LINE-1 retrotransposition in the nervous system.

Long interspersed element-1 (LINE-1 or L1) is a repetitive DNA retrotransposon capable of duplication by a copy-and-paste genetic mechanism. Scattered throughout mammalian genomes, L1 is typically quiescent in most somatic cell types. In developing neurons, however, L1 can express and retrotranspose at high frequency. The L1 element can insert into various genomic locations including intragenic regions. These insertions can alter the dynamic of the neuronal transcriptome by changing the expression pattern of several nearby genes. The consequences of L1 genomic alterations in somatic cells are still under investigation, but the high level of mutagenesis within neurons suggests that each neuron is genetically unique. Furthermore, some neurological diseases, such as Rett syndrome and ataxia telangiectasia, misregulate L1 retrotransposition, which could contribute to some pathological aspects. In this review, we survey the literature related to neurodevelopmental retrotransposition and discuss possible relevance to neuronal function, evolution, and neurological disease.

dRFEtools: dynamic recursive feature elimination for omics.

MOTIVATION: Advances in technology have generated larger omics datasets with potential applications for machine learning. In many datasets, however, cost and limited sample availability result in an excessively higher number of features as compared to observations. Moreover, biological processes are associated with networks of core and peripheral genes, while traditional feature selection approaches capture only core genes. RESULTS: To overcome these limitations, we present dRFEtools that implements dynamic recursive feature elimination (RFE), reducing computational time with high accuracy compared to standard RFE, expanding dynamic RFE to regression algorithms, and outputting the subsets of features that hold predictive power with and without peripheral features. dRFEtools integrates with scikit-learn (the popular Python machine learning platform) and thus provides new opportunities for dynamic RFE in large-scale omics data while enhancing its interpretability. AVAILABILITY AND IMPLEMENTATION: dRFEtools is freely available on PyPI at https://pypi.org/project/drfetools/ or on GitHub https://github.com/LieberInstitute/dRFEtools, implemented in Python 3, and supported on Linux, Windows, and Mac OS.

Cytoplasmic synthesis of endogenous Alu complementary DNA via reverse transcription and implications in age-related macular degeneration.

Alu retroelements propagate via retrotransposition by hijacking long interspersed nuclear element-1 (L1) reverse transcriptase (RT) and endonuclease activities. Reverse transcription of Alu RNA into complementary DNA (cDNA) is presumed to occur exclusively in the nucleus at the genomic integration site. Whether Alu cDNA is synthesized independently of genomic integration is unknown. Alu RNA promotes retinal pigmented epithelium (RPE) death in geographic atrophy, an untreatable type of age-related macular degeneration. We report that Alu RNA-induced RPE degeneration is mediated via cytoplasmic L1-reverse-transcribed Alu cDNA independently of retrotransposition. Alu RNA did not induce cDNA production or RPE degeneration in L1-inhibited animals or human cells. Alu reverse transcription can be initiated in the cytoplasm via self-priming of Alu RNA. In four health insurance databases, use of nucleoside RT inhibitors was associated with reduced risk of developing atrophic macular degeneration (pooled adjusted hazard ratio, 0.616; 95% confidence interval, 0.493-0.770), thus identifying inhibitors of this Alu replication cycle shunt as potential therapies for a major cause of blindness.

Developmental excitation-inhibition imbalance underlying psychoses revealed by single-cell analyses of discordant twins-derived cerebral organoids.

Despite extensive genetic and neuroimaging studies, detailed cellular mechanisms underlying schizophrenia and bipolar disorder remain poorly understood. Recent progress in single-cell RNA sequencing (scRNA-seq) technologies enables identification of cell-type-specific pathophysiology. However, its application to psychiatric disorders is challenging because of methodological difficulties in analyzing human brains and the confounds due to a lifetime of illness. Brain organoids derived from induced pluripotent stem cells (iPSCs) of the patients are a powerful avenue to investigate the pathophysiological processes. Here, we generated iPSC-derived cerebral organoids from monozygotic twins discordant for psychosis. scRNA-seq analysis of the organoids revealed enhanced GABAergic specification and reduced cell proliferation following diminished Wnt signaling in the patient, which was confirmed in iPSC-derived forebrain neuronal cells. Two additional monozygotic twin pairs discordant for schizophrenia also confirmed the excess GABAergic specification of the patients' neural progenitor cells. With a well-controlled genetic background, our data suggest that unbalanced specification of excitatory and inhibitory neurons during cortical development underlies psychoses.

Altered serotonergic circuitry in SSRI-resistant major depressive disorder patient-derived neurons.

Disrupted serotonergic neurotransmission has long been implicated in major depressive disorder (MDD), for which selective serotonin reuptake inhibitors (SSRIs) are the first line of treatment. However, a significant percentage of patients remain SSRI-resistant and it is unclear whether and how alterations in serotonergic neurons contribute to SSRI resistance in these patients. Induced pluripotent stem cells (iPSCs) facilitate the study of patient-specific neural subtypes that are typically inaccessible in living patients, enabling the discovery of disease-related phenotypes. In our study of a well-characterized cohort of over 800 MDD patients, we generated iPSCs and serotonergic neurons from three extreme SSRI-remitters (R) and SSRI-nonremitters (NR). We studied serotonin (5-HT) biochemistry and observed no significant differences in 5-HT release and reuptake or in genes related to 5-HT biochemistry. NR patient-derived serotonergic neurons exhibited altered neurite growth and morphology downstream of lowered expression of key Protocadherin alpha genes as compared to healthy controls and Rs. Furthermore, knockdown of Protocadherin alpha genes directly regulated iPSC-derived neurite length and morphology. Our results suggest that intrinsic differences in serotonergic neuron morphology and the resulting circuitry may contribute to SSRI resistance in MDD patients.

Differential L1 regulation in pluripotent stem cells of humans and apes.

Identifying cellular and molecular differences between human and non-human primates (NHPs) is essential to the basic understanding of the evolution and diversity of our own species. Until now, preserved tissues have been the main source for most comparative studies between humans, chimpanzees (Pan troglodytes) and bonobos (Pan paniscus). However, these tissue samples do not fairly represent the distinctive traits of live cell behaviour and are not amenable to genetic manipulation. We propose that induced pluripotent stem (iPS) cells could be a unique biological resource to determine relevant phenotypical differences between human and NHPs, and that those differences could have potential adaptation and speciation value. Here we describe the generation and initial characterization of iPS cells from chimpanzees and bonobos as new tools to explore factors that may have contributed to great ape evolution. Comparative gene expression analysis of human and NHP iPS cells revealed differences in the regulation of long interspersed element-1 (L1, also known as LINE-1) transposons. A force of change in mammalian evolution, L1 elements are retrotransposons that have remained active during primate evolution. Decreased levels of L1-restricting factors APOBEC3B (also known as A3B) and PIWIL2 (ref. 7) in NHP iPS cells correlated with increased L1 mobility and endogenous L1 messenger RNA levels. Moreover, results from the manipulation of A3B and PIWIL2 levels in iPS cells supported a causal inverse relationship between levels of these proteins and L1 retrotransposition. Finally, we found increased copy numbers of species-specific L1 elements in the genome of chimpanzees compared to humans, supporting the idea that increased L1 mobility in NHPs is not limited to iPS cells in culture and may have also occurred in the germ line or embryonic cells developmentally upstream to germline specification during primate evolution. We propose that differences in L1 mobility may have differentially shaped the genomes of humans and NHPs and could have continuing adaptive significance.

Horizontal Gene Transfer Building Prokaryote Genomes: Genes Related to Exchange Between Cell and Environment are Frequently Transferred.

Horizontal gene transfer (HGT) has a major impact on the evolution of prokaryotic genomes, as it allows genes evolved in different contexts to be combined in a single genome, greatly enhancing the ways evolving organisms can explore the gene content space and adapt to the environment. A systematic analysis of HGT in a large number of genomes is of key importance in understanding the impact of HGT in the evolution of prokaryotes. We developed a method for the detection of genes that potentially originated by HGT based on the comparison of BLAST scores between homologous genes to 16S rRNA-based phylogenetic distances between the involved organisms. The approach was applied to 697 prokaryote genomes and estimated that in average approximately 15% of the genes in prokaryote genomes originated by HGT, with a clear correlation between the proportion of predicted HGT genes and the size of the genome. The methodology was strongly supported by evolutionary relationships, as tested by the direct phylogenetic reconstruction of many of the HGT candidates. Studies performed with Escherichia coli W3110 genome clearly show that HGT proteins have fewer interactions when compared to those predicted as vertical inherited, an indication that the number of protein partners imposes limitations to horizontal transfer. A detailed functional classification confirms that genes related to protein translation are vertically inherited, whereas interestingly, transport and binding proteins are strongly enriched among HGT genes. Because these genes are related to the cell exchange with their environment, their transfer most likely contributed to successful adaptation throughout evolution.

Lifespan analysis of repeat expression reveals age-dependent upregulation of HERV-K in the neurotypical human brain.

DNA repetitive sequences (or repeats) comprise over 50% of the human genome and have a crucial regulatory role, specifically regulating transcription machinery. The human brain is the tissue with the highest detectable repeat expression and dysregulations on the repeat activity are related to several neurological and neurodegenerative disorders, as repeat-derived products can stimulate a pro-inflammatory response. Even so, it is unclear how repeat expression acts on the aging neurotypical brain. Here, we leverage a large postmortem transcriptome cohort spanning the human lifespan to assess global repeat expression in the neurotypical brain. We identified 21,696 differentially expressed repeats (DERs) that varied across seven age bins (Prenatal; 0-15; 16-29; 30-39; 40-49; 50-59; 60+) across the caudate nucleus (n=271), dorsolateral prefrontal cortex (n=304), and hippocampus (n=310). Interestingly, we found that long interspersed nuclear elements and long terminal repeats (LTRs) DERs were the most abundant repeat families when comparing infants to early adolescence (0-15) with older adults (60+). Of these differentially regulated LTRs, we identified 17 shared across all brain regions, including increased expression of HERV-K-int in older adult brains (60+). Co-expression analysis from each of the three brain regions also showed repeats from the HERV subfamily were intramodular hubs in its subnetworks. While we do not observe a strong global relationship between repeat expression and age, we identified HERV-K as a repeat signature associated with the aging neurotypical brain. Our study is the first global assessment of repeat expression in the neurotypical brain.

Analysis of gene expression in the postmortem brain of neurotypical Black Americans reveals contributions of genetic ancestry.

Ancestral differences in genomic variation affect the regulation of gene expression; however, most gene expression studies have been limited to European ancestry samples or adjusted to identify ancestry-independent associations. Here, we instead examined the impact of genetic ancestry on gene expression and DNA methylation in the postmortem brain tissue of admixed Black American neurotypical individuals to identify ancestry-dependent and ancestry-independent contributions. Ancestry-associated differentially expressed genes (DEGs), transcripts and gene networks, while notably not implicating neurons, are enriched for genes related to the immune response and vascular tissue and explain up to 26% of heritability for ischemic stroke, 27% of heritability for Parkinson disease and 30% of heritability for Alzheimer's disease. Ancestry-associated DEGs also show general enrichment for the heritability of diverse immune-related traits but depletion for psychiatric-related traits. We also compared Black and non-Hispanic white Americans, confirming most ancestry-associated DEGs. Our results delineate the extent to which genetic ancestry affects differences in gene expression in the human brain and the implications for brain illness risk.

Sex affects transcriptional associations with schizophrenia across the dorsolateral prefrontal cortex, hippocampus, and caudate nucleus.

Schizophrenia is a complex neuropsychiatric disorder with sexually dimorphic features, including differential symptomatology, drug responsiveness, and male incidence rate. Prior large-scale transcriptome analyses for sex differences in schizophrenia have focused on the prefrontal cortex. Analyzing BrainSeq Consortium data (caudate nucleus: n = 399, dorsolateral prefrontal cortex: n = 377, and hippocampus: n = 394), we identified 831 unique genes that exhibit sex differences across brain regions, enriched for immune-related pathways. We observed X-chromosome dosage reduction in the hippocampus of male individuals with schizophrenia. Our sex interaction model revealed 148 junctions dysregulated in a sex-specific manner in schizophrenia. Sex-specific schizophrenia analysis identified dozens of differentially expressed genes, notably enriched in immune-related pathways. Finally, our sex-interacting expression quantitative trait loci analysis revealed 704 unique genes, nine associated with schizophrenia risk. These findings emphasize the importance of sex-informed analysis of sexually dimorphic traits, inform personalized therapeutic strategies in schizophrenia, and highlight the need for increased female samples for schizophrenia analyses.

Human archetypal pluripotent stem cells differentiate into trophoblast stem cells via endogenous BMP5/7 induction without transitioning through naive state.

Primary human trophoblast stem cells (TSCs) and TSCs derived from human pluripotent stem cells (hPSCs) can potentially model placental processes in vitro. Yet, the pluripotent states and factors involved in the differentiation of hPSCs to TSCs remain poorly understood. In this study, we demonstrate that the primed pluripotent state can generate TSCs by activating pathways such as Epidermal Growth Factor (EGF) and Wingless-related integration site (WNT), and by suppressing tumor growth factor beta (TGFß), histone deacetylases (HDAC), and Rho-associated protein kinase (ROCK) signaling pathways, all without the addition of exogenous Bone morphogenetic protein 4 (BMP4)-a condition we refer to as the TS condition. We characterized this process using temporal single-cell RNA sequencing to compare TS conditions with differentiation protocols involving BMP4 activation alone or BMP4 activation in conjunction with WNT inhibition. The TS condition consistently produced a stable, proliferative cell type that closely mimics first-trimester placental cytotrophoblasts, marked by the activation of endogenous retroviral genes and the absence of amnion expression. This was observed across multiple cell lines, including various primed induced pluripotent stem cell (iPSC) and embryonic stem cell (ESC) lines. Primed-derived TSCs can proliferate for over 30 passages and further specify into multinucleated syncytiotrophoblasts and extravillous trophoblast cells. Our research establishes that the differentiation of primed hPSCs to TSC under TS conditions triggers the induction of TMSB4X, BMP5/7, GATA3, and TFAP2A without progressing through a naive state. These findings propose that the primed hPSC state is part of a continuum of potency with the capacity to differentiate into TSCs through multiple routes.

Transcriptome analysis of the acoelomate human parasite Schistosoma mansoni.

Schistosoma mansoni is the primary causative agent of schistosomiasis, which affects 200 million individuals in 74 countries. We generated 163,000 expressed-sequence tags (ESTs) from normalized cDNA libraries from six selected developmental stages of the parasite, resulting in 31,000 assembled sequences and 92% sampling of an estimated 14,000 gene complement. By analyzing automated Gene Ontology assignments, we provide a detailed view of important S. mansoni biological systems, including characterization of metazoa-specific and eukarya-conserved genes. Phylogenetic analysis suggests an early divergence from other metazoa. The data set provides insights into the molecular mechanisms of tissue organization, development, signaling, sexual dimorphism, host interactions and immune evasion and identifies novel proteins to be investigated as vaccine candidates and potential drug targets.

Genetic and environmental contributions to ancestry differences in gene expression in the human brain.

Ancestral differences in genomic variation are determining factors in gene regulation; however, most gene expression studies have been limited to European ancestry samples or adjusted for ancestry to identify ancestry-independent associations. We instead examined the impact of genetic ancestry on gene expression and DNA methylation (DNAm) in admixed African/Black American neurotypical individuals to untangle effects of genetic and environmental factors. Ancestry-associated differentially expressed genes (DEGs), transcripts, and gene networks, while notably not implicating neurons, are enriched for genes related to immune response and vascular tissue and explain up to 26% of heritability for ischemic stroke, 27% of heritability for Parkinson's disease, and 30% of heritability for Alzhemier's disease. Ancestry-associated DEGs also show general enrichment for heritability of diverse immune-related traits but depletion for psychiatric-related traits. The cell-type enrichments and direction of effects vary by brain region. These DEGs are less evolutionarily constrained and are largely explained by genetic variations; roughly 15% are predicted by DNAm variation implicating environmental exposures. We also compared Black and White Americans, confirming most of these ancestry-associated DEGs. Our results highlight how environment and genetic background affect genetic ancestry differences in gene expression in the human brain and affect risk for brain illness.

Recapitulation of Perturbed Striatal Gene Expression Dynamics of Donors' Brains With Ventral Forebrain Organoids Derived From the Same Individuals With Schizophrenia.

OBJECTIVE: Schizophrenia is a brain disorder that originates during neurodevelopment and has complex genetic and environmental etiologies. Despite decades of clinical evidence of altered striatal function in affected patients, studies examining its cellular and molecular mechanisms in humans are limited. To explore neurodevelopmental alterations in the striatum associated with schizophrenia, the authors established a method for the differentiation of induced pluripotent stem cells (iPSCs) into ventral forebrain organoids (VFOs). METHODS: VFOs were generated from postmortem dural fibroblast-derived iPSCs of four individuals with schizophrenia and four neurotypical control individuals for whom postmortem caudate genotypes and transcriptomic data were profiled in the BrainSeq neurogenomics consortium. Individuals were selected such that the two groups had nonoverlapping schizophrenia polygenic risk scores (PRSs). RESULTS: Single-cell RNA sequencing analyses of VFOs revealed differences in developmental trajectory between schizophrenia and control individuals in which inhibitory neuronal cells from the patients exhibited accelerated maturation. Furthermore, upregulated genes in inhibitory neurons in schizophrenia VFOs showed a significant overlap with upregulated genes in postmortem caudate tissue of individuals with schizophrenia compared with control individuals, including the donors of the iPSC cohort. CONCLUSIONS: The findings suggest that striatal neurons derived from high-PRS individuals with schizophrenia carry abnormalities that originated during early brain development and that the VFO model can recapitulate disease-relevant cell type-specific neurodevelopmental phenotypes in a dish.

Variation in TAF1 expression in female carrier induced pluripotent stem cells and human brain ontogeny has implications for adult neostriatum vulnerability in X-linked Dystonia Parkinsonism.

X-linked Dystonia-Parkinsonism (XDP) is an inherited, X-linked, adult-onset movement disorder characterized by degeneration in the neostriatum. No therapeutics alter disease progression. The mechanisms underlying regional differences in degeneration and adult onset are unknown. Developing therapeutics requires a deeper understanding of how XDP-relevant features vary in health and disease. XDP is possibly due, in part, to a partial loss of TAF1 function. A disease-specific SINE-VNTR-Alu (SVA) retrotransposon insertion occurs within intron 32 of TAF1, a subunit of TFIID involved in transcription initiation. While all XDP males are usually clinically affected, females are heterozygous carriers generally not manifesting the full syndrome. As a resource for disease modeling, we characterized eight iPSC lines from three XDP female carrier individuals for X chromosome inactivation status and identified clonal lines that express either the wild-type X or XDP haplotype. Furthermore, we characterized XDP-relevant transcript expression in neurotypical humans, and found that SVA-F expression decreases after 30 years of age in the brain and that TAF1 is decreased in most female samples. Uniquely in the caudate nucleus, TAF1 expression is not sexually dimorphic and decreased after adolescence. These findings indicate that regional-, age- and sex-specific mechanisms regulate TAF1, highlighting the importance of disease-relevant models and postmortem tissue. We propose that the decreased TAF1 expression in the adult caudate may synergize with the XDP-specific partial loss of TAF1 function in patients, thereby passing a minimum threshold of TAF1 function, and triggering degeneration in the neostriatum.Significance StatementXDP is an inherited, X-linked, adult-onset movement disorder characterized by degeneration in the neostriatum. No therapeutics alter disease progression. Developing therapeutics requires a deeper understanding of how XDP-relevant features vary in health and disease. XDP is possibly due to a partial loss of TAF1 function. While all XDP males are usually affected, females are heterozygous carriers generally not manifesting the full syndrome. As a resource for disease modeling, we characterized eight stem cell lines from XDP female carrier individuals. Furthermore, we found that, uniquely in the caudate nucleus, TAF1 expression decreases after adolescence in healthy humans. We hypothesize that the decrease of TAF1 after adolescence in human caudate, in general, may underlie the vulnerability of the adult neostriatum in XDP.

Analysis of the caudate nucleus transcriptome in individuals with schizophrenia highlights effects of antipsychotics and new risk genes.

Most studies of gene expression in the brains of individuals with schizophrenia have focused on cortical regions, but subcortical nuclei such as the striatum are prominently implicated in the disease, and current antipsychotic drugs target the striatum's dense dopaminergic innervation. Here, we performed a comprehensive analysis of the genetic and transcriptional landscape of schizophrenia in the postmortem caudate nucleus of the striatum of 443 individuals (245 neurotypical individuals, 154 individuals with schizophrenia and 44 individuals with bipolar disorder), 210 from African and 233 from European ancestries. Integrating expression quantitative trait loci analysis, Mendelian randomization with the latest schizophrenia genome-wide association study, transcriptome-wide association study and differential expression analysis, we identified many genes associated with schizophrenia risk, including potentially the dopamine D2 receptor short isoform. We found that antipsychotic medication has an extensive influence on caudate gene expression. We constructed caudate nucleus gene expression networks that highlight interactions involving schizophrenia risk. These analyses provide a resource for the study of schizophrenia and insights into risk mechanisms and potential therapeutic targets.

Age-dependent instability of mature neuronal fate in induced neurons from Alzheimer's patients.

Sporadic Alzheimer's disease (AD) exclusively affects elderly people. Using direct conversion of AD patient fibroblasts into induced neurons (iNs), we generated an age-equivalent neuronal model. AD patient-derived iNs exhibit strong neuronal transcriptome signatures characterized by downregulation of mature neuronal properties and upregulation of immature and progenitor-like signaling pathways. Mapping iNs to longitudinal neuronal differentiation trajectory data demonstrated that AD iNs reflect a hypo-mature neuronal identity characterized by markers of stress, cell cycle, and de-differentiation. Epigenetic landscape profiling revealed an underlying aberrant neuronal state that shares similarities with malignant transformation and age-dependent epigenetic erosion. To probe for the involvement of aging, we generated rejuvenated iPSC-derived neurons that showed no significant disease-related transcriptome signatures, a feature that is consistent with epigenetic clock and brain ontogenesis mapping, which indicate that fibroblast-derived iNs more closely reflect old adult brain stages. Our findings identify AD-related neuronal changes as age-dependent cellular programs that impair neuronal identity.

Generation of four postmortem dura-derived iPS cell lines from four control individuals with genotypic and brain-region-specific transcriptomic data available through the BrainSEQ consortium.

In this study, we established induced pluripotent stem (iPS) cell lines from postmortem dura-derived fibroblasts of four control individuals with low polygenic risk score for psychiatric disorders including schizophrenia and bipolar disorder. The fibroblasts were reprogrammed into iPS cells using episomal vectors carrying OCT3/4, SOX2, KLF4, L-Myc, LIN28 and shRNA-p53. All iPS cell lines showed the same genotype with parental postmortem brain tissues, expressed pluripotency markers, and exhibited the differentiation potency into three embryonic germ layers.

Pathological priming causes developmental gene network heterochronicity in autistic subject-derived neurons.

Autism spectrum disorder (ASD) is thought to emerge during early cortical development. However, the exact developmental stages and associated molecular networks that prime disease propensity are elusive. To profile early neurodevelopmental alterations in ASD with macrocephaly, we monitored subject-derived induced pluripotent stem cells (iPSCs) throughout the recapitulation of cortical development. Our analysis revealed ASD-associated changes in the maturational sequence of early neuron development, involving temporal dysregulation of specific gene networks and morphological growth acceleration. The observed changes tracked back to a pathologically primed stage in neural stem cells (NSCs), reflected by altered chromatin accessibility. Concerted over-representation of network factors in control NSCs was sufficient to trigger ASD-like features, and circumventing the NSC stage by direct conversion of ASD iPSCs into induced neurons abolished ASD-associated phenotypes. Our findings identify heterochronic dynamics of a gene network that, while established earlier in development, contributes to subsequent neurodevelopmental aberrations in ASD.

Laterally transferred genomic islands in Xanthomonadales related to pathogenicity and primary metabolism.

Lateral gene transfer (LGT) is considered as one of the drivers in bacterial genome evolution, usually associated with increased fitness and/or changes in behavior, especially if one considers pathogenic vs. non-pathogenic bacterial groups. The genomes of two phytopathogens, Xanthomonas campestris pv. campestris and Xanthomonas axonopodis pv. citri, were previously inspected for genome islands originating from LGT events, and, in this work, potentially early and late LGT events were identified according to their altered nucleotide composition. The biological role of the islands was also assessed, and pathogenicity, virulence and secondary metabolism pathways were functions highly represented, especially in islands that were found to be recently transferred. However, old islands are composed of a high proportion of genes related to cell primary metabolic functions. These old islands, normally undetected by traditional atypical composition analysis, but confirmed as product of LGT by atypical phylogenetic reconstruction, reveal the role of LGT events by replacing core metabolic genes normally inherited by vertical processes.