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A Functionally Conserved yet Dynamically Evolving Toolkit Underpinning Molluscan Biomineralization: Insights From Shell and Radula.

The molluscan shell and radula constitute pivotal molluscan innovations, each characterized by distinct functions and diverse forms, regulated by the highly specific biomineralization regulatory networks. Despite their paramount importance, the conserved components and adaptive evolutionary processes governing these regulatory networks remain unresolved. To address this knowledge gap, we advocate for the integration of data from less-explored lineages, such as Scaphopoda, as an essential step. This study presents the inaugural comprehensive transcriptome analysis of Pictodentalium vernedei, a representative species of Scaphopoda distinguished by a unique and evolutionarily conserved shell morphology and radula structure. Furthermore, comparative transcriptome/genome analyses are employed to unravel the conservatism and evolutionary innovation of the involved biomineralization regulatory elements. Our findings underscore the central role of secretomes in governing biomineralization processes, and we identified a fundamental set of 26 domains within molluscan secretomes, forming an essential functional protein domain repertoire necessary for the transformation of inorganic ions into biomineralized structures. This core biomineralization toolkit has undergone independent expansion and lineage-specific recruitment, giving rise to novel, modular domain architectures. This may be essential for the functional specialization and morphological diversification of shell and radula structures. These evolutionary processes are driven by the independent co-option of ancient genes and the emergence of novel de novo genes. This comprehensive investigation not only contributes insights into the evolution of molluscan biomineralization structures but also establishes avenues for further scholarly exploration.

Animals

Variations in abundance: genome-wide responses to genetic variation and vice versa.

How do naturally occurring polymorphisms in DNA sequence relate to variation in gene expression? Recent work to map genetic sources of expression variation has shown a surprising balance between cis and trans effects. Other work suggests some chromosomal clustering of genes by expression pattern. A synthesis of approaches may provide new insight in to adaptive mechanisms in evolution and the population basis of complex traits.

Animals

Multilocus enzymes in man.

Two or more separate loci may be concerned in coding for enzyme proteins with the same or very similar catalytic properties. The phenomenon appears to be remarkably common and rough estimates suggest that perhaps 50% or more of structural loci coding for enzyme proteins are concerned with these so-called multilocus enzymes. The separate enzymes in such a multilocus set ressemble each other not only in their enzyme properties, but also in subunit structures and in molecular sizes. However there are often remarkable differences in expression of the several loci of a set from tissue to tissue. Such sets of loci probably arose in evolution by gene duplications with subsequent divergence by point mutations. Tissue differentiation in expression suggests that, pari passu with the divergence of the structural genes, there was an evolution of genetic regulating systems controlling their tissue expression.

Biological Evolution

Molecular evolution and immune expression analysis of ELF transcription factors in Lethenteron reissneri.

As important members of the ETS superfamily, the E74-like factor (ELF) transcription factor family regulates gene transcription through a conserved ETS domain and plays critical roles in immune regulation. However, the evolutionary characteristics and functions of this family in lampreys (Lethenteron reissneri) remain unclear. In this study, the ELF gene family of lampreys (Lr-ELF1, Lr-ELF2, Lr-ELF3, and Lr-ELF5) was systematically identified, and their molecular evolutionary features and immune response functions were investigated. Phylogenetic analysis revealed evolutionary characteristics reflecting the transition from jawless to jawed vertebrates. Domain architecture, gene structure, and three-dimensional structural analyses indicated that these genes appear to be conserved among vertebrates, with their three-dimensional structures showing high similarity to the core secondary structural elements of human homologous proteins. Synteny analysis demonstrated significant differences in the genomic neighborhoods of ELF genes between lampreys and jawed vertebrates. Quantitative real-time PCR (qRT-PCR) was performed with three biological and three technical replicates; relative expression levels were calculated using the ΔCt method, and statistical analysis was carried out with GraphPad Prism 9. Quantitative real-time PCR (qRT-PCR) results suggested that the ELF gene family may be involved in immune defense. This study not only enriches our understanding of the evolution of ELF genes but also provides new insights into the roles of lamprey ELFs in immune defense.

Animals

Genome-wide identification, characterization, evolutionary analysis, and expression profiling of the FCS-like zinc finger (FLZ) gene family in soybean (Glycine max L.) under abiotic stresses.

Drought and salinity limit soybean yield. Despite their role in the SnRK1 energy-sensing complex, a systematic study of FCS-Like Zinc Finger (FLZ) proteins in soybean has not been reported. We performed a genome-wide identification of the GmFLZ gene family, identifying 40 members distributed across 18 of the 20 soybean chromosomes. Phylogenetic analysis of 87 FLZ proteins from Glycine max, Arabidopsis thaliana, and Oryza sativa revealed four major evolutionary clades, suggesting that diversification predates the separation of monocots and dicots. Structural analysis identified ten conserved motifs, with Motifs 1 and 2 present in all family members. Gene duplication analysis identified 304 paralogous pairs, most arising from segmental duplication. Ka/Ks analysis indicated localized positive selection in six gene pairs and purifying selection in 97.9% of pairs. Tissue-specific expression profiling across nine tissues showed that GmFLZ5, GmFLZ15, GmFLZ25, and GmFLZ34 had the highest expression levels detected across the GmFLZ family, with GmFLZ5 the most highly expressed member in leaves, nodules, and stem and showing moderate expression in pod, root, and root hairs, whereas GmFLZ18, GmFLZ23, and GmFLZ37 showed root-preferential expression. RT-qPCR validation under drought (20% PEG-6000) and salt (200 mM NaCl) treatments in the Giza 5 cultivar showed that 36 and 34 of the 40 GmFLZ genes, respectively, exhibited at least a two-fold change in expression, with GmFLZ21 and GmFLZ35 among the most strongly induced under salt stress. These findings provide an evolutionary and functional framework for the GmFLZ family and identify candidate genes for future functional studies in soybean stress tolerance.

Glycine max

Endogenous primate and feline type C viruses.

1. Endogenous type C viruses have been detected in a wide variety of mammalian species. Multiple copies of related, but not identical, virogene sequences can be found in the DNA of these species. 2. The endogenous type C virogenes are subject to the pressures of natural selection, and closely related species possess related virogene sequences. These genes evolve as cellular entities diverging from one another in a manner which correlates well with taxonomic relatedness of the species. 3. The endogenous type C viruses of baboons and domestic cats are related, but they can be distinguished by biologic and immunologic criteria and by partial nucleic acid sequence homology. Virogene sequences in the DNA of Old World monkeys and domestic cats also show a degree of relatedness not shared by the unique sequence DNA of these species. The data suggest that progenitors of domestic cats were exogenously infected by a type C virus that also gave rise to present-day endogenous type C viruses of Old World monkeys. 4. The genomes of exogenously infectious viruses replicating in permissive host cells appear to evolve much more rapidly than endogenous virogenes which replicate as cellular genes. Laboratory strains of efficiently oncogenic type C viruses are presumed to be derived from activated endogenous viruses which have been selected for virulence and which, in certain cases, have acquired the capacity to replicate in the host's own cells. 5. The ubiquitous presence of endogenous type C viruses among vertegrates and their preservation throughout millions of years of evolution suggests that these genes express normal physiologic functions which provide a selective advantage to the species.

Animals

Long-range mRNA folding shapes expression and sequence of bacterial genes.

Bacterial gene expression is strongly influenced by local mRNA secondary structure, yet the impact of long-range folding remains poorly understood. Here, we show that sequences hundreds of nucleotides from the mRNA 5' end can act as potent repressors of gene expression through long-range base pairing to the ribosome binding site (RBS), subjecting anti-RBS sequences to negative selection. Using massively parallel reporter assays in Bacillus subtilis, we identify anti-RBS sequences as among the strongest determinants of reduced mRNA abundance across the transcript body. We demonstrate that distal anti-RBS elements engage in long-range folding with the Shine-Dalgarno sequence, blocking ribosome entry and promoting mRNA decay. Consistent with these repressive effects, anti-RBS-like sequences are depleted throughout diverse bacterial coding sequences but not from leaderless transcripts, and introducing distal anti-RBS to native genes reduces expression. Our findings establish that long-range mRNA folding is a conserved force shaping gene expression and constrains coding sequence evolution.

Bacillus subtilis

Tandem duplication-driven expansion and UV-B stress adaptation of the LHC gene family in Artemisia annua L.

BACKGROUND: Artemisia annua L., is the primary natural source of the antimalarial drug artemisinin. In nature, fluctuating light is a major environmental stress that affects plant growth and artemisinin biosynthesis. Although the light-harvesting chlorophyll a/b-binding (LHC) superfamily plays a key role in mediating plant responses to fluctuating light, systematic research of this gene family in A. annua has not yet been conducted, limiting our understanding of light adaptation in this medicinally important species. RESULTS: This study investigated the evolutionary dynamics and functional adaptation of the light-harvesting chlorophyll a/b-binding (LHC) superfamily in A. annua, with a focus on the early light‑induced protein (ELIP) subfamily. Comparative genomics of 24 plant species showed that the LHC superfamily recently expanded in the examined Asteraceae lineages through duplication events. In A. annua, 229 LHC genes identified from four haplotype genomes comprised 205 allelic and 24 haplotype-specific loci, with the ELIP subfamily expanding significantly via tandem duplication. Notably, compared to non-Asteraceae plants, ELIPs exhibited a uniform single-exon architecture, indicating it is a genomic feature unique to Asteraceae plants. Population genomics of 41 individuals showed dynamic copy number variations ranging from 1 to 4 copies per locus. Interestingly, a structurally disrupted ELIP allele remained transcriptionally active and produced long aberrant transcripts, showing that this subfamily is still actively evolving. Under UV-B stress, AaELIP loci showed synchronized induction trend but differed in expression levels, suggesting a division into major and auxiliary roles within the expanded tandem cluster. Overall, while the response of ELIPs to light stress is evolutionarily conserved, this dramatic expansion and structural streamlining of AaELIPs may represent a key evolutionary adaptation that enhances the plant's ability to cope with intense light and radiation stress. CONCLUSIONS: Collectively, this study demonstrates a significant expansion of the LHC superfamily in A. annua, especially within the ELIP subfamily, as well as its robust response to UV-B treatment, underscoring the essential role of ELIPs in mediating light stress responses. These findings provide a valuable foundation for future research to uncover the molecular mechanisms underlying A. annua's adaptation to complex light environments.

Artemisia annua

Evolutionary characterization and expression profiling of ACC and FASN genes in Chinese mitten crab Eriocheir sinensis.

Acetyl-CoA carboxylase (ACC) and fatty acid synthase (FASN) are rate-limiting enzymes in the fatty acid biosynthetic pathway, yet their evolutionary relationships, sequence features, and expression profiles remain poorly understood in crustaceans, particularly in the economically important Chinese mitten crab (Eriocheir sinensis). Here, we identified and systematically analyzed ACC and FASN genes in E. sinensis using comparative genomic analyses across 43 species. ACC was highly conserved as a single-copy gene in invertebrates, in contrast to the multiple paralogs observed in vertebrates. Similarly, FASN was generally maintained as a single-copy gene across most taxa but exhibited lineage-specific expansion in certain insect groups. Phylogenetic and structural analysis revealed strong conservation of both genes within crustaceans, supported by multiple conserved motifs and canonical functional domains. Expression profiling showed predominant expression in the hepatopancreas and midgut, suggesting their potential involvement in crustacean lipid metabolism. During the molting cycle, ACC and FASN exhibited higher expression levels during stages C and D, suggesting an increased capacity for fatty acid biosynthesis before molting. In addition, dietary lipid levels experiment revealed that ACC and FASN expression responded dynamically to dietary lipid availability, with increased expression at moderate lipid levels but reduced expression under excessive lipid supplementation, indicating a possible adaptive transcriptional response to lipid status. Collectively, this study provides insights into the evolutionary conservation and expression dynamics of ACC and FASN and improves our understanding of lipid metabolic adaptation in crustaceans.

Animals

Evolution of the differential regulation of duplicate genes after polyploidization.

In the 50 million years since the polyploidization event that gave rise to the catostomid family of fishes the duplicate genes encoding isozymes have undergone different fates. Ample opportunity has been available for regulatory evolution of these duplicate genes. Approximately half the duplicate genes have lost their expressions during this time. Of the duplicate genes remaining, the majority have diverged to different extents in their expression within and among adult tissues. The pattern of divergence of duplicate gene expression is consistent with the accumulation of mutations at regulatory genes. The absence of a correlation of extent of divergence of gene expression with the level of genetic variability for isozymes at these loci is consistent with the view that the rates of regulatory gene and structural gene evolution are uncoupled. The magnitude of divergence of duplicate gene expressions varies among tissues, enzymes, and species. Little correlation was found with the extent of divergence of duplicate gene expression within a species and its degree of morphological "conservatism", although species pairs which are increasingly taxonomically distant are less likely to share specific patterns of differential gene expression. Probable phylogenetic times of origin of several patterns of differential gene expression have been proposed. Some patterns of differential gene expression have evolved in recent evolutionary times and are specific to one or a few species, whereas at least one pattern of differential gene expression is present in nearly all species and probably arose soon after the polyploidization event. Multilocus isozymes, formed by polyploidization, provide a useful model system for studying the forces responsible for the maintenance of duplicate genes and the evolution of these once identical genes to new spatially and temporally specific patterns of regulation.

Animals

Genome-wide analysis of polymerase III-transcribed Alu elements suggests cell-type-specific enhancer function.

Alu elements are one of the most successful families of transposons in the human genome. A portion of Alu elements is transcribed by RNA Pol III, whereas the remaining ones are part of Pol II transcripts. Because Alu elements are highly repetitive, it has been difficult to identify the Pol III-transcribed elements and quantify their expression levels. In this study, we generated high-resolution, long-genomic-span RAMPAGE data in 155 biosamples all with matching RNA-seq data and built an atlas of 17,249 Pol III-transcribed Alu elements. We further performed an integrative analysis on the ChIP-seq data of 10 histone marks and hundreds of transcription factors, whole-genome bisulfite sequencing data, ChIA-PET data, and functional data in several biosamples, and our results revealed that although the human-specific Alu elements are transcriptionally repressed, the older, expressed Alu elements may be exapted by the human host to function as cell-type-specific enhancers for their nearby protein-coding genes.

Alu Elements

A General Principle of Neuronal Evolution Reveals a Human-Accelerated Neuron Type Potentially Underlying the High Prevalence of Autism in Humans.

The remarkable ability of a single genome sequence to encode a diverse collection of distinct cell types, including the thousands of cell types found in the mammalian brain, is a key characteristic of multicellular life. While it has been observed that some cell types are far more evolutionarily conserved than others, the factors driving these differences in the evolutionary rate remain unknown. Here, we hypothesized that highly abundant neuronal cell types may be under greater selective constraint than rarer neuronal types, leading to variation in their rates of evolution. To test this, we leveraged recently published cross-species single-nucleus RNA-sequencing datasets from three distinct regions of the mammalian neocortex. We found a strikingly consistent relationship where more abundant neuronal subtypes show greater gene expression conservation between species, which replicated across three independent datasets covering >106 neurons from six species. Based on this principle, we discovered that the most abundant type of neocortical neurons-layer 2/3 intratelencephalic excitatory neurons-has evolved exceptionally quickly in the human lineage compared to other apes. Surprisingly, this accelerated evolution was accompanied by the dramatic down-regulation of autism-associated genes, which was likely driven by polygenic positive selection specific to the human lineage. In summary, we introduce a general principle governing neuronal evolution and suggest that the exceptionally high prevalence of autism in humans may be a direct result of natural selection for lower expression of a suite of genes that conferred a fitness benefit to our ancestors while also rendering an abundant class of neurons more sensitive to perturbation.

Humans

Pan-genome characterization of the maize 4CL gene family and its dynamic responses to abiotic stress.

1.Pan-genome analysis across 26 maize inbred lines identified 13 Zm4CL genes (nine core and four near-core) classified into three evolutionary clades.2.Structural variations (SVs) are significantly associated with the expression and altered conserved protein domains of key Zm4CL genes.3.Zm4CL genes exhibit distinct tissue-specific expression patterns and dynamic enzymatic and transcriptional responses to stresses, particularly cold and drought.4-Coumarate:CoA ligase (4CL) is a key enzyme in the phenylpropanoid pathway and plays important roles in plant growth, development, and responses to environmental stresses. However, a comprehensive pan-genome analysis of the 4CL gene family in maize is still lacking. In this study, 13 Zm4CL genes were identified from a maize pan-genome comprising 26 diverse inbred lines, including nine core genes and four near-core genes. Phylogenetic analysis classified these genes into three evolutionary clades, while Ka/Ks analysis indicated that most members have been maintained under purifying selection, although several genes exhibited greater evolutionary divergence and relatively relaxed evolutionary constraints. Structural variation (SV) analysis revealed significant associations between SVs and the expression of Zm4CL2 and Zm4CL3, while sequence comparisons suggested that SVs were also associated with alterations in conserved protein domains in some genotypes. Transcriptome analyses revealed distinct tissue-specific expression patterns and diverse transcriptional responses to abiotic and biotic stresses. Enzyme activity assays showed that cold stress significantly increased 4CL activity at 12 h, whereas heat, salt, and alkali stresses caused an initial decrease followed by recovery, while drought had no significant effect. Time-course RT-qPCR further validated dynamic expression changes of representative Zm4CL genes under cold and drought stresses. Overall, this study provides a comprehensive pan-genome framework for understanding the evolutionary conservation, regulatory diversification, and stress-responsive characteristics of the maize Zm4CL gene family, providing valuable resources for future functional studies and the genetic improvement of stress tolerance in maize.

Zea mays

Co-option of ancestral stem regulators drove recurrent evolution of underground storage organs.

Geophytes are plants that produce underground storage organs such as tubers, rhizomes, and bulbs, to facilitate asexual reproduction and withstand a myriad of environmental challenges. While the potato (Solanum tuberosum L.) serves as the primary model for studying tuberization, the genetic mechanisms encoding this trait across diverse angiosperm lineages remains unclear. This study utilized a phylogenomic-transcriptomic approach to compare tuber development across nine tuberizing species with five nontuberizing sister taxa. We identified orthologs of key potato tuberization genes that exhibit similar expression in the stolons or tubers of these distant relatives. In nontuberizing species, these orthologs exhibit distinct expression profiles and are primarily expressed in the stem. This suggests that the independent evolution of tubers across angiosperms resulted from shifts in the expression of preexisting genes that led to their co-option. This process, also known as exaptation, occurs when existing genetic suites are recruited for entirely new biological functions. This mechanism stands in contrast to the repeated loss or gain of genes, which has been associated with the origin of other adaptive plant traits. Furthermore, the co-option of the same genes was observed in species with other stem-derived storage organs, such as rhizomes and runners. These findings reveal a conserved evolutionary model for the development of stem-derived geophyte organs that evolved independently across the flowering plants over the past 160 My.

Plant Tubers

Mixed Evidence that Dosage Sensitive Genes Drive Global Dosage Compensation in Flour Beetles.

Heteromorphic sex chromosomes create inherent gene dosage differences between males and females because one sex carries a single copy of the X chromosome while the other carries two. Many species have evolved mechanisms that equalize X-linked gene expression between the sexes and, in some cases, restore ancestral autosomal levels, a process known as dosage compensation. Although chromosome-wide compensation is common in male heterogametic (XY) insects, regulatory outcomes vary across taxa and sex chromosome systems, leaving the evolutionary forces shaping sex chromosome regulation unresolved. One hypothesis proposes that the extent to which genes are sensitive to changes in gene dose determines whether complete compensation evolves. We tested predictions of this insensitive sex chromosome hypothesis (ISCH) across five flour beetle species using comparative transcriptomics and genome-wide RNAi-derived measures of gene-by-gene sensitivity. Including an X-autosome fusion in Tribolium confusum allowed direct assessment of expression evolution following a transition from a diploid autosome to a hemizygous Neo-X. Across all five species, we detect complete chromosome-wide dosage compensation and balance between the sexes in somatic tissues, including the Neo-X region. Consistent with ISCH predictions, neither the ancestral Shared-X nor the Neo-X is depleted of genes that are sensitive to RNAi-based expression disruption. However, contrary to expectations, at the level of individual genes, we find little evidence that more sensitive genes exhibit reduced expression divergence. These results suggest that chromosome-wide compensation can be maintained by global regulatory mechanisms that persist through sex chromosome turnover, even when gene-by-gene constraints are weak. Understanding the molecular basis of these mechanisms remains a central challenge in sex chromosome evolution.

Animals

Expression of De Novo Open Reading Frames in Natural Populations of Drosophila melanogaster.

De novo genes, which originate from noncoding DNA, are known to have a high rate of turnover over short evolutionary timescales, such as within a species. Thus, their expression is often lineage- or genetic background-specific. However, little is known about their levels and breadth of expression as populations of a species diverge. In this study, we utilized publicly available RNA-seq data to examine the expression of newly evolved open reading frames (neORFs) in comparison to non- and protein-coding genes in Drosophila melanogaster populations from the derived species range in Europe and the ancestral range in sub-Saharan Africa. Our datasets included two adult tissue types as well as whole bodies at two temperatures for both sexes and three larval/prepupal developmental stages in a single tissue and sex, which allowed us to examine neORF expression and divergence across multiple sample types as well as sex and population. We detected a relatively large proportion (approximately 50%) of annotated neORFs as expressed in the population samples, with neORFs often showing greater expression divergence between populations than non- or protein-coding genes. However, differential expression of neORFs between populations tended to occur in a sample type-specific manner. On the other hand, neORFs displayed less sex-biased expression than the other two gene classes, with the majority of sex-biased neORFs detected in whole bodies, which may be attributable to the presence of the gonads. We also found that neORFs shared among multiple lines in the original set of inbred lines in which they were first detected were more likely to be both expressed and differentially expressed in the new population samples, suggesting that neORFs at a higher frequency (i.e. present in more individuals) within a species are more likely to be functional.

Animals

Identification and characterization of G protein-coupled receptors in the nocturnal halictid bee Megalopta genalis.

G protein-coupled receptors (GPCRs) are one of the largest families of membrane proteins in insects, regulating vision, neural signal transduction, and various physiological behaviors. Megalopta genalis exhibits a unique facultatively eusocial lifestyle and possesses adaptations for nocturnal activity; however, its GPCR family has not yet been systematically characterized. In this study, we performed genome-wide identification, phylogenetic analysis, and expression profiling of GPCRs in M. genalis by integrating genomic annotation and transcriptomic analysis. The results showed that a total of 99 GPCRs were identified in the genome of M. genalis, which were classified into four major families. Here, we show that M. genalis has undergone lineage-specific GPCR repertoire remodeling, marked by the expansion of novel orphan receptors and the systematic loss of multiple receptor subtypes, such as the neuropeptide receptors MIP-R and NPFR. Moreover, opsins have formed a diverse array of combinations and non-GPCR odorant receptors have undergone significant expansion via tandem duplication. Together, these features may represent part of the molecular repertoire associated with the adaptation of M. genalis to a nocturnal lifestyle. Furthermore, transcriptomic analysis revealed distinct spatiotemporal expression divergence within each of the Mth/Mthl and Fz GPCR families, suggesting functional specialization across development and adult tissues. This study provides the first systematic identification and initial functional characterization of GPCRs in M. genalis, revealing an evolutionary pattern characterized by the coexistence of contraction and expansion within the GPCR family. These findings lay a foundation for further studies aimed at elucidating the roles of these GPCRs in regulating M. genalis physiology and behavior.

Animals

Myeloma proteins from NZB and BALB/c mice: structural and functional differences.

Structural and functional analyses of myeloma immunoglobulins from inbred BALB/c mice and humans have provided important insights into the structure of the antibody molecule and the expression and evolution of antibody genes. One important question concerning these analyses is whether the myeloma process selects, in a nonrandom manner, the lymphocytes to be transformed. The availability of myeloma tumors in a second inbred strain of mouse, NZB, permits us to approach this question. In this respect. the NH2-terminal amino acid sequences of 27 kappa light chains as well as data relating to the antigen-binding properties and immunoglobulin class distribution of NZB myeloma proteins are presented and compared with similar data from the BALB/c mouse. These studies suggest that the myeloma proteins from the BALB/c and NZB mice constitute two populations of immunoglobulins with distinct functional and structural properties. The implication of this observation are discussed.

Amino Acid Sequence