Reconstruction algorithms for DNA-storage systems.

Motivated by DNA storage systems, this work presents the DNA reconstruction problem, in which a length-n string, is passing through the DNA-storage channel, which introduces deletion, insertion and substitution errors. This channel generates multiple noisy copies of the transmitted string which are called traces. A DNA reconstruction algorithm is a mapping which receives t traces as an input and produces an estimation of the original string. The goal in the DNA reconstruction problem is to minimize the edit distance between the original string and the algorithm's estimation. In this work, we present several new algorithms for this problem. Our algorithms look globally on the entire sequence of the traces and use dynamic programming algorithms, which are used for the shortest common supersequence and the longest common subsequence problems, in order to decode the original string. Our algorithms do not require any limitations on the input and the number of traces, and more than that, they perform well even for error probabilities as high as 0.27. The algorithms have been tested on simulated data, on data from previous DNA storage experiments, and on a new synthesized dataset, and are shown to outperform previous algorithms in reconstruction accuracy.

Genomic Markers Associated with Cytomegalovirus DNAemia in Kidney Transplant Recipients.

Human cytomegalovirus (CMV) is a major pathogen after solid organ transplantation, leading to high morbidity and mortality. Transplantation from a CMV-seropositive donor to a CMV-seronegative recipient (D+/R-) is associated with high risk of CMV disease. However, that risk is not uniform, suggesting a role for host factors in immune control of CMV. To identify host genetic factors that control CMV DNAemia post transplantation, we performed a whole-exome association study in two cohorts of D+/R- kidney transplant recipients. Quantitative CMV DNA was measured for at least one year following transplantation. Several CMV-protective single-nucleotide polymorphisms (SNPs) were identified in the first cohort (72 patients) but were not reproducible in the second cohort (126 patients). A meta-analysis of both cohorts revealed several SNPs that were significantly associated with protection from CMV DNAemia. The copy number variation of several genes was significantly different between recipients with and without CMV DNAemia. Amongst patients with CMV DNAemia in the second cohort, several variants of interest (p < 5 × 10-5), the most common of which was NLRC5, were associated with peak viral load. We provide new predictive genetic markers for protection of CMV DNAemia. These markers should be validated in larger cohorts.

Unexpected gender differences in progressive supranuclear palsy reveal efficacy for davunetide in women.

Progressive supranuclear palsy (PSP) is a pure tauopathy, implicating davunetide, enhancing Tau-microtubule interaction, as an ideal drug candidate. However, pooling patient data irrespective of sex concluded no efficacy. Here, analyzing sex-dependency in a 52 week-long- PSP clinical trial (involving over 200 patients) demonstrated clear baseline differences in brain ventricular volumes, a secondary endpoint. Dramatic baseline ventricular volume-dependent/volume increase correlations were observed in 52-week-placebo-treated females (r = 0.74, P = 2.36-9), whereas davunetide-treated females (like males) revealed no such effects. Assessment of primary endpoints, by the PSP Rating Scale (PSPRS) and markedly more so by the Schwab and England Activities of Daily Living (SEADL) scale, showed significantly faster deterioration in females, starting at trial week 13 (P = 0.01, and correlating with most other endpoints by week 52). Twice daily davunetide treatments slowed female disease progression and revealed significant protection according to the SEADL scale as early as at 39 weeks (P = 0.008), as well as protection of the bulbar and limb motor domains considered by the PSPRS, including speaking and swallowing difficulties caused by brain damage, and deterioration of fine motor skills, respectably (P = 0.01), at 52 weeks. Furthermore, at 52 weeks of trial, the exploratory Geriatric Depression Scale (GDS) significantly correlated with the SEADL scale deterioration in the female placebo group and demonstrated davunetide-mediated protection of females. Female-specific davunetide-mediated protection of ventricular volume corresponded to clinical efficacy. Together with the significantly slower disease progression seen in men, the results reveal sex-based drug efficacy differences, demonstrating the neuroprotective and disease-modifying impact of davunetide treatment for female PSP patients.

mRNA splicing is modulated by intronic microRNAs.

Splicing of transcripts is catalyzed by the spliceosome, a mega-complex consisting of hundreds of proteins and five snRNAs, which employs direct interactions. When U1 snRNA forms high-affinity binding, namely more than eight base pairs, with the 5'SS, the result is usually a suppressing effect on the splicing activity. This likely occurs due to the inefficient unwinding of U1/5'SS base-pairing or other regulatory obstructions. Here, we show in vitro and in patient-derived cell lines that pre-microRNAs can modulate the splicing reaction by interacting with U1 snRNA. This leads to reduced binding affinity to the 5'SS, and hence promotes the inclusion of exons containing 5'SS, despite sequence-based high affinity to U1. Application of the mechanism resulted in correction of the splicing defect in the disease-causing VCAN gene from an individual with Wagner syndrome. This pre-miRNA/U1 interaction can regulate the expression of alternatively spliced exons, thus extending the scope of mechanisms regulating splicing.

Hippocampal differential expression underlying the neuroprotective effect of delta-9-tetrahydrocannabinol microdose on old mice.

Delta-9-tetrahydrocannabinol (THC) is the primary psychoactive compound of the cannabis plant and an exogenous ligand of the endocannabinoid system. In previous studies, we demonstrated that a single microdose of THC (0.002 mg/kg, 3-4 orders of magnitude lower than the standard dose for rodents) exerts distinct, long-term neuroprotection in model mice subjected to acute neurological insults. When administered to old, healthy mice, the THC microdose induced remarkable long-lasting (weeks) improvement in a wide range of cognitive functions, including significant morphological and biochemical brain alterations. To elucidate the mechanisms underlying these effects, we analyzed the gene expression of hippocampal samples from the model mice. Samples taken 5 days after THC treatment showed significant differential expression of genes associated with neurogenesis and brain development. In samples taken 5 weeks after treatment, the transcriptional signature was shifted to that of neuronal differentiation and survival. This study demonstrated the use of hippocampal transcriptome profiling in uncovering the molecular basis of the atypical, anti-aging effects of THC microdose treatment in old mice.

Immunosuppression as a Hub for SARS-CoV-2 Mutational Drift.

The clinical course of coronavirus disease 2019 (COVID-19), caused by severe acute respiratory syndrome coronavirus-2 (SARS-CoV-2), is largely determined by host factors, with a wide range of outcomes. Despite an extensive vaccination campaign and high rates of infection worldwide, the pandemic persists, adapting to overcome antiviral immunity acquired through prior exposure. The source of many such major adaptations is variants of concern (VOCs), novel SARS-CoV-2 variants produced by extraordinary evolutionary leaps whose origins remain mostly unknown. In this study, we tested the influence of factors on the evolutionary course of SARS-CoV-2. Electronic health records of individuals infected with SARS-CoV-2 were paired to viral whole-genome sequences to assess the effects of host clinical parameters and immunity on the intra-host evolution of SARS-CoV-2. We found slight, albeit significant, differences in SARS-CoV-2 intra-host diversity, which depended on host parameters such as vaccination status and smoking. Only one viral genome had significant alterations as a result of host parameters; it was found in an immunocompromised, chronically infected woman in her 70s. We highlight the unusual viral genome obtained from this woman, which had an accelerated mutational rate and an excess of rare mutations, including near-complete truncating of the accessory protein ORF3a. Our findings suggest that the evolutionary capacity of SARS-CoV-2 during acute infection is limited and mostly unaffected by host characteristics. Significant viral evolution is seemingly exclusive to a small subset of COVID-19 cases, which typically prolong infections in immunocompromised patients. In these rare cases, SARS-CoV-2 genomes accumulate many impactful and potentially adaptive mutations; however, the transmissibility of such viruses remains unclear.

Comprehensive Genetic Analysis of Druze Provides Insights into Carrier Screening.

BACKGROUND: Druze individuals, like many genetically homogeneous and isolated populations, harbor recurring pathogenic variants (PV) in autosomal recessive (AR) disorders. METHODS: Variant calling of whole-genome sequencing (WGS) of 40 Druze from the Human Genome Diversity Project (HGDP) was performed (HGDP-cohort). Additionally, we performed whole exome sequencing (WES) of 118 Druze individuals: 38 trios and 2 couples, representing geographically distinct clans (WES-cohort). Rates of validated PV were compared with rates in worldwide and Middle Eastern populations, from the gnomAD and dbSNP datasets. RESULTS: Overall, 34 PVs were identified: 30 PVs in genes underlying AR disorders, 3 additional PVs were associated with autosomal dominant (AD) disorders, and 1 PV with X-linked-dominant inherited disorder in the WES cohort. CONCLUSIONS: The newly identified PVs associated with AR conditions should be considered for incorporation into prenatal-screening options offered to Druze individuals after an extension and validation of the results in a larger study.

Primary Melanoma miRNA Trafficking Induces Lymphangiogenesis.

Melanoma, the deadliest cutaneous tumor, initiates within the epidermis; during progression, cells invade into the dermis and become metastatic through the lymphatic and blood system. Before melanoma cell invasion into the dermis, an increased density of dermal lymphatic vessels is observed, generated by a mechanism which is not fully understood. In this study, we show that, while at the primary epidermal stage (in situ), melanoma cells secrete extracellular vesicles termed melanosomes, which are uptaken by dermal lymphatic cells, leading to transcriptional and phenotypic pro-lymphangiogenic changes. Mechanistically, melanoma-derived melanosomes traffic mature let-7i to lymphatic endothelial cells, which mediate pro-lymphangiogenic phenotypic changes by the induction of type I IFN signaling. Furthermore, transcriptome analysis upon treatment with melanosomes or let-7i reveals the enhancement of IFI6 expression in lymphatic cells. Because melanoma cells metastasize primarily via lymphatic vessels, our data suggest that blocking lymphangiogenesis by repressing either melanosome release or type I IFN signaling will prevent melanoma progression to the deadly metastatic stage.

Multiple Copies of microRNA Binding Sites in Long 3'UTR Variants Regulate Axonal Translation.

Rapid responses to changes within subcellular compartments of highly polarized cells, such as neuron axons, depend on local translation and post-transcriptional regulation. The mechanism by which microRNAs (miRNAs) regulate this process is not fully understood. Here, using live cell imaging and RNA sequencing analysis, we demonstrated how miRNAs can differentially control hundreds of transcripts at the subcellular level. We demonstrated that the seed match length of the miRNA target-sequence regulates both mRNA stability and protein translation rates. While longer seed matches have an increased inhibitory effect, transcriptome analysis did not reveal differences in seed match length between axonal and somata mRNAs of motor neurons. However, mRNA variants with longer 3'UTR are enriched in axons and contain multiple repeats of specific miRNA target sequences. Finally, we demonstrated that the long 3'UTR mRNA variant of the motor protein Kif5b is enriched explicitly in motor neuron axons and contains multiple sequence repeats for binding miR-129-5p. This subsequently results in the differential post-transcriptional regulation of kif5b and its synthesis in axons. Thus, we suggest that the number of miRNA binding sites at the 3'UTR of the mRNA, rather than the miRNA seed match length, regulates the axonal transcriptome.

Elevated Expression of RGS2 May Underlie Reduced Olfaction in COVID-19 Patients.

Anosmia is common in COVID-19 patients, lasting for weeks or months following recovery. The biological mechanism underlying olfactory deficiency in COVID-19 does not involve direct damage to nasal olfactory neurons, which do not express the proteins required for SARS-CoV-2 infection. A recent study suggested that anosmia results from downregulation of olfactory receptors. We hypothesized that anosmia in COVID-19 may also reflect SARS-CoV-2 infection-driven elevated expression of regulator of G protein signaling 2 (RGS2), a key regulator of odorant receptors, thereby silencing their signaling. To test our hypothesis, we analyzed gene expression of nasopharyngeal swabs from SARS-CoV-2 positive patients and non-infected controls (two published RNA-sequencing datasets, 580 individuals). Our analysis found upregulated RGS2 expression in SARS-CoV-2 positive patients (FC = 14.5, Padj = 1.69 × 10-5 and FC = 2.4; Padj = 0.001, per dataset). Additionally, RGS2 expression was strongly correlated with PTGS2, IL1B, CXCL8, NAMPT and other inflammation markers with substantial upregulation in early infection. These observations suggest that upregulated expression of RGS2 may underlie anosmia in COVID-19 patients. As a regulator of numerous G-protein coupled receptors, RGS2 may drive further neurological symptoms of COVID-19. Studies are required for clarifying the cellular mechanisms by which SARS-CoV-2 infection drives the upregulation of RGS2 and other genes implicated in inflammation. Insights on these pathway(s) may assist in understanding anosmia and additional neurological symptoms reported in COVID-19 patients.

Transient cell-in-cell formation underlies tumor relapse and resistance to immunotherapy.

Despite the remarkable successes of cancer immunotherapies, the majority of patients will experience only partial response followed by relapse of resistant tumors. While treatment resistance has frequently been attributed to clonal selection and immunoediting, comparisons of paired primary and relapsed tumors in melanoma and breast cancers indicate that they share the majority of clones. Here, we demonstrate in both mouse models and clinical human samples that tumor cells evade immunotherapy by generating unique transient cell-in-cell structures, which are resistant to killing by T cells and chemotherapies. While the outer cells in this cell-in-cell formation are often killed by reactive T cells, the inner cells remain intact and disseminate into single tumor cells once T cells are no longer present. This formation is mediated predominantly by IFNγ-activated T cells, which subsequently induce phosphorylation of the transcription factors signal transducer and activator of transcription 3 (STAT3) and early growth response-1 (EGR-1) in tumor cells. Indeed, inhibiting these factors prior to immunotherapy significantly improves its therapeutic efficacy. Overall, this work highlights a currently insurmountable limitation of immunotherapy and reveals a previously unknown resistance mechanism which enables tumor cells to survive immune-mediated killing without altering their immunogenicity.

Cancer immunotherapies use the body's own immune system to fight off cancer. But, despite some remarkable success stories, many patients only see a temporary improvement before the immunotherapy stops being effective and the tumours regrow. It is unclear why this occurs, but it may have to do with how the immune system attacks cancer cells. Immunotherapies aim to activate a special group of cells known as killer T-cells, which are responsible for the immune response to tumours. These cells can identify cancer cells and inject toxic granules through their membranes, killing them. However, killer T-cells are not always effective. This is because cancer cells are naturally good at avoiding detection, and during treatment, their genes can mutate, giving them new ways to evade the immune system. Interestingly, when scientists analysed the genes of tumour cells before and after immunotherapy, they found that many of the genes that code for proteins recognized by T-cells do not change significantly. This suggests that tumours' resistance to immune attack may be physical, rather than genetic. To investigate this hypothesis, Gutwillig et al. developed several mouse tumour models that stop responding to immunotherapy after initial treatment. Examining cells from these tumours revealed that when the immune system attacks, they reorganise by getting inside one another. This allows some cancer cells to hide under many layers of cell membrane. At this point killer T-cells can identify and inject the outer cell with toxic granules, but it cannot reach the cells inside. This ability of cancer cells to hide within one another relies on them recognising when the immune system is attacking. This happens because the cancer cells can detect certain signals released by the killer T-cells, allowing them to hide. Gutwillig et al. identified some of these signals, and showed that blocking them stopped cancer cells from hiding inside each other, making immunotherapy more effective. This new explanation for how cancer cells escape the immune system could guide future research and lead to new cancer treatments, or approaches to boost existing treatments. Understanding the process in more detail could allow scientists to prevent it from happening, by revealing which signals to block, and when, for best results.

Transcriptomic Analysis of Human Fragile X Syndrome Neurons Reveals Neurite Outgrowth Modulation by the TGFß/BMP Pathway.

Fragile X Syndrome (FXS) is the main genetic reason for intellectual disability and is caused by the silencing of fragile X mental retardation protein (FMRP), an RNA-binding protein regulating the translation of many neuronal mRNAs. Neural differentiation of FX human embryonic stem cells (hESC) mimics the neurodevelopment of FXS fetuses and thus serves as a good model to explore the mechanisms underlining the development of FXS. Isogenic hESC clones with and without the FX mutation that share the same genetic background were in vitro differentiated into neurons, and their transcriptome was analyzed by RNA sequencing. FX neurons inactivating FMR1 expression presented delayed neuronal development and maturation, concomitant with dysregulation of the TGFß/BMP signaling pathway, and genes related to the extracellular matrix. Migration assay showed decreased neurite outgrowth in FX neurons that was rescued by inhibition of the TGFß/BMP signaling pathway. Our results provide new insights into the molecular pathway by which loss of FMRP affects neuronal network development. In FX neurons, the lack of FMRP dysregulates members of the BMP signaling pathway associated with ECM organization which, in a yet unknown mechanism, reduces the guidance of axonal growth cones, probably leading to the aberrant neuronal network function seen in FXS.

Lower serum alpha 1 antitrypsin levels in patients with severe COVID-19 compared with patients hospitalized due to non-COVID-19 pneumonia.

INTRODUCTION: Alpha 1 antitrypsin (A1AT) is the major human blood serine protease inhibitor. Transmembrane serine protease 2 (TMPRSS2), which is crucial for SARS-CoV-2 cell entry, is inhibited by A1AT. Therefore, we hypothesized that individuals with diminished levels of A1AT may be more prone to SARS-CoV-2 infection and severe COVID-19 disease. Our aim in this study was to evaluate the level of A1AT in hospitalized COVID-19 patients in comparison to hospitalized patients with non-COVID-19 pneumonia. METHODS: We conducted an observational prospective study between October 2020 and April 2021 in Rabin Medical Centre in Israel. A1AT levels were measured from the routine serum samples of hospitalized patients with COVID-19 and non-COVID-19 pneumonia (control group). The primary outcome was A1AT level, secondary outcomes were clinical outcomes and predictors of morality. RESULTS: Overall, 145 patients were included in the study, 98 in the COVID-19 group and 47 in the control group. The median A1AT level was 222 mg/dL (interquartile range (IQR) 188-269) and 258 mg/dL (IQR 210-281) in the COVID-19 and control groups, respectively (p = .045). Multivariate analysis for independent risk factors for mortality among COVID-19 patients showed that diabetes mellitus (p = .02), older age (p = .04), and high A1AT levels (p = .04) were all associated with increased mortality. CONCLUSION: Patients admitted due to severe COVID-19 had lower A1AT levels in comparison to patients admitted due to non-COVID pneumonia. This observation may suggest an association between mildly diminished A1AT and higher risk of SARS-CoV-2 infection with severe COVID-19 disease.

Oral microbiota signatures in post-traumatic stress disorder (PTSD) veterans.

Post-traumatic stress disorder (PTSD) represents a global public health concern, affecting about 1 in 20 individuals. The symptoms of PTSD include intrusiveness (involuntary nightmares or flashbacks), avoidance of traumatic memories, negative alterations in cognition and mood (such as negative beliefs about oneself or social detachment), increased arousal and reactivity with irritable reckless behavior, concentration problems, and sleep disturbances. PTSD is also highly comorbid with anxiety, depression, and substance abuse. To advance the field from subjective, self-reported psychological measurements to objective molecular biomarkers while considering environmental influences, we examined a unique cohort of Israeli veterans who participated in the 1982 Lebanon war. Non-invasive oral 16S RNA sequencing was correlated with psychological phenotyping. Thus, a microbiota signature (i.e., decreased levels of the bacteria sp_HMT_914, 332 and 871 and Noxia) was correlated with PTSD severity, as exemplified by intrusiveness, arousal, and reactivity, as well as additional psychopathological symptoms, including anxiety, hostility, memory difficulties, and idiopathic pain. In contrast, education duration correlated with significantly increased levels of sp_HMT_871 and decreased levels of Bacteroidetes and Firmicutes, and presented an inverted correlation with adverse psychopathological measures. Air pollution was positively correlated with PTSD symptoms, psychopathological symptoms, and microbiota composition. Arousal and reactivity symptoms were correlated with reductions in transaldolase, an enzyme controlling a major cellular energy pathway, that potentially accelerates aging. In conclusion, the newly discovered bacterial signature, whether an outcome or a consequence of PTSD, could allow for objective soldier deployment and stratification according to decreases in sp_HMT_914, 332, 871, and Noxia levels, coupled with increases in Bacteroidetes levels. These findings also raise the possibility of microbiota pathway-related non-intrusive treatments for PTSD.

TWIST1 methylation by SETD6 selectively antagonizes LINC-PINT expression in glioma.

Gliomas are one of the most common and lethal brain tumors among adults. One process that contributes to glioma progression and recurrence is the epithelial to mesenchymal transition (EMT). EMT is regulated by a set of defined transcription factors which tightly regulate this process, among them is the basic helix-loop-helix family member, TWIST1. Here we show that TWIST1 is methylated on lysine-33 at chromatin by SETD6, a methyltransferase with expression levels correlating with poor survival in glioma patients. RNA-seq analysis in U251 glioma cells suggested that both SETD6 and TWIST1 regulate cell adhesion and migration processes. We further show that TWIST1 methylation attenuates the expression of the long-non-coding RNA, LINC-PINT, thereby promoting EMT in glioma. Mechanistically, TWIST1 methylation represses the transcription of LINC-PINT by increasing the occupancy of EZH2 and the catalysis of the repressive H3K27me3 mark at the LINC-PINT locus. Under un-methylated conditions, TWIST1 dissociates from the LINC-PINT locus, allowing the expression of LINC-PINT which leads to increased cell adhesion and decreased cell migration. Together, our findings unravel a new mechanistic dimension for selective expression of LINC-PINT mediated by TWIST1 methylation.

Structure-function conservation between the methyltransferases SETD3 and SETD6.

Among the protein lysine methyltransferases family members, it appears that SETD6 is highly similar and closely related to SETD3. The two methyltransferases show high similarity in their structure, which raised the hypothesis that they share cellular functions. Using a proteomic screen, we identified 52 shared interacting-proteins. Gene Ontology (GO) analysis of the shared proteins revealed significant enrichment of proteins involved in transcription. Our RNA-seq data of SETD6 KO and SETD3 KO HeLa cells identified ∼100 up-regulated and down-regulated shared genes. We have also identified a substantial number of genes that changed dramatically in the double KO cells but did not significantly change in the single KO cells. GO analysis of these genes revealed enrichment of apoptotic genes. Accordingly, we show that the double KO cells displayed high apoptotic levels, suggesting that SETD6 and SETD3 inhibit apoptosis. Collectively, our data strongly suggest a functional link between SETD6 and SETD3 in the regulation of apoptosis.

Population differences in antibody response to SARS-CoV-2 infection and BNT162b2 vaccination.

The concentration of SARS-CoV-2-specific serum antibodies, elicited by vaccination or infection, is a primary determinant of anti-viral immunity, which correlates with protection against infection and COVID-19. Serum samples were obtained from 25 897 participants and assayed for anti-SARS-CoV-2 spike protein RBD IgG antibodies. The cohort was composed of newly vaccinated BNT162b2 recipients, in the first month or 6 months after vaccination, COVID-19 patients and a general sample of the Israeli population. Antibody levels of BNT162b2 vaccine recipients were negatively correlated with age, with a prominent decrease in recipients over 55 years old, which was most significant in males. This trend was observable within the first month and 6 months after vaccination, while younger participants were more likely to maintain stable levels of serum antibodies. The antibody concentration of participants previously infected with SARS-CoV-2 was lower than the vaccinated and had a more complex, non-linear relation to age, sex and COVID-19 symptoms. Taken together, our data supports age and sex as primary determining factors for both the magnitude and durability of humoral response to SARS-CoV-2 infection and the COVID-19 vaccine. Our results could inform vaccination policies, prioritizing the most susceptible populations for repeated vaccination.

Altered White Matter and microRNA Expression in a Murine Model Related to Williams Syndrome Suggests That miR-34b/c Affects Brain Development via Ptpru and Dcx Modulation.

Williams syndrome (WS) is a multisystem neurodevelopmental disorder caused by a de novo hemizygous deletion of ~26 genes from chromosome 7q11.23, among them the general transcription factor II-I (GTF2I). By studying a novel murine model for the hypersociability phenotype associated with WS, we previously revealed surprising aberrations in myelination and cell differentiation properties in the cortices of mutant mice compared to controls. These mutant mice had selective deletion of Gtf2i in the excitatory neurons of the forebrain. Here, we applied diffusion magnetic resonance imaging and fiber tracking, which showed a reduction in the number of streamlines in limbic outputs such as the fimbria/fornix fibers and the stria terminalis, as well as the corpus callosum of these mutant mice compared to controls. Furthermore, we utilized next-generation sequencing (NGS) analysis of cortical small RNAs' expression (RNA-Seq) levels to identify altered expression of microRNAs (miRNAs), including two from the miR-34 cluster, known to be involved in prominent processes in the developing nervous system. Luciferase reporter assay confirmed the direct binding of miR-34c-5p to the 3'UTR of PTPRU-a gene involved in neural development that was elevated in the cortices of mutant mice relative to controls. Moreover, we found an age-dependent variation in the expression levels of doublecortin (Dcx)-a verified miR-34 target. Thus, we demonstrate the substantial effect a single gene deletion can exert on miRNA regulation and brain structure, and advance our understanding and, hopefully, treatment of WS.

Novel ADNP Syndrome Mice Reveal Dramatic Sex-Specific Peripheral Gene Expression With Brain Synaptic and Tau Pathologies.

BACKGROUND: ADNP is essential for embryonic development. As such, de novo ADNP mutations lead to an intractable autism/intellectual disability syndrome requiring investigation. METHODS: Mimicking humans, CRISPR (clustered regularly interspaced short palindromic repeats)-Cas9 editing produced mice carrying heterozygous Adnp p.Tyr718∗ (Tyr), a paralog of the most common ADNP syndrome mutation. Phenotypic rescue was validated by treatment with the microtubule/autophagy-protective ADNP fragment NAPVSIPQ (NAP). RESULTS: RNA sequencing of spleens, representing a peripheral biomarker source, revealed Tyr-specific sex differences (e.g., cell cycle), accentuated in females (with significant effects on antigen processing and cellular senescence) and corrected by NAP. Differentially expressed, NAP-correctable transcripts, including the autophagy and microbiome resilience-linked FOXO3, were also deregulated in human patient-derived ADNP-mutated lymphoblastoid cells. There were also Tyr sex-specific microbiota signatures. Phenotypically, Tyr mice, similar to patients with ADNP syndrome, exhibited delayed development coupled with sex-dependent gait defects. Speech acquisition delays paralleled sex-specific mouse syntax abnormalities. Anatomically, dendritic spine densities/morphologies were decreased with NAP amelioration. These findings were replicated in the Adnp+/- mouse, including Foxo3 deregulation, required for dendritic spine formation. Grooming duration and nociception threshold (autistic traits) were significantly affected only in males. Early-onset tauopathy was accentuated in males (hippocampus and visual cortex), mimicking humans, and was paralleled by impaired visual evoked potentials and correction by acute NAP treatment. CONCLUSIONS: Tyr mice model ADNP syndrome pathology. The newly discovered ADNP/NAP target FOXO3 controls the autophagy initiator LC3 (microtubule-associated protein 1 light chain 3), with known ADNP binding to LC3 augmented by NAP, protecting against tauopathy. NAP amelioration attests to specificity, with potential for drug development targeting accessible biomarkers.

Transcriptional Profiling of Mouse Eosinophils Identifies Distinct Gene Signatures Following Cellular Activation.

Eosinophils are multifunctional, evolutionary conserved leukocytes that are involved in a plethora of responses ranging from regulation of tissue homeostasis to host defense and cancer. Eosinophils have been studied mostly in the context of Type 2 inflammatory responses such as those found in allergy. Nonetheless, it is now evident that they participate in Type 1 inflammatory responses and can respond to Type 1 cytokines such as IFN-γ. Recent data suggest that the pleotropic roles of eosinophils are due to heterogeneous responses to environmental cues. Despite this, the activation profile of eosinophils, in response to various stimuli is yet to be defined. To better understand the transcriptional spectrum of eosinophil activation, we exposed eosinophils to Type 1 (e.g. IFN-γ, E. coli) vs. Type 2 (e.g. IL-4) conditions and subjected them to global RNA sequencing. Our analyses show that IL-4, IFN-γ, E. coli and IFN-γ in the presence of E. coli (IFN-γ/E. coli)-stimulated eosinophils acquire distinct transcriptional profiles, which polarize them towards what we termed Type 1 and Type 2 eosinophils. Bioinformatics analyses using Gene Ontology based on biological processes revealed that different stimuli induced distinct pathways in eosinophils. These pathways were confirmed using functional assays by assessing cytokine/chemokine release (i.e. CXCL9, CCL24, TNF-α and IL-6) from eosinophils following activation. In addition, analysis of cell surface markers highlighted CD101 and CD274 as potential cell surface markers that distinguish between Type 1 and Type 2 eosinophils, respectively. Finally, the transcriptome signature of Type 1 eosinophils resembled that of eosinophils that were obtained from mice with experimental colitis whereas the transcriptome signature of Type 2 eosinophils resembled that of eosinophils from experimental asthma. Our data demonstrate that eosinophils are polarized to distinct "Type 1" and "Type 2" phenotypes following distinct stimulations. These findings provide fundamental knowledge regarding the heterogeneity of eosinophils and support the presence of transcriptional differences between Type 1 and Type 2 cells that are likely reflected by their pleotropic activities in diverse disease settings.