New Dominant-Negative IL6ST Variants Expand the Immunological and Clinical Spectrum of GP130-Dependent Hyper-IgE Syndrome.

Patients with autosomal dominant (AD) hyper-IgE syndrome (HIES) suffer from a constellation of manifestations including recurrent bacterial and fungal infections, severe atopy, and skeletal abnormalities. This condition is typically caused by monoallelic dominant-negative (DN) STAT3 variants. In 2020, we described 12 patients from eight kindreds with DN IL6ST variants resulting in a new form of AD HIES. These variants encoded truncated GP130 receptors, with intact extracellular and transmembrane domains, but lacking the intracellular recycling motif and the four STAT3-binding residues, resulting in an inability to recycle and activate STAT3. We report here two new DN variants of IL6ST in three unrelated families with HIES-AD. The biochemical and clinical impacts of these variants are different from those of the previously reported variants. The p.(Ser731Valfs*8) variant, identified in seven patients from two families, lacks the recycling motif and all the STAT3-binding residues, but its levels on the cell surface are only slightly increased and it underlies mild biological phenotypes with variable clinical expressivity. The p.(Arg768*) variant, identified in a single patient, lacks the recycling motif and the three most distal STAT3-binding residues. This variant accumulates at the cell surface and underlies severe biological and clinical phenotypes. The p.(Ser731Valfs*8) variant shows that a DN GP130 expressed at near normal levels on the cell surface can underlie heterogeneous clinical presentations, ranging from mild to severe. The p.(Arg768*) variant demonstrates that a truncated GP130 protein retaining one STAT3-binding residue can underlie severe HIES.

Immune deficiency, autoimmune disease and intellectual disability: A pleiotropic disorder caused by biallelic variants in the TPP2 gene.

Four individuals from two families presented with a multisystemic condition of suspected genetic origin that was diagnosed only after genome analysis. The main phenotypic features were immune system dysregulation (severe immunodeficiency with autoimmunity) and intellectual disability. The four individuals were found to be homozygous for a 4.4 Kb deletion removing exons 20-23 (NM_003291.4) of the TPP2 gene, predicting a frameshift with premature termination of the protein. The deletion was located on a shared chromosome 13 haplotype indicating a Swiss founder mutation. Tripeptidyl peptidase 2 (TPP2) is a protease involved in HLA/antigen complex processing and amino acid homeostasis. Biallelic variants in TPP2 have been described in 10 individuals with variable features including immune deficiency, autoimmune cytopenias, and intellectual disability or chronic sterile brain inflammation mimicking multiple sclerosis. Our observations further delineate this severe condition not yet included in the OMIM catalog. Timely recognition of TPP2 deficiency is crucial since (1) immune surveillance is needed and hematopoietic stem cell transplantation may be necessary, and (2) for provision of genetic counselling. Additionally, enzyme replacement therapy, as already established for TPP1 deficiency, might be an option in the future.

Peripubertal stress-induced heightened aggression: modulation of the glucocorticoid receptor in the central amygdala and normalization by mifepristone treatment.

Despite the enormous negative impact of excessive aggression for individuals and societies, there is a paucity of treatments. Here, using a peripubertal stress model of heightened aggression in rats, we investigated the involvement of the glucocorticoid system and tested the effectiveness of antiglucocorticoid treatment to normalize behavior. We assessed peripubertal stress-induced changes in glucocorticoid (GR) and mineralocorticoid (MR) gene expression in different amygdala nuclei and hippocampus, and report a specific increase in GR mRNA expression in the central amygdala (CeA). Administration of mifepristone (10 mg/kg), a GR antagonist, before stressor exposure at peripuberty prevented the habituation of plasma corticosterone responses observed throughout the stress protocol. This treatment also prevented the increase in aggression and GR expression in the CeA observed in peripubertally stressed rats at adulthood. Viral downregulation of CeA GR expression at adulthood led to reduced aggression. Subsequently, we showed that a brief, 3-day, treatment with mifepristone at adulthood was effective to normalize the abnormal aggression phenotype in peripubertally stressed rats. Our results support a key role for GR actions during peripubertal stress for the long-term programming of heightened aggression. Strikingly, they also support the translational interest of testing the effectiveness of mifepristone treatment to diminish reactive aggression in early adversity-related human psychopathologies.

[Next generation sequencing : a diagnostic tool for inherited immune defects]. / Séquençage à haut débit : outil de diagnostic des déficits immunitaires héréditaires.

Establishing the definitive diagnosis in the case of inherited immune defects (IID) is often challenging because the clinical features can be heterogeneous, atypical and overlapping different disease entities. The next generation sequencing technology (NGS) allows identifying genetic variants that are responsible for the observed clinical presentations. The use of NGS applied to the genes mutated in IIDs or known to be involved in the development, differentiation and regulation of the immune system allows to target hundreds of relevant genes in well characterized patients suspected of carrying inherited immune defects. This approach answers both diagnostic and research needs, facilitates the understanding of the mechanisms that underlie IIDs, and ultimately leads to the discovery of new therapeutic targets.

Etablir un diagnostic précis est souvent difficile dans le cas des déficits immunitaires héréditaires (DIH) car les manifestations cliniques sont hétérogènes et parfois atypiques ou communes à des maladies distinctes. Le séquençage à haut débit, aussi appelé next generation sequencing (NGS), permet d'identifier des variants génétiques responsables de caractéristiques cliniques. Le recours au NGS pour découvrir les gènes mutés dans les DIH ou connus pour être impliqués dans le développement, la différenciation et la régulation du système immunitaire, permet de cibler des centaines de gènes d'intérêt sur des patients suspectés de DIH pour lesquels le tableau clinique est documenté. Cette approche répond à la fois à des besoins de diagnostic et de recherche pour comprendre les mécanismes qui régissent les DIH et découvrir de nouveaux outils ou cibles thérapeutiques.

Neuroligin-2 Expression in the Prefrontal Cortex is Involved in Attention Deficits Induced by Peripubertal Stress.

Emerging evidence indicates that attention deficits, which are frequently observed as core symptoms of neuropsychiatric disorders, may be elicited by early life stress. However, the mechanisms mediating these stress effects remain unknown. The prefrontal cortex (PFC) has been implicated in the regulation of attention, including dysfunctions in GABAergic transmission, and it is highly sensitive to stress. Here, we investigated the involvement of neuroligin-2 (NLGN-2), a synaptic cell adhesion molecule involved in the stabilization and maturation of GABAergic synapses, in the PFC in the link between stress and attention deficits. First, we established that exposure of rats to stress during the peripubertal period impairs attention in the five-choice serial reaction time task and results in reductions in the GABA-synthesizing enzyme glutamic acid decarboxylase in different PFC subregions (ie, prelimbic (PL), infralimbic, and medial and ventral orbitofrontal (OFC) cortex) and in NLGN-2 in the PL cortex. In peripubertally stressed animals, NLGN-2 expression in the PL and OFC cortex correlated with attention measurements. Subsequently, we found that adeno-associated virus-induced rescue of NLGN-2 in the PFC reverses the stress-induced attention deficits regarding omitted trials. Therefore, our findings highlight peripuberty as a period that is highly vulnerable to stress, leading to the development of attention deficits and a dysfunction in the PFC GABAergic system and NLGN-2 expression. Furthermore, NLGN-2 is underscored as a promising target to treat stress-induced cognitive alterations, and in particular attentional deficits as manifested by augmented omissions in a continuous performance task.

CRHR1 links peripuberty stress with deficits in social and stress-coping behaviors.

Stressful life events during childhood and adolescence are important risk factors for the development of psychopathologies later in life. The corticotropin releasing hormone (CRH) and the CRH receptor 1 (CRHR1) have been implicated in the link between early life adversity and adult anxiety and depression, with rodent studies identifying the very early postnatal period as highly susceptible to this programming. Here, we investigated whether stress exposure during the peripubertal period - comprising juvenility and puberty - is effective in inducing long-lasting changes in the expression of CRHR1 and CRHR2 in the hippocampus and amygdala, and whether treating animals with a CRHR1 antagonist following stress exposure could reverse behavioral alterations induced by peripuberty stress. We show that peripuberty stress leads to enhanced expression of the Crhr1, but not Crhr2, gene in the hippocampal CA1 and the central nucleus of the amygdala, in association with social deficits in the social exploration test and increased stress-coping behaviors in the forced swim test. Treatment with the CRHR1 antagonist NBI30775 (10 mg/kg) daily for 1 week (from P43 to P49), immediately following peripuberty stress exposure, prevented the occurrence of those psychopathological behaviors at adulthood. These findings highlight peripuberty as a period of plasticity for the enduring modulation of the CRHR1 system and support a growing body of data implicating the CRHR1 system in the programming effects of early life stress on eventual psychopathology. They also support recent evidence indicating that temporarily tackling CRHR1 during development might represent a therapeutic opportunity to correct behavioral trajectories linking early stress to adult psychopathology.

The interplay of conditional NCAM-knockout and chronic unpredictable stress leads to increased aggression in mice.

The neural cell adhesion molecule (NCAM) is a key regulator of brain plasticity. Substantial evidence indicates that NCAM is down-regulated by exposure to sustained stress and chronic stress seems to lead to increased aggression. In addition, constitutional NCAM deletion in mice has been shown to lead to increased intermale aggression and altered emotionality Forebrain-specific postnatal NCAM knockout was previously shown to impair cognitive function, particularly when animals were exposed to subchronic stress, but the effects on emotional and social behavior remain unclear. In this study, we investigated the potential interplay of a forebrain-specific postnatal NCAM deletion and exposure to different lengths of repeated stress (i.e. subchronic: 14 days; chronic: 29 days) on aggressive and emotional behavior. Our results show that postnatal deletion of NCAM in the forebrain leads to increased aggression and altered emotionality depending on the duration of stress, whereas conditional NCAM knockout has no basal impact on these behaviors. These findings support the involvement of NCAM in the regulation of emotional and aggressive behaviors, suggesting that diminished NCAM expression might be a critical vulnerability factor for the development of these behavioral alterations under repeated exposure to stress.

Hippocampal neuroligin-2 overexpression leads to reduced aggression and inhibited novelty reactivity in rats.

Disturbances of the excitation/inhibition (E/I) balance in the brain were recently suggested as potential factors underlying disorders like autism and schizophrenia resulting in associated behavioral alterations including changes in social and emotional behavior as well as abnormal aggression. Neuronal cell adhesion molecules (nCAMs) and mutations in these genes were found to be strongly implicated in the pathophysiology of these disorders. Neuroligin2 (nlgn2) is a postsynaptic cell adhesion molecule, which is predominantly expressed at inhibitory synapses and required for synapse specification and stabilization. Changes in the expression of nlgn2 were shown to result in alterations of social behavior as well as altered inhibitory synaptic transmission, hence modifying the E/I balance. In our study, we focused on the role of nlgn2 in the dorsal hippocampus in the regulation of emotional and social behaviors. To this purpose, we injected an AAV construct overexpressing nlgn2 in the hippocampus of rats and investigated the effects on behavior and on markers for the E/I ratio. We could show an increase in GAD65, a GABA-synthesizing protein in neuronal terminals, and furthermore, reduced exploration of novel stimuli and less offensive behavior. Our data suggest nlgn2 in the hippocampus to be strongly implicated in maintaining the E/I balance in the brain and thereby modulating social and emotional behavior.

Alpha2-adrenergic receptor activation regulates cortical interneuron migration.

Monoamines such as serotonin and dopamine have been shown to regulate cortical interneuron migration but very little is known regarding noradrenaline. Similarly to other monoamines, noradrenaline is detected during embryonic cortical development and adrenergic receptors are expressed in transient embryonic zones of the pallium that contain migrating neurons. Evidence of a functional role for the adrenergic system in interneuron migration is lacking. In this study we first investigated the expression pattern of adrenergic receptors in mouse cortical interneuron subtypes preferentially derived from the caudal ganglionic eminences, and found that they expressed different subtypes of adrenergic receptors. To directly monitor the effects of adrenergic receptor stimulation on interneuron migration we used time-lapse recordings in cortical slices and observed that alpha2 adrenergic receptors (adra2) receptor activation inhibits the migration of cortical interneurons in a concentration-dependent and reversible manner. Furthermore, we observed that following adra2 activation the directionality of migrating interneurons was significantly modified, suggesting that adra2 stimulation could modulate their responsiveness to guidance cues. Finally the distribution of cortical interneurons was altered in vivo in adra2a/2c-knockout mice. These results support the general hypothesis that adrenergic dysregulation occurring during embryonic development alters cellular processes involved in the formation of cortical circuits.

Loss of intestinal crypt progenitor cells owing to inactivation of both Notch1 and Notch2 is accompanied by derepression of CDK inhibitors p27Kip1 and p57Kip2.

The crucial role of individual Notch receptors and the mechanism by which they maintain intestinal crypt progenitor cells were assessed by using a series of inducible gut-specific Notch mutant mice. We found that Notch1 and Notch2 receptors function redundantly in the gut, as only simultaneous loss of both receptors results in complete conversion of proliferating crypt progenitors into post-mitotic goblet cells. This conversion correlates with the loss of Hes1 expression and derepression of the cyclin-dependent kinase (CDK) inhibitors p27Kip1 and p57Kip2. We also found that the promoter of both CDK inhibitor genes is occupied by the Notch effector Hes1 in wild-type crypt progenitor cells. Thus, our results indicate that Notch-mediated Hes1 expression contributes to the maintenance of the proliferative crypt compartment of the small intestine by transcriptionally repressing two CDK inhibitors.

From gut homeostasis to cancer.

The mammalian intestine has one of the highest turnover rates in the body. The intestinal epithelium is completely renewed in less than a week. It is divided into spatially distinct compartments in the form of finger-like projections and invaginations that are dedicated to specific functions. Intestinal cells are constantly produced from a stem cell reservoir that gives rise to proliferating transient amplifying cells, which subsequently differentiate and migrate to the correct compartment before dying after having fulfilled their physiological function. In recent years, a substantial body of evidence has accumulated to support the concept that signaling pathways known to be crucial for embryonic development of multiple organisms play a critical role in tightly regulating and controlling the self-renewing process of the intestine. Moreover, the same pathways appear to be deregulated in several hereditary and sporadic colorectal cancer syndromes due to activating and/or inactivating mutations of key components of such pathways. In this review we discuss recent findings demonstrating that differentiation and homeostasis of the intestine are controlled by developmental pathways such as Wnt, Notch, TGF-beta and Hedgehog, and illustrate how their deregulation contributes to intestinal neoplasia.

Synthetic RGD-containing alpha-helical coiled coil peptides promote integrin-dependent cell adhesion.

Integrin receptors are the main mediators of cell adhesion to the extracellular matrix. They bind to their ligands by interacting with short amino acid sequences, such as the RGD sequence. Soluble, small RGD-based peptides have been used to block integrin-binding to ligands, thereby interfering with cell adhesion, migration and survival, while substrate-immobilized RGD sequences have been used to enhance cell binding to artificial surfaces. This approach has several important medical applications, e.g. in suppression of tumor angiogenesis or stimulation of bone formation around implants. However, the relatively weak affinity of short RGD-containing peptides often results in incomplete integrin inhibition or ineffective ligation. In this work, we designed and synthesized several new multivalent RGD-containing molecules and tested their ability to inhibit or to promote integrin-dependent cell adhesion when used in solution or immobilized on substrates, respectively. These molecules consist of an oligomeric structure formed by alpha-helical coiled coil peptides fused at their amino-terminal ends with an RGD-containing fragment. When immobilized on a substrate, these peptides specifically promoted integrin alphaVbeta3-dependent cell adhesion, but when used in solution, they blocked alphaVbeta3-dependent cell adhesion to the natural substrates fibronectin and vitronectin. One of the peptides was nearly 10-fold more efficient than fibronectin or vitronectin in promoting cell adhesion, and almost 100-fold more efficient than a linear RGD tripeptide in blocking adhesion. These results indicate that alpha-helical coiled coil peptides carrying an amino-terminal RGD motif can be used as soluble antagonists or surface-immobilized agonists to efficiently inhibit or promote integrin alphaVbeta3-mediated cell adhesion, respectively.

Notch/gamma-secretase inhibition turns proliferative cells in intestinal crypts and adenomas into goblet cells.

The self-renewing epithelium of the small intestine is ordered into stem/progenitor crypt compartments and differentiated villus compartments. Recent evidence indicates that the Wnt cascade is the dominant force in controlling cell fate along the crypt-villus axis. Here we show a rapid, massive conversion of proliferative crypt cells into post-mitotic goblet cells after conditional removal of the common Notch pathway transcription factor CSL/RBP-J. We obtained a similar phenotype by blocking the Notch cascade with a gamma-secretase inhibitor. The inhibitor also induced goblet cell differentiation in adenomas in mice carrying a mutation of the Apc tumour suppressor gene. Thus, maintenance of undifferentiated, proliferative cells in crypts and adenomas requires the concerted activation of the Notch and Wnt cascades. Our data indicate that gamma-secretase inhibitors, developed for Alzheimer's disease, might be of therapeutic benefit in colorectal neoplastic disease.