Reciprocal conversion between annual and polycarpic perennial flowering behavior in the Brassicaceae.

The development of perennial crops holds great promise for sustainable agriculture and food security. However, the evolution of the transition between perenniality and annuality is poorly understood. Here, using two Brassicaceae species, Crucihimalaya himalaica and Erysimum nevadense, as polycarpic perennial models, we reveal that the transition from polycarpic perennial to biennial and annual flowering behavior is a continuum determined by the dosage of three closely related MADS-box genes. Diversification of the expression patterns, functional strengths, and combinations of these genes endows species with the potential to adopt various life-history strategies. Remarkably, we find that a single gene among these three is sufficient to convert winter-annual or annual Brassicaceae plants into polycarpic perennial flowering plants. Our work delineates a genetic basis for the evolution of diverse life-history strategies in plants and lays the groundwork for the generation of diverse perennial Brassicaceae crops in the future.

Aneuploidy Causes Non-genetic Individuality.

Phenotypic variability is a hallmark of diseases involving chromosome gains and losses, such as Down syndrome and cancer. Allelic variances have been thought to be the sole cause of this heterogeneity. Here, we systematically examine the consequences of gaining and losing single or multiple chromosomes to show that the aneuploid state causes non-genetic phenotypic variability. Yeast cell populations harboring the same defined aneuploidy exhibit heterogeneity in cell-cycle progression and response to environmental perturbations. Variability increases with degree of aneuploidy and is partly due to gene copy number imbalances, suggesting that subtle changes in gene expression impact the robustness of biological networks and cause alternate behaviors when they occur across many genes. As inbred trisomic mice also exhibit variable phenotypes, we further propose that non-genetic individuality is a universal characteristic of the aneuploid state that may contribute to variability in presentation and treatment responses of diseases caused by aneuploidy.

Bypassing Negative Epistasis on Yield in Tomato Imposed by a Domestication Gene.

Selection for inflorescence architecture with improved flower production and yield is common to many domesticated crops. However, tomato inflorescences resemble wild ancestors, and breeders avoided excessive branching because of low fertility. We found branched variants carry mutations in two related transcription factors that were selected independently. One founder mutation enlarged the leaf-like organs on fruits and was selected as fruit size increased during domestication. The other mutation eliminated the flower abscission zone, providing "jointless" fruit stems that reduced fruit dropping and facilitated mechanical harvesting. Stacking both beneficial traits caused undesirable branching and sterility due to epistasis, which breeders overcame with suppressors. However, this suppression restricted the opportunity for productivity gains from weak branching. Exploiting natural and engineered alleles for multiple family members, we achieved a continuum of inflorescence complexity that allowed breeding of higher-yielding hybrids. Characterizing and neutralizing similar cases of negative epistasis could improve productivity in many agricultural organisms. VIDEO ABSTRACT.

New Horizons for Dissecting Epistasis in Crop Quantitative Trait Variation.

Uncovering the genes, variants, and interactions underlying crop diversity is a frontier in plant genetics. Phenotypic variation often does not reflect the cumulative effect of individual gene mutations. This deviation is due to epistasis, in which interactions between alleles are often unpredictable and quantitative in effect. Recent advances in genomics and genome-editing technologies are elevating the study of epistasis in crops. Using the traits and developmental pathways that were major targets in domestication and breeding, we highlight how epistasis is central in guiding the behavior of the genetic variation that shapes quantitative trait variation. We outline new strategies that illuminate how quantitative epistasis from modified gene dosage defines background dependencies. Advancing our understanding of epistasis in crops can reveal new principles and approaches to engineering targeted improvements in agriculture.

Developmental trajectories of GABAergic cortical interneurons are sequentially modulated by dynamic FoxG1 expression levels.

GABAergic inhibitory interneurons, originating from the embryonic ventral forebrain territories, traverse a convoluted migratory path to reach the neocortex. These interneuron precursors undergo sequential phases of tangential and radial migration before settling into specific laminae during differentiation. Here, we show that the developmental trajectory of FoxG1 expression is dynamically controlled in these interneuron precursors at critical junctures of migration. By utilizing mouse genetic strategies, we elucidate the pivotal role of precise changes in FoxG1 expression levels during interneuron specification and migration. Our findings underscore the gene dosage-dependent function of FoxG1, aligning with clinical observations of FOXG1 haploinsufficiency and duplication in syndromic forms of autism spectrum disorders. In conclusion, our results reveal the finely tuned developmental clock governing cortical interneuron development, driven by temporal dynamics and the dose-dependent actions of FoxG1.

X-chromosome inactivation: a historic topic that's still hot.

The last edition of the X-chromosome inactivation (XCI) meeting was held as an EMBO workshop in Berlin on 19-22 June 2023. The conference took place at the Harnack-haus in the Dahlem district, birthplace of the first modern research campus, where notable scientists such as Lise Meitner, Hans Krebs and, briefly, Albert Einstein conducted their research. This special edition, also accessible online, was organized by Rafael Galupa (Centre for Integrative Biology of Toulouse, France), Joost Gribnau (Erasmus MC Rotterdam, The Netherlands), Claire Rougeulle (Université Paris Cité/CNRS, Epigenetics and Cell Fate Center, Paris, France), Edda Schulz (Max Planck Institute for Molecular Genetics, Berlin, Germany) and James Turner (The Francis Crick Institute, London, UK). Originally scheduled for 2021, to commemorate the 60th anniversary of Mary Lyon's hypothesis on X-chromosome inactivation in mammals and the 30th anniversary of XIST/Xist discovery, the meeting had to be postponed because of the COVID-19 pandemic. Seven years after the latest XCI meeting in London, the enthusiasm and expectations of the community were at their highest, bringing together over 160 scientists from around the world to share and discuss their research. Eighty posters and more than 40 talks were presented at this event, in a collegial and collaborative atmosphere. A historical session and several breakout discussions were also organized, as well as the now traditional boat trip, all thanks to great organization. Here, we debrief readers on this fantastic conference.

A new Down syndrome rat model races forward.

Animal models of Down syndrome (DS) provide an essential resource for understanding genetic, cellular, and molecular contributions to traits associated with trisomy 21 (Ts21). Recent genetic enhancements in the development of DS models, including the new TcHSA21rat model (Kazuki et al.), have potential to transform our understanding of and potential therapies for Ts21.

TREM2 variants that cause early dementia and increase Alzheimer's disease risk affect gene splicing.

Loss-of-function variants in the triggering receptor expressed on myeloid cells 2 (TREM2) are responsible for a spectrum of neurodegenerative disorders. In the homozygous state, they cause severe pathologies with early onset dementia, such as Nasu-Hakola disease and behavioural variants of frontotemporal dementia (FTD), whereas heterozygous variants increase the risk of late-onset Alzheimer's disease (AD) and FTD. For over half of TREM2 variants found in families with recessive early onset dementia, the defect occurs at the transcript level via premature termination codons or aberrant splicing. The remaining variants are missense alterations thought to affect the protein; however, the underlying pathogenic mechanism is less clear. In this work, we tested whether these disease-associated TREM2 variants contribute to the pathology via altered splicing. Variants scored by SpliceAI algorithm were tested by a full-size TREM2 splicing reporter assay in different cell lines. The effect of variants was quantified by qRT-/RT-PCR and western blots. Nanostring nCounter was used to measure TREM2 RNA in the brains of NHD patients who carried spliceogenic variants. Exon skipping events were analysed from brain RNA-Seq datasets available through the Accelerating Medicines Partnership for Alzheimer's Disease Consortium. We found that for some Nasu-Hakola disease and early onset FTD-causing variants, splicing defects were the primary cause (D134G) or likely contributor to pathogenicity (V126G and K186N). Similar but milder effects on splicing of exons 2 and 3 were demonstrated for A130V, L133L and R136W enriched in patients with dementia. Moreover, the two most frequent missense variants associated with AD/FTD risk in European and African ancestries (R62H, 1% in Caucasians and T96K, 12% in Africans) had splicing defects via excessive skipping of exon 2 and overproduction of a potentially antagonistic TREM2 protein isoform. The effect of R62H on exon 2 skipping was confirmed in three independent brain RNA-Seq datasets. Our findings revealed an unanticipated complexity of pathogenic variation in TREM2, in which effects on post-transcriptional gene regulation and protein function often coexist. This necessitates the inclusion of computational and experimental analyses of splicing and mRNA processing for a better understanding of genetic variation in disease.

New Perspectives on Genomic Imprinting, an Essential and Multifaceted Mode of Epigenetic Control in the Developing and Adult Brain.

Mammalian evolution entailed multiple innovations in gene regulation, including the emergence of genomic imprinting, an epigenetic regulation leading to the preferential expression of a gene from its maternal or paternal allele. Genomic imprinting is highly prevalent in the brain, yet, until recently, its central roles in neural processes have not been fully appreciated. Here, we provide a comprehensive survey of adult and developmental brain functions influenced by imprinted genes, from neural development and wiring to synaptic function and plasticity, energy balance, social behaviors, emotions, and cognition. We further review the widespread identification of parental biases alongside monoallelic expression in brain tissues, discuss their potential roles in dosage regulation of key neural pathways, and suggest possible mechanisms underlying the dynamic regulation of imprinting in the brain. This review should help provide a better understanding of the significance of genomic imprinting in the normal and pathological brain of mammals including humans.

Dlk1 dosage regulates hippocampal neurogenesis and cognition.

Neurogenesis in the adult brain gives rise to functional neurons, which integrate into neuronal circuits and modulate neural plasticity. Sustained neurogenesis throughout life occurs in the subgranular zone (SGZ) of the dentate gyrus in the hippocampus and is hypothesized to be involved in behavioral/cognitive processes such as memory and in diseases. Genomic imprinting is of critical importance to brain development and normal behavior, and exemplifies how epigenetic states regulate genome function and gene dosage. While most genes are expressed from both alleles, imprinted genes are usually expressed from either the maternally or the paternally inherited chromosome. Here, we show that in contrast to its canonical imprinting in nonneurogenic regions, Delta-like homolog 1 (Dlk1) is expressed biallelically in the SGZ, and both parental alleles are required for stem cell behavior and normal adult neurogenesis in the hippocampus. To evaluate the effects of maternally, paternally, and biallelically inherited mutations within the Dlk1 gene in specific behavioral domains, we subjected Dlk1-mutant mice to a battery of tests that dissociate and evaluate the effects of Dlk1 dosage on spatial learning ability and on anxiety traits. Importantly, reduction in Dlk1 levels triggers specific cognitive abnormalities that affect aspects of discriminating differences in environmental stimuli, emphasizing the importance of selective absence of imprinting in this neurogenic niche.

TCF3 haploinsufficiency defined by immune, clinical, gene-dosage, and murine studies.

BACKGROUND: TCF3 is a transcription factor contributing to early lymphocyte differentiation. Germline monoallelic dominant negative and biallelic loss-of-function (LOF) null TCF3 mutations cause a fully penetrant severe immunodeficiency. We identified 8 individuals from 7 unrelated families with monoallelic LOF TCF3 variants presenting with immunodeficiency with incomplete clinical penetrance. OBJECTIVE: We sought to define TCF3 haploinsufficiency (HI) biology and its association with immunodeficiency. METHODS: Patient clinical data and blood samples were analyzed. Flow cytometry, Western blot analysis, plasmablast differentiation, immunoglobulin secretion, and transcriptional activity studies were conducted on individuals carrying TCF3 variants. Mice with a heterozygous Tcf3 deletion were analyzed for lymphocyte development and phenotyping. RESULTS: Individuals carrying monoallelic LOF TCF3 variants showed B-cell defects (eg, reduced total, class-switched memory, and/or plasmablasts) and reduced serum immunoglobulin levels; most but not all presented with recurrent but nonsevere infections. These TCF3 LOF variants were either not transcribed or translated, resulting in reduced wild-type TCF3 protein expression, strongly suggesting HI pathophysiology for the disease. Targeted RNA sequencing analysis of T-cell blasts from TCF3-null, dominant negative, or HI individuals clustered away from healthy donors, implying that 2 WT copies of TCF3 are needed to sustain a tightly regulated TCF3 gene-dosage effect. Murine TCF3 HI resulted in a reduction of circulating B cells but overall normal humoral immune responses. CONCLUSION: Monoallelic LOF TCF3 mutations cause a gene-dosage-dependent reduction in wild-type protein expression, B-cell defects, and a dysregulated transcriptome, resulting in immunodeficiency. Tcf3+/- mice partially recapitulate the human phenotype, underscoring the differences between TCF3 in humans and mice.

The Consequences of Abnormal Gene Dosage: Lessons from Chromosome 18.

Accurate interpretation of genomic copy number variation (CNV) remains a challenge and has important consequences for both congenital and late-onset disease. Hemizygosity dosage characterization of the genes on chromosome 18 reveals a spectrum of outcomes ranging from no clinical effect, to risk factors for disease, to both low- and high-penetrance disease. These data are important for accurate and predictive clinical management. Additionally, the potential mechanisms of reduced penetrance due to dosage compensation are discussed as a key to understanding avenues for potential treatment. This review describes the chromosome 18 findings, and discusses the molecular mechanisms that allow haploinsufficiency, reduced penetrance, and dosage compensation.

Iteron control of oriV function in IncP-1 plasmid RK2.

Replication control of many plasmids is mediated by the balance between the positive and negative effects of Rep protein binding repeated sequences (iterons) associated with the replication origin, oriV. Negative control is thought to be mediated by dimeric Rep protein linking iterons in a process termed "handcuffing". The well-studied oriV region of RK2 contains 9 iterons arranged as a singleton (iteron 1), a group of 3 (iterons 2-4) and a group of 5 (iterons 5-9), but only iterons 5 to 9 are essential for replication. An additional iteron (iteron 10), oriented in the opposite direction, is also involved and reduces copy-number nearly two-fold. Since iterons 1 and 10 share an identical upstream hexamer (5' TTTCAT 3') it has been hypothesised that they form a TrfA-mediated loop facilitated by their inverted orientation. Here we report that contrary to the hypothesis, flipping one or other so they are in direct orientation results in marginally lower rather than higher copy-number. In addition, following mutagenesis of the hexamer upstream of iteron 10, we report that the Logo for the hexamer "upstream" of the regulatory iterons (1 to 4 and 10) differs from that of the essential iterons, suggesting functional differences in their interaction with TrfA.

"Liu-Liang-Chung" syndrome with multiple congenital anomalies and the distinctive craniofacial features caused by dominant ZEB2 gene gain mutation.

BACKGROUND: Contiguous gene gain syndrome including entire ZEB2 may be a novel syndrome. In the past, there were no easily distinct and recognizable features as a guide for precise clinical and genetic diagnosis of the syndrome. CASE PRESENTATION: We report a novel case with the syndrome with a novel de novo 22.16 Mb duplication at 2q21.2-q24.1. The syndrome is characterized by multiple anomalies including the same typical craniofacial phenotype that is entirely different from Mowat-Wilson syndrome (MWS), and other quite similar features of MWS consisting of development delay, congenital heart disease, abdominal abnormalities, urogenital abnormalities, behavioral problems and so on, in which the distinctive craniofacial features can be more easily recognized. CONCLUSIONS: Contiguous gene gain syndrome including entire ZEB2 characterized with similar multiple congenital anomalies of MWS and the distinctive craniofacial features is mainly caused by large 2q22 repeats including ZEB2 leading to dominant singe ZEB2 gene gain mutation, which is recommended to be named "Liu-Liang-Chung" syndrome. We diagnose this novel syndrome to distinguish it from MWS. Some variable additional features in the syndrome including remarkable growth and development retardation and protruding ears were recognized for the first time.

Alpha Globin Gene Copy Number Is Associated with Prevalent Chronic Kidney Disease and Incident End-Stage Kidney Disease among Black Americans.

BACKGROUND: α-Globin is expressed in endothelial cells of resistance arteries, where it limits endothelial nitric oxide signaling and enhances α-adrenergic-mediated vasoconstriction. α-Globin gene (HBA) copy number is variable in people of African descent and other populations worldwide. Given the protective effect of nitric oxide in the kidney, we hypothesized that HBA copy number would be associated with kidney disease risk. METHODS: Community-dwelling Black Americans aged ≥45 years old were enrolled in a national longitudinal cohort from 2003 through 2007. HBA copy number was measured using droplet digital PCR. The prevalence ratio (PR) of CKD and the relative risk (RR) of incident reduced eGFR were calculated using modified Poisson multivariable regression. The hazard ratio (HR) of incident ESKD was calculated using Cox proportional hazards multivariable regression. RESULTS: Among 9908 participants, HBA copy number varied from 2 to 6. In analyses adjusted for demographic, clinical, and genetic risk factors, a one-copy increase in HBA was associated with 14% greater prevalence of CKD (PR, 1.14; 95% CI, 1.07 to 1.21; P<0.0001). While HBA copy number was not associated with incident reduced eGFR (RR, 1.06; 95% CI, 0.94 to 1.19; P=0.38), the hazard of incident ESKD was 32% higher for each additional copy of HBA (HR, 1.32; 95% CI, 1.09 to 1.61; P=0.005). CONCLUSIONS: Increasing HBA copy number was associated with a greater prevalence of CKD and incidence of ESKD in a national longitudinal cohort of Black Americans.

Steroidogenic Factor 1, a Goldilocks Transcription Factor from Adrenocortical Organogenesis to Malignancy.

Steroidogenic factor-1 (SF-1, also termed Ad4BP; NR5A1 in the official nomenclature) is a nuclear receptor transcription factor that plays a crucial role in the regulation of adrenal and gonadal development, function and maintenance. In addition to its classical role in regulating the expression of P450 steroid hydroxylases and other steroidogenic genes, involvement in other key processes such as cell survival/proliferation and cytoskeleton dynamics have also been highlighted for SF-1. SF-1 has a restricted pattern of expression, being expressed along the hypothalamic-pituitary axis and in steroidogenic organs since the time of their establishment. Reduced SF-1 expression affects proper gonadal and adrenal organogenesis and function. On the other hand, SF-1 overexpression is found in adrenocortical carcinoma and represents a prognostic marker for patients' survival. This review is focused on the current knowledge about SF-1 and the crucial importance of its dosage for adrenal gland development and function, from its involvement in adrenal cortex formation to tumorigenesis. Overall, data converge towards SF-1 being a key player in the complex network of transcriptional regulation within the adrenal gland in a dosage-dependent manner.

Multiple optimizations of recombinant plasmid for improving expression of Hepatitis B core antigen in Escherichia coli.

Hepatitis B core antigen (HBcAg) can self-assemble into virus-like particles (VLPs) when expressed in Escherichia coli. We optimized the different of the expression plasmid pBV220, including the ribosome bind site (RBS), spacer region, promoter and replication origin (ori), as well as the hbc gene dosage, to enhance HBcAg transcription and translation in E. coli. The optimized construct with a customized RBS6, 6 nt spacer, T7 promoter and pUCori significantly increased the levels of HBc36GFP fusion protein to 3.4-folds compared to the control. Thereafter, we substituted hbc36gfp gene with different copies of the hbc gene and tested the effects of gene dosage on HBcAg expression. The HBcAg-VLPs yield obtained using an engineered strain with three copies of hbc was 842.1 ± 46.8 µg/mL, which was 2.2-folds higher compared to that in the control strain. Thus, our study provides a simple and effective strategy for improving HBcAg expression in E. coli. Since the HBcAg-VLPs are promising carriers for presenting foreign antigen epitopes, an in vitro expression system that can generate high levels of HBcAg-VLPs can serve as a promising tool for developing novel HBV vaccines and drugs.

Why haploinsufficiency persists.

Haploinsufficiency describes the decrease in organismal fitness observed when a single copy of a gene is deleted in diploids. We investigated the origin of haploinsufficiency by creating a comprehensive dosage sensitivity data set for genes under their native promoters. We demonstrate that the expression of haploinsufficient genes is limited by the toxicity of their overexpression. We further show that the fitness penalty associated with excess gene copy number is not the only determinant of haploinsufficiency. Haploinsufficient genes represent a unique subset of genes sensitive to copy number increases, as they are also limiting for important cellular processes when present in one copy instead of two. The selective pressure to decrease gene expression due to the toxicity of overexpression, combined with the pressure to increase expression due to their fitness-limiting nature, has made haploinsufficient genes extremely sensitive to changes in gene expression. As a consequence, haploinsufficient genes are dosage stabilized, showing much more narrow ranges in cell-to-cell variability of expression compared with other genes in the genome. We propose a dosage-stabilizing hypothesis of haploinsufficiency to explain its persistence over evolutionary time.

A comprehensive genomic scan reveals gene dosage balance impacts on quantitative traits in Populus trees.

Gene dosage variation and the associated changes in gene expression influence a wide variety of traits, ranging from cancer in humans to yield in plants. It is also expected to affect important traits of ecological and agronomic importance in forest trees, but this variation has not been systematically characterized or exploited. Here we performed a comprehensive scan of the Populus genome for dosage-sensitive loci affecting quantitative trait variation for spring and fall phenology and biomass production. The study population was a large collection of clonally propagated F1 hybrid lines of Populus that saturate the genome 10-fold with deletions and insertions (indels) of known sizes and positions. As a group, the phenotypic means of the indel lines consistently differed from control nonindel lines, with an overall negative effect of both insertions and deletions on all biomass-related traits but more diverse effects and an overall wider phenotypic distribution of the indel lines for the phenology-related traits. We also investigated the correlation between gene dosage at specific chromosomal locations and phenotype, to identify dosage quantitative trait loci (dQTL). Such dQTL were detected for most phenotypes examined, but stronger effect dQTL were identified for the phenology-related traits than for the biomass traits. Our genome-wide screen for dosage sensitivity in a higher eukaryote demonstrates the importance of global genomic balance and the impact of dosage on life history traits.

Engineering precursor and co-factor supply to enhance D-pantothenic acid production in Bacillus megaterium.

High-yielding chemical and chemo-enzymatic methods of D-pantothenic acid (DPA) synthesis are limited by using poisonous chemicals and DL-pantolactone racemic mixture formation. Alternatively, the safe microbial fermentative route of DPA production was found promising but suffered from low productivity and precursor supplementation. In this study, Bacillus megaterium was metabolically engineered to produce DPA without precursor supplementation. In order to provide a higher supply of precursor D-pantoic acid, key genes involved in its synthesis are overexpressed, resulting strain was produced 0.53 ± 0.08 g/L DPA was attained in shake flasks. Cofactor CH2-THF was found to be vital for DPA biosynthesis and was regenerated through the serine-glycine degradation pathway. Enhanced supply of another precursor, ß-alanine was achieved by codon optimization and dosing of the limiting L-asparate-1-decarboxylase (ADC). Co-expression of Pantoate-ß-alanine ligase, ADC, phosphoenolpyruvate carboxylase, aspartate aminotransferase and aspartate ammonia-lyase enhanced DPA concentration to 2.56 ± 0.05 g/L at shake flasks level. Fed-batch fermentation in a bioreactor with and without the supplementation of ß-alanine increased DPA concentration to 19.52 ± 0.26 and 4.78 ± 0.53 g/L, respectively. This present study successfully demonstrated a rational approach combining precursor supply engineering with cofactor regeneration for the enhancement of DPA titer in recombinant B. megaterium.