density lipoprotein (ox-LDL) for 24 h, and then intracellular lipid droplets and cell apoptosis were quantified, and SCGN and ER stress markers were identified by western blot analysis. Furthermore, small interfer RNA (siRNA)-mediated SCGN knockdown and recombinant plasmid-mediated SCGN restoration experiments were performed to confirm the role of SCGN in ER stress and associated cell apoptosis. Finally, the interaction of SCGN with ATF4 was computationally predicted and then validated by a co-immunoprecipitation assay. We found that ox-LDL treatment increased the levels of ER stress markers, such as phosphorylated protein kinase-like endoplasmic reticulum kinase, phosphorylated eukaryotic initiation factor 2 alpha, activating transcription factor 4 (ATF4), and transcription factor CCAAT-enhancer-binding protein homologous protein, and promoted MIN6 cell apoptosis; in addition, the expression of SCGN was downregulated. siRNA-mediated SCGN knockdown exacerbated ß-cell ER stress by increasing ATF4 expression. Pretreatment of MIN6 cells with the recombinant SCGN partly antagonized ox-LDL-induced ER stress and apoptosis. Furthermore, a co-immunoprecipitation assay revealed an interaction between SCGN and ATF4 in MIN6 cells. Taken together, these results demonstrated that pancreatic ß-cell apoptosis induced by ox-LDL treatment can be attributed, in part, to an SCGN/ATF4-dependent ER stress response.

Secretagogin expression in the mouse olfactory bulb under sensory impairments.

The interneurons of the olfactory bulb (OB) are characterized by the expression of different calcium-binding proteins, whose specific functions are not fully understood. This is the case of one of the most recently discovered, the secretagogin (SCGN), which is expressed in interneurons of the glomerular and the granule cell layers, but whose function in the olfactory pathway is still unknown. To address this question, we examined the distribution, generation and activity of SCGN-positive interneurons in the OB of two complementary models of olfactory impairments: Purkinje Cell Degeneration (PCD) and olfactory-deprived mice. Our results showed a significant increase in the density of SCGN-positive cells in the inframitral layers of olfactory-deprived mice as compared to control animals. Moreover, BrdU analyses revealed that these additional SCGN-positive cells are not newly formed. Finally, the neuronal activity, estimated by c-Fos expression, increased in preexisting SCGN-positive interneurons of both deprived and PCD mice -being higher in the later- in comparison with control animals. Altogether, our results suggest that the OB possesses different compensatory mechanisms depending on the type of alteration. Particularly, the SCGN expression is dependent of olfactory stimuli and its function may be related to a compensation against a reduction in sensory inputs.

Structural and mechanistic insights into secretagogin-mediated exocytosis.

Secretagogin (SCGN) is a hexa-EF-hand protein that is highly expressed in the pancreas, brain, and gastrointestinal tract. SCGN is known to modulate regulated exocytosis in multiple cell lines and tissues; however, its exact functions and underlying mechanisms remain unclear. Here, we report that SCGN interacts with the plasma membrane SNARE SNAP-25, but not the assembled SNARE complex, in a Ca2+-dependent manner. The crystal structure of SCGN in complex with a SNAP-25 fragment reveals that SNAP-25 adopts a helical structure and binds to EF-hands 5 and 6 of SCGN. SCGN strongly inhibits SNARE-mediated vesicle fusion in vitro by binding to SNAP-25. SCGN promotes the plasma membrane localization of SNAP-25, but not Syntaxin-1a, in SCGN-expressing cells. Finally, SCGN controls neuronal growth and brain development in zebrafish, likely via interacting with SNAP-25 or its close homolog, SNAP-23. Our results thus provide insights into the regulation of SNAREs and suggest that aberrant synapse functions underlie multiple neurological disorders caused by SCGN deficiency.

The core clock gene, Bmal1, and its downstream target, the SNARE regulatory protein secretagogin, are necessary for circadian secretion of glucagon-like peptide-1.

OBJECTIVES: The incretin hormone glucagon-like peptide-1 (GLP-1) is secreted from intestinal L-cells upon nutrient intake. While recent evidence has shown that GLP-1 is released in a circadian manner in rats, whether this occurs in mice and if this pattern is regulated by the circadian clock remain to be elucidated. Furthermore, although circadian GLP-1 secretion parallels expression of the core clock gene Bmal1, the link between the two remains largely unknown. Secretagogin (Scgn) is an exocytotic SNARE regulatory protein that demonstrates circadian expression and is essential for insulin secretion from ß-cells. The objective of the current study was to establish the necessity of the core clock gene Bmal1 and the SNARE protein SCGN as essential regulators of circadian GLP-1 secretion. METHODS: Oral glucose tolerance tests were conducted at different times of the day on 4-hour fasted C57BL/6J, Bmal1 wild-type, and Bmal1 knockout mice. Mass spectrometry, RNA-seq, qRT-PCR and/or microarray analyses, and immunostaining were conducted on murine (m) and human (h) primary L-cells and mGLUTag and hNCI-H716 L-cell lines. At peak and trough GLP-1 secretory time points, the mGLUTag cells were co-stained for SCGN and a membrane-marker, ChIP was used to analyze BMAL1 binding sites in the Scgn promoter, protein interaction with SCGN was tested by co-immunoprecipitation, and siRNA was used to knockdown Scgn for GLP-1 secretion assay. RESULTS: C57BL/6J mice displayed a circadian rhythm in GLP-1 secretion that peaked at the onset of their feeding period. Rhythmic GLP-1 release was impaired in Bmal1 knockout (KO) mice as compared to wild-type controls at the peak (p < 0.05) but not at the trough secretory time point. Microarray identified SNARE and transport vesicle pathways as highly upregulated in mGLUTag L-cells at the peak time point of GLP-1 secretion (p < 0.001). Mass spectrometry revealed that SCGN was also increased at this time (p < 0.001), while RNA-seq, qRT-PCR, and immunostaining demonstrated Scgn expression in all human and murine primary L-cells and cell lines. The mGLUTag and hNCI-H716 L-cells exhibited circadian rhythms in Scgn expression (p < 0.001). The ChIP analysis demonstrated increased binding of BMAL1 only at the peak of Scgn expression (p < 0.01). Immunocytochemistry showed the translocation of SCGN to the cell membrane after stimulation at the peak time point only (p < 0.05), while CoIP showed that SCGN was pulled down with SNAP25 and ß-actin, but only the latter interaction was time-dependent (p < 0.05). Finally, Scgn siRNA-treated cells demonstrated significantly blunted GLP-1 secretion (p < 0.01) in response to stimulation at the peak time point only. CONCLUSIONS: These data demonstrate, for the first time, that mice display a circadian pattern in GLP-1 secretion, which is impaired in Bmal1 knockout mice, and that Bmal1 regulation of Scgn expression plays an essential role in the circadian release of the incretin hormone GLP-1.

Brain-wide genetic mapping identifies the indusium griseum as a prenatal target of pharmacologically unrelated psychostimulants.

Psychostimulant use is an ever-increasing socioeconomic burden, including a dramatic rise during pregnancy. Nevertheless, brain-wide effects of psychostimulant exposure are incompletely understood. Here, we performed Fos-CreERT2-based activity mapping, correlated for pregnant mouse dams and their fetuses with amphetamine, nicotine, and caffeine applied acutely during midgestation. While light-sheet microscopy-assisted intact tissue imaging revealed drug- and age-specific neuronal activation, the indusium griseum (IG) appeared indiscriminately affected. By using GAD67gfp/+ mice we subdivided the IG into a dorsolateral domain populated by γ-aminobutyric acidergic interneurons and a ventromedial segment containing glutamatergic neurons, many showing drug-induced activation and sequentially expressing Pou3f3/Brn1 and secretagogin (Scgn) during differentiation. We then combined Patch-seq and circuit mapping to show that the ventromedial IG is a quasi-continuum of glutamatergic neurons (IG-Vglut1+) reminiscent of dentate granule cells in both rodents and humans, whose dendrites emanate perpendicularly toward while their axons course parallel with the superior longitudinal fissure. IG-Vglut1+ neurons receive VGLUT1+ and VGLUT2+ excitatory afferents that topologically segregate along their somatodendritic axis. In turn, their efferents terminate in the olfactory bulb, thus being integral to a multisynaptic circuit that could feed information antiparallel to the olfactory-cortical pathway. In IG-Vglut1+ neurons, prenatal psychostimulant exposure delayed the onset of Scgn expression. Genetic ablation of Scgn was then found to sensitize adult mice toward methamphetamine-induced epilepsy. Overall, our study identifies brain-wide targets of the most common psychostimulants, among which Scgn+/Vglut1+ neurons of the IG link limbic and olfactory circuits.

Secretagogin Binding Prevents α-Synuclein Fibrillation.

Secretagogin (SCGN) is a secreted calcium sensor that has emerged as a potential multifunctional protein of neuroendocrine cells. A significantly reduced level of expression of SCGN has been reported in the hippocampus of a mouse model of Alzheimer's disease (AD) and in Parkinson's patients, although the biochemical implications and mechanistic underpinnings of the altered SCGN expression in neurodegenerative diseases remain unknown. We have pursued the interaction of SCGN with α-synuclein that we discovered in impartial pull-down analyses to decode the SCGN interactome. SCGN physically binds α-synuclein and rescues it from detrimental fibrillation. Correspondingly, it is observed that a significant reduction in the cytotoxicity of α-synuclein fibrils is caused by SCGN. We map these antifibrillar attributes to the central region and C-terminal domain of SCGN, while the N-terminal domain is not essential for this activity. On the basis of these results, a broader neuroprotective function of SCGN by proficient chaperone action is proposed. An intriguing correlation of this interaction with a reduced level of expression of SCGN in neurodegenerative diseases shall inspire further studies of the physiological role of SCGN in precluding pathological protein aggregation.

A microRNA­24­to­secretagogin regulatory pathway mediates cholesterol­induced inhibition of insulin secretion.

Hypercholesterolemia is a key factor leading to ß­cell dysfunction, but its underlying mechanisms remain unclear. Secretagogin (Scgn), a Ca2+ sensor protein that is expressed at high levels in the islets, has been shown to play a key role in regulating insulin secretion through effects on the soluble N­ethylmaleimide­sensitive factor attachment receptor protein complexes. However, further studies are required to determine whether Scgn plays a role in hypercholesterolemia­associated ß­cell dysfunction. The present study investigated the involvement of a microRNA­24 (miR­24)­to­Scgn regulatory pathway in cholesterol­induced ß­cell dysfunction. In the present study, MIN6 cells were treated with increasing concentrations of cholesterol and then, the cellular functions and changes in the miR­24­to­Scgn signal pathway were observed. Excessive uptake of cholesterol in MIN6 cells increased the expression of miR­24, resulting in a reduction in Sp1 expression by directly targeting its 3' untranslated region. As a transcriptional activator of Scgn, downregulation of Sp1 decreased Scgn levels and subsequently decreased the phosphorylation of focal adhesion kinase and paxillin, which is regulated by Scgn. Therefore, the focal adhesions in insulin granules were impaired and insulin exocytosis was reduced. The present study concluded that a miR­24­to­Scgn pathway participates in the mechanism regulating cholesterol accumulation­induced ß­cell dysfunction.

Secretagogin expression in the vertebrate brainstem with focus on the noradrenergic system and implications for Alzheimer's disease.

Calcium-binding proteins are widely used to distinguish neuronal subsets in the brain. This study focuses on secretagogin, an EF-hand calcium sensor, to identify distinct neuronal populations in the brainstem of several vertebrate species. By using neural tube whole mounts of mouse embryos, we show that secretagogin is already expressed during the early ontogeny of brainstem noradrenaline cells. In adults, secretagogin-expressing neurons typically populate relay centres of special senses and vegetative regulatory centres of the medulla oblongata, pons and midbrain. Notably, secretagogin expression overlapped with the brainstem column of noradrenergic cell bodies, including the locus coeruleus (A6) and the A1, A5 and A7 fields. Secretagogin expression in avian, mouse, rat and human samples showed quasi-equivalent patterns, suggesting conservation throughout vertebrate phylogeny. We found reduced secretagogin expression in locus coeruleus from subjects with Alzheimer's disease, and this reduction paralleled the loss of tyrosine hydroxylase, the enzyme rate limiting noradrenaline synthesis. Residual secretagogin immunoreactivity was confined to small submembrane domains associated with initial aberrant tau phosphorylation. In conclusion, we provide evidence that secretagogin is a useful marker to distinguish neuronal subsets in the brainstem, conserved throughout several species, and its altered expression may reflect cellular dysfunction of locus coeruleus neurons in Alzheimer's disease.