Development Of Phenotype Research Articles

Neurological and psychiatric characterization of rapid-onset dystonia-parkinsonism over time.

The onset of symptoms in Rapid-onset dystonia-parkinsonism (RDP) is typically over days to weeks and is often triggered by stressors like fever or childbirth. Limited information is available on how the motor and nonmotor symptoms evolve over the course of the disease. Our longitudinal study analyzed data from a cohort of RDP patients, documenting their symptoms across multiple visits. We characterized the phenotypic evolution of 14 individuals positive for ATP1A3 mutations (7 females, 7 males; mean examination age=37 years, mean age of onset=20 years). We focused on neurologic, cognitive, and neuropsychological data collected during in-person visits (mean interval between testing=5½ years). Initially, all participants exhibited bulbar symptoms. Headaches were noted in 50%, seizures in 31%, and tremors in 36%. At follow-up, 29% of those initially without headaches developed them, 22% without prior seizures experienced them, and 56% previously without tremors developed them. No improvements were seen in those with headaches; however, seizures and tremors improved in 25% and 80% of cases, respectively. For Burke-Fahn-Marsden Dystonia Rating Scale, Unified Parkinson's Disease Rating Scale, and International Cooperative Ataxia Rating Scale scores, improvement consisted of the reduction of the symptom. Cognitive functions improved from mildly impaired to low-average, and psychiatric evaluations indicated mild anxiety levels, slight increases in obsessive-compulsive behaviors, and decreased depression scores over time. This longitudinal analysis highlights the complex evolution of RDP, demonstrating significant variability in motor function and other symptoms such as headaches, seizures, and tremors.

Homospermidine synthase evolution and the origin(s) of pyrrolizidine alkaloids in Apocynaceae.

Enzymes that are encoded by paralogous genes and produce identical specialized metabolites in distantly related plant lineages are strong evidence of parallel phenotypic evolution. Inference of phenotypic homology for metabolites produced by orthologous genes is less straightforward, since orthologs may be recruited in parallel into novel pathways. In prior research on pyrrolizidine alkaloids (PAs), specialized metabolites of Apocynaceae, the evolution of homospermidine synthase (HSS), an enzyme of PA biosynthesis, was reconstructed and a single origin of PAs inferred because HSS enzymes of all known PA-producing Apocynaceae species are orthologous and descended from an ancestral enzyme with the motif (VXXXD) of an optimized HSS. We increased sampling, tested the effect of amino acid motif on HSS function, revisited motif evolution, and tested for selection to infer evolution of HSS function and its correlation with phenotype. Some evidence supports a single origin of PAs: an IXXXD HSS-like gene, similar in function to VXXXD HSS, evolved in the shared ancestor of all PA-producing species; loss of HSS function occurred multiple times via pseudogenization and perhaps via evolution of an IXXXN motif. Other evidence indicates multiple origins: the VXXXD motif, highly correlated with the PA phenotype, evolved two or four times independently; the ancestral IXXXD gene was not under positive selection, while some VXXXD genes were; and substitutions at sites experiencing positive selection occurred on multiple branches in the HSS-like gene tree. The complexity of the genotype-function-phenotype map confounds the inference of PA homology from HSS-like gene evolution in Apocynaceae.

Optogenetic control of transgene expression in Marchantia polymorpha

AbstractPremiseThe model liverwort Marchantia polymorpha is an emerging testbed species for plant metabolic engineering but lacks well‐characterized inducible promoters, which are necessary to minimize biochemical and physiological disruption when over‐accumulating target products. Here, we demonstrate the functionality of the light‐inducible plant‐usable light‐switch elements (PULSE) optogenetic system in Marchantia and exemplify its use through the light‐inducible overproduction of the bioplastic poly‐3‐hydroxybutyrate (PHB).MethodsThe PULSE system was used to drive expression of luciferase as a reporter and characterize its induction in transgenic M. polymorpha. Additionally, PULSE was used to drive expression of the PHB biosynthetic pathway; the accumulation of PHB under light‐inducible control was compared to constitutive overexpression.ResultsPULSE was fully functional and minimally leaky in M. polymorpha. The presence of the PULSE construct, even in the absence of induction, resulted in a developmental phenotype. Constitutive and inducible expression resulted in similar PHB accumulation levels.DiscussionPHB biosynthesis in plants is known to adversely affect plant health, but placing its production under optogenetic control alleviated negative effects on biomass accumulation in some instances. The work presented here represents a significant expansion of the toolbox for the metabolic engineering of M. polymorpha.

Functional innovation through new genes as a general evolutionary process.

In the past decade, our understanding of how new genes originate in diverse organisms has advanced substantially, and more than a dozen molecular mechanisms for generating initial gene structures were identified, in addition to gene duplication. These new genes have been found to integrate into and modify pre-existing gene networks primarily through mutation and selection, revealing new patterns and rules with stable origination rates across various organisms. This progress has challenged the prevailing belief that new proteins evolve from pre-existing genes, as new genes may arise de novo from noncoding DNA sequences in many organisms, with high rates observed in flowering plants. New genes have important roles in phenotypic and functional evolution across diverse biological processes and structures, with detectable fitness effects of sexual conflict genes that can shape species divergence. Such knowledge of new genes can be of translational value in agriculture and medicine.

Prognostic models of immune-related cell death and stress unveil mechanisms driving macrophage phenotypic evolution in colorectal cancer

BackgroundTumor microenvironment (TME), particularly immune cell infiltration, programmed cell death (PCD) and stress, has increasingly become a focal point in colorectal cancer (CRC) treatment. Uncovering the intricate crosstalk between these factors can enhance our understanding of CRC, guide therapeutic strategies, and improve patient prognosis.MethodsWe constructed an immune-related cell death and stress (ICDS) prognostic model utilizing machine learning methodologies. Furthermore, we performed enrichment analyses and deconvolution algorithms to elucidate the complex interactions between immune cell infiltration and the processes of PCD and stress within a substantial array of transcriptomic data from The Cancer Genome Atlas (TCGA) and Gene Expression Omnibus data base (GEO) related to CRC. Single-cell sequencing and biochemical experiments were used to validate the interaction between the model genes and programmed cell death in tumor cells.ResultsThe ICDS prognostic model exhibited robust predictive performance in seven independent cohorts, revealing an inverse correlation between model scores and patient prognosis. Meanwhile, the ICDS index was positively correlated with clinical stage. Model analysis indicated that patient subgroups with low ICDS index exhibited heightened immune activation features and elevated activity in PCD and stress pathways. Single-cell analysis further revealed that macrophages were the central drivers of immune characteristics underlying prognostic differences within the ICDS prognostic model. Pseudotime analysis and cellular experiments indicated that the model gene GAL3ST4 promotes the transition of macrophages toward an M2 pro-tumor phenotype. Furthermore, cell communication analysis and experimental validation revealed that the cuproptosis in tumor cells suppress GAL3ST4 expression, thereby inhibiting M2-like macrophage polarization.ConclusionIn summary, we constructed the ICDS prognostic model and uncovered the mechanism by which tumor cells downregulate GAL3ST4 expression via cuproptosis to inhibit M2-like macrophage polarization, providing new targets and biomarkers for CRC treatment and prognosis evaluation.

Genome Wide Identification of Structure Variations in Five Italian Turkey Populations

Structural variants (SVs) are one of the main sources of genetic variants and have a significant impact on phenotype evolution, disease susceptibility, and environmental adaptations. We used 73 whole genome sequencing (12x) to apply a mapping approach to identify SVs in five turkey populations. A notable degree of genetic isolation was observed between the Basilicata and Apulian populations, as indicated by principal component analysis and admixture results. A total of 11,733 SVs were detected, including 6712 deletions, 2671 duplications, 1430 inversions, and 920 translocations. The Variant Effect Predictor (VEP) analysis predicted various consequences of filtered SVs as follows: intron variants (35.8%), intergenic variants (9.6%), coding sequence variants (8.3%), downstream gene variants (7.5%), and transcript ablations (7.3%). Our functional annotation of genes overlapping with SVs was mainly enriched in recognized pathways governing positive regulation of nucleoplasm, protein binding, mitochondrion, negative regulation of cell population proliferation, identical protein binding, and calcium signaling. We produced a comprehensive SV catalog utilizing unique whole-genome turkey data. This SV catalog not only increases our understanding of genetic diversity in turkeys but also enhances our knowledge of the role of SVs in their phenotypic traits.

The widely used Ucp1-Cre transgene elicits complex developmental and metabolic phenotypes

Bacterial artificial chromosome transgenic models, including most Cre-recombinases, enable potent interrogation of gene function in vivo but require rigorous validation as limitations emerge. Due to its high relevance to metabolic studies, we perform comprehensive analysis of the Ucp1-CreEvdr line which is widely used for brown fat research. Hemizygotes exhibit major brown and white fat transcriptomic dysregulation, indicating potential altered tissue function. Ucp1-CreEvdr homozygotes also show high mortality, tissue specific growth defects, and craniofacial abnormalities. Mapping the transgene insertion site reveals insertion in chromosome 1 accompanied by large genomic alterations disrupting several genes expressed in a range of tissues. Notably, Ucp1-CreEvdr transgene retains an extra Ucp1 gene copy that may be highly expressed under high thermogenic burden. Our multi-faceted analysis highlights a complex phenotype arising from the presence of the Ucp1-CreEvdr transgene independently of intended genetic manipulations. Overall, comprehensive validation of transgenic mice is imperative to maximize discovery while mitigating unexpected, off-target effects.

Comparative transcriptomics in serial organs uncovers early and pan-organ developmental changes associated with organ-specific morphological adaptation

Mice have evolved a new dental plan with two additional cusps on the upper molar, while hamsters were retaining the ancestral plan. By comparing the dynamics of molar development with transcriptome time series, we found at least three early changes in mouse upper molar development. Together, they redirect spatio-temporal dynamics to ultimately form two additional cusps. The mouse lower molar has undergone much more limited phenotypic evolution. Nevertheless, its developmental trajectory evolved as much as that of the upper molar and co-evolved with it. Among the coevolving changes, some are clearly involved in the new upper molar phenotype. We found a similar level of coevolution in bat limbs. In conclusion, our study reveals how serial organ morphology has adapted through organ-specific developmental changes, as expected, but also through shared changes that have organ-specific effects on the final phenotype. This highlights the important role of developmental system drift in one organ to accommodate adaptation in another.

Punctuated versus gradual shifts in the multivariate evolutionary process: a test with paired radiations of scincid lizards.

As lineages become separated in time, they are expected to accumulate mutational (or developmental-genetic) differences that influence the macroevolutionary trajectories of those lineages even under similar environmental conditions. Here, we compare the dynamics of phenotypic evolution in radiations of scincid lizards from Australia and Madagascar that are separated by more than 100 million years of independent evolution and show rampant phenotypic parallelism. We collected linear measurements of the skull, limbs, and limb girdles from micro-CT scans of 94 Australian and 29 Malagasy species. Using multivariate comparative methods, we tested whether the underlying evolutionary covariance structure for this superficial parallelism was conserved and whether these patterns were consistent across distinct functional modules. Malagasy and most Australian skinks have similar covariance matrices for skull evolution. Results are ambiguous for limbs and limb girdles, as some trait subsets support different evolutionary processes and for other subsets, a shared covariance matrix could not be rejected. However, across most trait sets, the extremely speciose Australian genus Ctenotus exhibits a radically different covariance structure from all other lizards in these groups, including several closely related genera. The shift in Ctenotus demonstrates that the architecture of trait correlations can change at relatively shallow timescales and may explain the unique position of this clade in morphospace relative to other scincid lizards from both geographic regions. More generally, our results demonstrate that the multivariate evolutionary process can change dramatically in a relatively short period of time.

Double trouble: two retrotransposons triggered a cascade of invasions in Drosophila species within the last 50 years

Horizontal transfer of genetic material in eukaryotes has rarely been documented over short evolutionary timescales. Here, we show that two retrotransposons, Shellder and Spoink, invaded the genomes of multiple species of the melanogaster subgroup within the last 50 years. Through horizontal transfer, Spoink spread in D. melanogaster during the 1980s, while both Shellder and Spoink invaded D. simulans in the 1990s. Possibly following hybridization, D. simulans infected the island endemic species D. mauritiana (Mauritius) and D. sechellia (Seychelles) with both TEs after 1995. In the same approximate time-frame, Shellder also invaded D. teissieri, a species confined to sub-Saharan Africa. We find that the donors of Shellder and Spoink are likely American Drosophila species from the willistoni, cardini, and repleta groups. Thus, the described cascade of TE invasions could only become feasible after D. melanogaster and D. simulans extended their distributions into the Americas 200 years ago, likely aided by human activity. Our work reveals that cascades of TE invasions, likely initiated by human-mediated range expansions, could have an impact on the genomic and phenotypic evolution of geographically dispersed species. Within a few decades, TEs could invade many species, including island endemics, with distributions very distant from the donor of the TE.

Interaction between perfluoro-octanoic sulfonate and common antibiotics induces developmental anomalies and lethality in Xenopus laevis.

Perfluoroalkyl substances (PFAS) are persistent environmental contaminants previously used for industrial purposes as a non-stick coating and flame retardant. The stability of these molecules prevents their breakdown, which results in ground water contamination across the globe. Perfluoroalkyl substances molecules are known to bioaccumulate in various organisms. However, the health consequences remain unclear due to the large number of molecules in the PFAS family and different effects on various tissues. Here, we use the frog Xenopus laevis to investigate the developmental consequences of exposure to the PFAS molecule perfluoro-octanoic sulfonate (PFOS). We find that exposure to high levels of PFOS results in significant axial shortening of developing tadpoles. Further, we find that PFOS exposure results in a dose-dependent formation of a cellular mass in the dorsal fin. Unexpectedly, we found that these developmental phenotypes are exacerbated upon co-exposure with commonly used antibiotics. Specifically, PFOS and gentamicin co-treatment results in increased apoptosis, loss of cellular integrity, and increased overall lethality. Our results suggest a mechanism whereby gentamicin reaches levels that are toxic to mitochondria only in the presence of PFOS. These findings add to our understanding of PFOS exposure to vertebrate development and present an added concern with potential interactions with antibiotics.

Phenotypic evolution of SARS-CoV-2 spike during the COVID-19 pandemic

SARS-CoV-2 variants are mainly defined by mutations in their spike. It is therefore critical to understand how the evolutionary trajectories of spike affect virus phenotypes. So far, it has been challenging to comprehensively compare the many spikes that emerged during the pandemic in a single experimental platform. Here we generated a panel of recombinant viruses carrying different spike proteins from 27 variants circulating between 2020 and 2024 in the same genomic background. We then assessed several of their phenotypic traits both in vitro and in vivo. We found distinct phenotypic trajectories of spike among and between variants circulating before and after the emergence of Omicron variants. Spike of post-Omicron variants maintained enhanced tropism for the nasal epithelium and large airways but displayed, over time, several phenotypic traits typical of the pre-Omicron variants. Hence, spike with phenotypic features of both pre- and post-Omicron variants may continue to emerge in the future.

A Deep Learning Model for Accurate Segmentation of the Drosophila melanogaster Brain from Micro-CT Imaging.

Micro-CT data can be automatically segmented and quantified using a deep learning model trained on as few as 3 samples, facilitating rapid comparison of developmental phenotypes.

Herbivory and water availability interact to shape the adaptive landscape in the perennial forb, Boechera stricta.

Abiotic and biotic factors interact to influence phenotypic evolution; however, identifying the causal agents of selection that drive the evolution and expression of traits remains challenging. In a field common garden, we manipulated water availability and herbivore abundance across three years, and evaluated clinal variation in functional traits and phenology, plasticity, local adaptation, and selection using diverse accessions of the perennial forb, Boechera stricta. Consistent with expectations, drought stress exacerbated damage from herbivores. Foliar traits exhibited greater plasticity than phenological traits, which displayed more consistent genetic clines. Water availability and herbivory interacted to exert selection, even on traits like flowering duration, which showed no clinal variation and limited plasticity. Furthermore, the direction of selection on specific leaf area in response to water availability mirrored the genetic cline and plasticity, suggesting that variation in water levels across the landscape influences the evolution of this trait. Finally, both herbivory and water availability likely contribute to local adaptation. This work emphasizes the additive and synergistic roles of abiotic and biotic factors in shaping phenotypic variation across environmental gradients.

Developmental phenotype and quality of life in SLC13A5 citrate transporter disorder.

To describe the neurodevelopment and quality of life in SLC13A5 (solute carrier family 13 member 5) citrate transporter disorder (developmental and epileptic encephalopathy 25, DEE25), a rare genetic early infantile epileptic encephalopathy caused by deficiency of a sodium-citrate transporter, characterized by heavy seizure burden in the neonatal period. We analyzed longitudinal neurodevelopmental outcomes from a prospective natural history study of DEE25, using standardized assessments of Mullen Scales of Early Learning, Peabody Developmental Motor Scales, and Vineland Adaptive Behavior Scales. There was significant global impairment across the cohort, with variable quality of life and limited genotype-phenotype correlation. Patient-specific scores were stable across visits with evidence of modest gains in early childhood and static skills in adolescence and adulthood. There is a poor prognosis in terms of multiple measures of age-appropriate development.

The emergence and evolution of gene expression in genome regions replete with regulatory motifs.

Gene regulation is essential for life and controlled by regulatory DNA. Mutations can modify the activity of regulatory DNA, and also create new regulatory DNA, a process called regulatory emergence. Non-regulatory and regulatory DNA contain motifs to which transcription factors may bind. In prokaryotes, gene expression requires a stretch of DNA called a promoter, which contains two motifs called -10 and -35 boxes. However, these motifs may occur in both promoters and non-promoter DNA in multiple copies. They have been implicated in some studies to improve promoter activity, and in others to repress it. Here, we ask whether the presence of such motifs in different genetic sequences influences promoter evolution and emergence. To understand whether and how promoter motifs influence promoter emergence and evolution, we start from 50 'promoter islands', DNA sequences enriched with -10 and -35 boxes. We mutagenize these starting 'parent' sequences, and measure gene expression driven by 240,000 of the resulting mutants. We find that the probability that mutations create an active promoter varies more than 200-fold, and is not correlated with the number of promoter motifs. For parent sequences without promoter activity, mutations created over 1500 new -10 and -35 boxes at unique positions in the library, but only ~0.3% of these resulted in de-novo promoter activity. Only ~13% of all -10 and -35 boxes contribute to de-novo promoter activity. For parent sequences with promoter activity, mutations created new -10 and -35 boxes in 11 specific positions that partially overlap with preexisting ones to modulate expression. We also find that -10 and -35 boxes do not repress promoter activity. Overall, our work demonstrates how promoter motifs influence promoter emergence and evolution. It has implications for predicting and understanding regulatory evolution, de novo genes, and phenotypic evolution.

Chromosome-level genome assembly and annotation of a sea toad (Chaunax sp.)

The sea toad genus Chaunax is a group of small benthic fishes that predominantly inhabiting the deep seas of the Atlantic, Indian, and Pacific Oceans. Although they have the potential to make excellent systems for studies of evolutionary adaptation to deep-sea environments, genomic research on Chaunax has been hindered by a scarcity of high-quality genomic resources. We present a chromosome-scale genome assembly of a Chaunax specimen generated using PacBio long-read sequencing and high-throughput chromosome conformation capture technology. The size of the assembled genome was 706.94 Mb, with a contig N50 of 15.24 Mb and scaffold N50 of 29.42 Mb. Approximately 96.11% of assembled sequences were anchored and oriented onto 24 pseudo-chromosomes. The genome contained 213.47 Mb repetitive sequences, 25,280 protein-coding genes, and 5,090 non-coding RNAs. The high ratio of complete BUSCO genes (97.20%) indicates high quality of genome assembly. The chromosomal-level reference genome of Chaunax sp. provides a preliminary molecular basis for understanding deep-sea adaptation and phenotypic evolution as well as an important reference for whole-genome sequencing of related species.

Integrative multi-omics analysis reveals the contribution of neoVTX genes to venom diversity of Synanceia verrucosa

BackgroundAnimal venom systems are considered as valuable model for investigating the molecular mechanisms underlying phenotypic evolution. Stonefish are the most venomous and dangerous fish because of severe human envenomation and occasionally fatalities, whereas the genomic background of their venom has not been fully explored compared with that in other venomous animals.ResultsIn this study, we followed modern venomic pipelines to decode the Synanceia verrucosa venom components. A catalog of 478 toxin genes was annotated based on our assembled chromosome-level genome. Integrative analysis of the high-quality genome, the transcriptome of the venom gland, and the proteome of crude venom revealed mechanisms underlying the venom complexity in S. verrucosa. Six tandem-duplicated neoVTX subunit genes were identified as the major source for the neoVTX protein production. Further isoform sequencing revealed massive alternative splicing events with a total of 411 isoforms demonstrated by the six genes, which further contributed to the venom diversity. We then characterized 12 dominantly expressed toxin genes in the venom gland, and 11 of which were evidenced to produce the venom protein components, with the neoVTX proteins as the most abundant. Other major venom proteins included a presumed CRVP, Kuntiz-type serine protease inhibitor, calglandulin protein, and hyaluronidase. Besides, a few of highly abundant non-toxin proteins were also characterized and they were hypothesized to function in housekeeping or hemostasis maintaining roles in the venom gland. Notably, gastrotropin like non-toxin proteins were the second highest abundant proteins in the venom, which have not been reported in other venomous animals and contribute to the unique venom properties of S. verrucosa.ConclusionsThe results identified the major venom composition of S. verrucosa, and highlighted the contribution of neoVTX genes to the diversity of venom composition through tandem-duplication and alternative splicing. The diverse neoVTX proteins in the venom as lethal particles are important for understanding the adaptive evolution of S. verrucosa. Further functional studies are encouraged to exploit the venom components of S. verrucosa for pharmaceutical innovation.

Social preferences in chickens–effects of domestication and tameness

It has been suggested that evolution of domesticated phenotypes may have evolved as a result of correlated selection responses to reduced fear, a prominent feature in early domestication. To investigate whether domestication changes in social preferences can be attributed to increased tameness, we studied two lines of Red Junglefowl, ancestors of domesticated chickens, bidirectionally selected during 12 generations for high (HF) or low (LF) fear of humans and compared the differences between these lines to those between unselected Red Junglefowl (RJF) and domesticated White Leghorn egg layers (WL). One bird at a time was observed on its own for 12 min in an arena with one adjacent pen behind netting on each side. One of the adjacent pens contained familiar birds, and the other contained unfamiliar birds. Towards the end of the observation period, a sudden stressful stimulus was displayed, and we compared the reactions of the breeds to this as well. Male RJF spent more time close to the unfamiliar birds than WL, and performed more agonistic behaviour, but this was not mirrored in the selected birds. For females, it was WL that performed the most agonistic behaviour. Both LF and WL males showed more non-agonistic social exploration than HF and RJF respectively. Male LF and WL emitted more food calls than HF and RJF respectively. We conclude that several differences between RJF and WL were mirrored in the selection lines, with LF behaving more like domesticated WL. This is in line with the hypothesis that selection for tameness may have driven domestication related changes in social behaviour against conspecifics.

Independent Genetic Mapping Experiments Identify Diverse Molecular Determinants of Host Adaptation in a Generalist Herbivore.

Interactions between plants and herbivores promote evolutionary change. Studying the evolution of herbivore mechanisms aimed to cope with different host plant species is a critical intersection between evolutionary biology and sustainable pest management. Generalist herbivores are of particular interest, as hybridization between genetically distinct populations can increase the standing genetic variation and therefore the adaptive potential of the species. Tetranychus urticae is a generalist arthropod known for its adaptive potential, evidenced in its immense host range and ability to develop metabolic resistance to xenobiotics. However, the molecular underpinnings associated with the potential of host adaptation and the consequences of host adaptation in this, and many other pests remain elusive. Here, we use two independent, empirical approaches to identify and map the genetic basis of host plant performance and adaptation in genetically distinct populations of T. urticae. In the first approach, we subject a genetically diverse mite population to tomato selection and map genomic regions linked to the phenotypic evolution of increased reproductive performance. In the second approach, we map genomic regions responsible for performance on tomato by comparing the genomes of pooled individuals from an F2 backcross between populations with high and low reproductive performance. Both approaches revealed specific and shared genomic regions associated with host plant performance and adaptation and key candidate genes were identified. Our findings highlight the power of spider mite genetic approaches to identify the complex genetic basis of host adaptation in generalist herbivores.