Retention of duplicated genes in evolution.

Gene duplication is a prevalent phenomenon across the tree of life. The processes that lead to the retention of duplicated genes are not well understood. Functional genomics approaches in model organisms, such as yeast, provide useful tools to test the mechanisms underlying retention with functional redundancy and divergence of duplicated genes, including fates associated with neofunctionalization, subfunctionalization, back-up compensation, and dosage amplification. Duplicated genes may also be retained as a consequence of structural and functional entanglement. Advances in human gene editing have enabled the interrogation of duplicated genes in the human genome, providing new tools to evaluate the relative contributions of each of these factors to duplicate gene retention and the evolution of genome structure.

Diverse triterpene skeletons are derived from the expansion and divergent evolution of 2,3-oxidosqualene cyclases in plants.

Triterpenoids are one of the largest groups of secondary metabolites and exhibit diverse structures, which are derived from C30 skeletons that are biosynthesized via the isoprenoid pathway by cyclization of 2,3-oxidosqualene. Triterpenoids have a wide range of biological activities, and are used in functional foods, drugs, and as industrial materials. Due to the low content levels in their native plants and limited feasibility and efficiency of chemical synthesis, heterologous biosynthesis of triterpenoids is the most promising strategy. Herein, we classified 121 triterpene alcohols/ketones according to their conformation and ring numbers, among which 51 skeletons have been experimentally characterized as the products of oxidosqualene cyclases (OSCs). Interestingly, 24 skeletons that have not been reported from nature source were generated by OSCs in heterologous expression. Comprehensive evolutionary analysis of the identified 152 OSCs from 75 species in 25 plant orders show that several pentacyclic triterpene synthases repeatedly originated in multiple plant lineages. Comparative analysis of OSC catalytic reaction revealed that stabilization of intermediate cations, steric hindrance, and conformation of active center amino acid residues are primary factors affecting triterpene formation. Optimization of OSC could be achieved by changing of side-chain orientations of key residues. Recently, methods, such as rationally design of pathways, regulation of metabolic flow, compartmentalization engineering, etc., were introduced in improving chassis for the biosynthesis of triterpenoids. We expect that extensive study of natural variation of large number of OSCs and catalytical mechanism will provide basis for production of high level of triterpenoids by application of synthetic biology strategies.

Functional reorganization of North American wintering avifauna.

Wintering birds serve as vital climate sentinels, yet they are often overlooked in studies of avian diversity change. Here, we provide a continental-scale characterization of change in multifaceted wintering avifauna and examine the effects of climate change on these dynamics. We reveal a strong functional reorganization of wintering bird communities marked by a north-south gradient in functional diversity change, along with a superimposed mild east-west gradient in trait composition change. Assemblages in the northern United States saw contractions of the functional space and increases in functional evenness and originality, while the southern United States saw smaller contractions of the functional space and stasis in evenness and originality. Shifts in functional diversity were underlined by significant reshuffling in trait composition, particularly pronounced in the western and northern United States. Finally, we find strong contributions of climate change to this functional reorganization, underscoring the importance of wintering birds in tracking climate change impacts on biodiversity.

Functional and regulatory diversity of homeobox-leucine zipper transcription factors BnaHB6 under dehydration and salt stress in Brassica napus L.

The plant-specific homeodomain-leucine zipper I subfamily is involved in the regulation of various biological processes, particularly growth, development and stress response. In the present study, we characterized four BnaHB6 homologues from Brassica napus. All BnaHB6 proteins have transcriptional activation activity. Structural and functional data indicate the complex role of BnaHB6 genes in regulating biological processes, with some functions conserved and others diverged. Transcriptional analyzes revealed that they are induced in a similar manner in different tissues but show different expression patterns in response to stress and circadian rhythm. Only the BnaA09HB6 and BnaC08HB6 genes are expressed under dehydration and salt stress, and in darkness. The partial transcriptional overlap of BnaHB6s with the evolutionarily related genes BnaHB5 and BnaHB16 was also observed. Transgenic Arabidopsis thaliana plants expressing a single proBnaHB6::GUS partially confirmed the expression results. Bioinformatic analysis allowed the identification of TF-binding sites in the BnaHB6 promoters that may control their expression under stress and circadian rhythm. ChIP-qPCR analysis revealed that BnaA09HB6 and BnaC08HB6 bind directly to the promoters of the target genes BnaABF4 and BnaDREB2A. Comparison of their expression patterns in the WT plants and the bnac08hb6 mutant showed that BnaC08HB6 positively regulates the expression of the BnaABF4 and BnaDREB2A genes under dehydration and salt stress. We conclude that four BnaHB6 homologues have distinct functions in response to stress despite high sequence similarity, possibly indicating different binding preferences with BnaABF4 and BnaDREB2A. We hypothesize that BnaC08HB6 and BnaA09HB6 function in a complex regulatory network under stress.

Multiple roles of brassinosteroid signaling in vascular development.

Brassinosteroids (BRs) are plant steroid hormones that control growth and stress responses. In the context of development, BRs play diverse roles in controlling cell differentiation and tissue patterning. The vascular system, which is essential for transporting water and nutrients throughout the plant body, initially establishes a tissue pattern during primary development and then dramatically increases the number of vascular cells during secondary development. This complex developmental process is properly regulated by a network consisting of various hormonal signalling pathways. Genetic studies have revealed that mutants defective in BR biosynthesis or the BR signalling cascade exhibit a multifaceted vascular development phenotype. Furthermore, BR crosstalk with other plant hormones, including peptide hormones, coordinately regulates vascular development. Recently, the involvement of BR in vascular development, especially in xylem differentiation, has also been suggested in plant species other than the model plant Arabidopsis thaliana. In this review, we briefly summarize the recent findings on the roles of BR in primary and secondary vascular development in Arabidopsis and other species.

Functional Divergence and Origin of the Vertebrate Praja Family.

The Praja family is an E3 ubiquitin ligase, promoting polyubiquitination and subsequent degradation of substrates. It comprises two paralogs, praja1 and praja2. Prior research suggests these paralogs have undergone functional divergence, with examples, such as their distinct roles in neurite outgrowth. However, the specific evolutionary trajectories of each paralog remain largely unexplored preventing mechanistic understanding of functional differences between paralogs. Here, we investigated the phylogeny and divergence of the vertebrate Praja family through molecular evolutionary analysis. Phylogenetic examination of the vertebrate praja revealed that praja1 and praja2 originated from the common ancestor of placentals via gene duplication, with praja1 evolving at twice the rate of praja2 shortly after the duplication. Moreover, a unique evolutionary trajectory for praja1 relative to other vertebrate Praja was indicated, as evidenced by principal component analysis on GC content, codon usage frequency, and amino acid composition. Subsequent motif/domain comparison revealed conserved N terminus and C terminus in praja1 and praja2, together with praja1-specific motifs, including nuclear localization signal and Ala-Gly-Ser repeats. The nuclear localization signal was demonstrated to be functional in human neuroblastoma SH-SY5Y cells using deletion mutant, while praja2 was exclusively expressed in the nucleus. These discoveries contribute to a more comprehensive understanding of the Praja family's phylogeny and suggest a functional divergence between praja1 and praja2. Specifically, the shift of praja1 into the nucleus implies the degradation of novel substrates located in the nucleus as an evolutionary consequence.

The evolution and functional divergence of FT-related genes in controlling flowering time in Brassica rapa ssp. rapa.

KEY MESSAGE: The BrrFT paralogues exhibit distinct expression patterns and play different roles in regulating flowering time, and BrrFT4 competes with BrrFT1 and BrrFT2 to interact with BrrFD proteins. Flowering time is an important agricultural trait for Brassica crops, and early bolting strongly affects the yield and quality of Brassica rapa ssp. rapa. Flowering Locus T paralogues play an important role in regulating flowering time. In this study, we identified FT-related genes in turnip by phylogenetic classification, and four BrrFT homoeologs that shared with high identities with BraFT genes were isolated. The different gene structures, promoter binding sites, and expression patterns observed indicated that these genes may play different roles in flowering time regulation. Further genetic and biochemical experiments showed that as for FT-like paralogues, BrrFT2 acted as the key floral inducer, and BrrFT1 seems to act as a mild 'florigen' protein. However, BrrFT4 acts as a floral repressor and antagonistically regulates flowering time by competing with BrrFT1 and BrrFT2 to bind BrrFD proteins. BrrFT3 may have experienced loss of function via base shift mutation. Our results revealed the potential roles of FT-related genes in flowering time regulation in turnip.

The type III polyketide synthase supergene family in plants: complex evolutionary history and functional divergence.

Type III polyketide synthases (PKSs) are key enzymes involved in the biosynthesis of a variety of plant specialized metabolites, including flavonoids, stilbenes, and sporopollenin, to name a few. These enzymes likely played vital roles in plant adaptation during their transition from aquatic to terrestrial habitats and their colonization of specific ecological environments. Members of this supergene family have diverse functions, but how type III PKSs and their functions have evolved remains poorly understood. Here, we conducted comprehensive phylogenomics analysis of the type III PKS supergene family in 60 species representing the major plant lineages and elucidated the classification, origin, and evolutionary history of each class. Molecular evolutionary analysis of the typical chalcone synthase and stilbene synthase types revealed evidence for strong positive natural selection in both the Pinaceae and Fabaceae lineages. The positively selected sites of these proteins include residues at the catalytic tunnel entrance and homodimer interface, which might have driven the functional divergence between the two types. Our results also suggest that convergent evolution of enzymes involved in plant flavonoid biosynthesis is quite common. The results of this study provide new insights into the origin, evolution, and functional diversity of plant type III PKSs. In addition, they serve as a guide for the enzymatic engineering of plant polyketides.

Evolution of isoform-level gene expression patterns across tissues during lotus species divergence.

Both gene duplication and alternative splicing (AS) drive the functional diversity of gene products in plants, yet the relative contributions of the two key mechanisms to the evolution of gene function are largely unclear. Here, we studied AS in two closely related lotus plants, Nelumbo lutea and Nelumbo nucifera, and the outgroup Arabidopsis thaliana, for both single-copy and duplicated genes. We show that most splicing events evolved rapidly between orthologs and that the origin of lineage-specific splice variants or isoforms contributed to gene functional changes during species divergence within Nelumbo. Single-copy genes contain more isoforms, have more AS events conserved across species, and show more complex tissue-dependent expression patterns than their duplicated counterparts. This suggests that expression divergence through isoforms is a mechanism to extend the expression breadth of genes with low copy numbers. As compared to isoforms of local, small-scale duplicates, isoforms of whole-genome duplicates are less conserved and display a less conserved tissue bias, pointing towards their contribution to subfunctionalization. Through comparative analysis of isoform expression networks, we identified orthologous genes of which the expression of at least some of their isoforms displays a conserved tissue bias across species, indicating a strong selection pressure for maintaining a stable expression pattern of these isoforms. Overall, our study shows that both AS and gene duplication contributed to the diversity of gene function during the evolution of lotus.

Divergence of active site motifs among different classes of Populus glutaredoxins results in substrate switches.

Enzymes are essential components of all biological systems. The key characteristics of proteins functioning as enzymes are their substrate specificities and catalytic efficiencies. In plants, most genes encoding enzymes are members of large gene families. Within such families, the contributions of active site motifs to the functional divergence of duplicate genes have not been well elucidated. In this study, we identified 41 glutaredoxin (GRX) genes in the Populus trichocarpa genome. GRXs are ubiquitous enzymes in plants that play important roles in developmental and stress tolerance processes. In poplar, GRX genes were divided into four classes based on clear differences in gene structure and expression pattern, subcellular localization, enzymatic activity, and substrate specificity of the encoded proteins. Using site-directed mutagenesis, this study revealed that the divergence of the active site motif among different classes of GRX proteins resulted in substrate switches and thus provided new insights into the molecular evolution of these important plant enzymes.

Functional traits explain waterbirds' host status, subtype richness, and community-level infection risk for avian influenza.

Species functional traits can influence pathogen transmission processes, and consequently affect species' host status, pathogen diversity, and community-level infection risk. We here investigated, for 143 European waterbird species, effects of functional traits on host status and pathogen diversity (subtype richness) for avian influenza virus at species level. We then explored the association between functional diversity and HPAI H5Nx occurrence at the community level for 2016/17 and 2021/22 epidemics in Europe. We found that both host status and subtype richness were shaped by several traits, such as diet guild and dispersal ability, and that the community-weighted means of these traits were also correlated with community-level risk of H5Nx occurrence. Moreover, functional divergence was negatively associated with H5Nx occurrence, indicating that functional diversity can reduce infection risk. Our findings highlight the value of integrating trait-based ecology into the framework of diversity-disease relationship, and provide new insights for HPAI prediction and prevention.

Evolution of TOP1 and TOP1MT Topoisomerases in Chordata.

Type IB topoisomerases relax the torsional stress associated with DNA metabolism in the nucleus and mitochondria and constitute important molecular targets of anticancer drugs. Vertebrates stand out among eukaryotes by having two Type IB topoisomerases acting specifically in the nucleus (TOP1) and mitochondria (TOP1MT). Despite their major importance, the origin and evolution of these paralogues remain unknown. Here, we examine the molecular evolutionary processes acting on both TOP1 and TOP1MT in Chordata, taking advantage of the increasing number of available genome sequences. We found that both TOP1 and TOP1MT evolved under strong purifying selection, as expected considering their essential biological functions. Critical active sites, including those associated with resistance to anticancer agents, were found particularly conserved. However, TOP1MT presented a higher rate of molecular evolution than TOP1, possibly related with its specialized activity on the mitochondrial genome and a less critical role in cells. We could place the duplication event that originated the TOP1 and TOP1MT paralogues early in the radiation of vertebrates, most likely associated with the first round of vertebrate tetraploidization (1R). Moreover, our data suggest that cyclostomes present a specialized mitochondrial Type IB topoisomerase. Interestingly, we identified two missense mutations replacing amino acids in the Linker region of TOP1MT in Neanderthals, which appears as a rare event when comparing the genome of both species. In conclusion, TOP1 and TOP1MT differ in their rates of evolution, and their evolutionary histories allowed us to better understand the evolution of chordates.

Defining the functional divergence of orthologous genes between human and mouse in the context of miRNA regulation.

Animal models have a certain degree of similarity with human in genes and physiological processes, which leads them to be valuable tools for studying human diseases and for assisting drug development. However, translational researches adopting animal models are largely restricted by the species heterogeneity, which is also a major reason for the failure of drug research. Currently, computational method for exploring the functional differences between orthologous genes is still insufficient. For this purpose, here, we presented an algorithm, functional divergence score (FDS), by comprehensively evaluating the functional differences between the microRNAs regulating the paired orthologous genes. Given that mouse is one of the most popular model animals, currently, FDS was designed to dissect the functional divergence of orthologous genes between human and mouse. The results showed that gene FDS value is significantly associated with gene evolutionary characteristics and can discover expression divergence of human-mouse orthologous genes. Moreover, FDS performed well in distinguishing the targets of approved drugs and the failed ones. These results suggest that FDS is a valuable tool to evaluate the functional divergence of paired human and mouse orthologous genes. In addition, for each orthologous gene pair, FDS can provide detailed differences in functions and phenotypes. Our study provided a useful tool for quantifying the functional difference between human and mouse, and the presented framework is easily to be extended to the orthologous genes between human and other species. An online server of FDS is available at http://www.cuilab.cn/fds/.

Climate change threats to the global functional diversity of freshwater fish.

Climate change impacts on freshwater ecosystems and freshwater biodiversity show strong spatial variability, highlighting the importance of a global perspective. While previous studies on biodiversity mostly focused on species richness, functional diversity, which is a better predictor of ecosystem functioning, has received much less attention. This study aims to comprehensively assess climate change threats to the functional diversity of freshwater fish across the world, considering three complementary metrics-functional richness, evenness and divergence. We built on existing spatially explicit projections of geographical ranges for 11,425 riverine fish species as affected by changes in streamflow and water temperature extremes at four warming levels (1.5°C, 2.0°C, 3.2°C and 4.5°C). To estimate functional diversity, we considered the following four continuous, morphological and physiological traits: relative head length, relative body depth, trophic level and relative growth rate. Together, these traits cover five ecological functions. We treated missing trait values in two different ways: we either removed species with missing trait values or imputed them. Depending on the warming level, 6%-25% of the locations globally face a complete loss of functional diversity when assuming no dispersal (6%-17% when assuming maximal dispersal), with hotspots in the Amazon and Paraná River basins. The three facets of functional diversity do not always follow the same pattern. Sometimes, functional richness is not yet affected despite species loss, while functional evenness and divergence are already reducing. Other times, functional richness reduces, while functional evenness and/or divergence increase instead. The contrasting patterns of the three facets of functional diversity show their complementarity among each other and their added value compared to species richness. With increasing climate change, impacts on freshwater communities accelerate, making early mitigation critically important.

Enhanced plasticity and reproductive fitness of floral and seed traits facilitate non-native species spread in mountain ecosystems.

Floral and seed traits, their relationships, and responses to abiotic constraints are considered the key determinants of the invasion success of non-native plant species. However, studies evaluating the pattern of floral and seed traits of non-native species in mountain ecosystems are lacking. In this study, we determined (a) whether the floral and seed traits of native and non-native species show similarity or dissimilarity across elevations in mountains, and (b) whether the non-native species follow different allometric patterns compared with native species. Functional variations between native and non-native species were assessed through floral and seed traits: flower count, flower display area, flower mass, specific flower area, seed count, and seed mass across an elevational gradient. Permanent plots (20 × 20 m) were laid at each 100 m elevation rise from 2000 to 4000 m a.s.l. for sampling of herbaceous plant species. The mean values of floral and seed traits such as flower display area, specific flower area, and seed count were significantly higher for non-native species compared to native species. A significant difference in trait values (flower display area, flower mass, seed count, and seed mass) between non-native species and native species was observed along the elevational gradient, except for flower count and specific flower area. The bivariate relationship revealed non-native species to exhibit a stronger relationship between flower display area ∼ flower mass, and flower display area ∼ seed mass traits than the native species. Non-native species showed enhanced reproductive ability under varying environmental conditions along an elevational gradient in mountain ecosystems. Greater flower display area and seed mass at lower elevations and a stronger overall trait-trait relationship among non-native species implied resource investment in pollinator visualization, flower mass, and seed quality over seed quantity. The study concludes that enhanced plasticity and reproductive fitness of floral and seed traits would consequently aid non-native species to adapt, become invasive, and displace native species in mountain ecosystems if the climatic barriers acting on non-native species are reduced with climate change.

Evolutionary expansion and functional divergence of sugar transporters in Saccharum (S. spontaneum and S. officinarum).

The sugar transporter (ST) family is considered to be the most important gene family for sugar accumulation, but limited information about the ST family in the important sugar-yielding crop Saccharum is available due to its complex genetic background. Here, 105 ST genes were identified and clustered into eight subfamilies in Saccharum spontaneum. Comparative genomics revealed that tandem duplication events contributed to ST gene expansions of two subfamilies, PLT and STP, in S. spontaneum, indicating an early evolutionary step towards high sugar content in Saccharum. The analyses of expression patterns were based on four large datasets with a total of 226 RNA sequencing samples from S. spontaneum and Saccharum officinarum. The results clearly demonstrated 50 ST genes had different spatiotemporal expression patterns in leaf tissues, 10 STs were specifically expressed in the stem, and 10 STs responded to the diurnal rhythm. Heterologous expression experiments in the defective yeast strain EBY.VW4000 indicated STP13, pGlcT2, VGT3, and TMT4 are the STs with most affinity for glucose/fructose and SUT1_T1 has the highest affinity to sucrose. Furthermore, metabolomics analysis suggested STP7 is a sugar starvation-induced gene and STP13 has a function in retrieving sugar in senescent tissues. PLT11, PLT11_T1, TMT3, and TMT4 contributed to breaking the limitations of the storage sink. SUT1, SUT1_T1, PLT11, TMT4, pGlcT2, and VGT3 responded for different functions in these two Saccharum species. This study demonstrated the evolutionary expansion and functional divergence of the ST gene family and will enable the further investigation of the molecular mechanism of sugar metabolism in Saccharum.

Degree of Functional Divergence in Duplicates Is Associated with Distinct Roles in Plant Evolution.

Gene duplication is a major mechanism to create new genes. After gene duplication, some duplicated genes undergo functionalization, whereas others largely maintain redundant functions. Duplicated genes comprise various degrees of functional diversification in plants. However, the evolutionary fate of high and low diversified duplicates is unclear at genomic scale. To infer high and low diversified duplicates in Arabidopsis thaliana genome, we generated a prediction method for predicting whether a pair of duplicate genes was subjected to high or low diversification based on the phenotypes of knock-out mutants. Among 4,017 pairs of recently duplicated A. thaliana genes, 1,052 and 600 are high and low diversified duplicate pairs, respectively. The predictions were validated based on the phenotypes of generated knock-down transgenic plants. We determined that the high diversified duplicates resulting from tandem duplications tend to have lineage-specific functions, whereas the low diversified duplicates produced by whole-genome duplications are related to essential signaling pathways. To assess the evolutionary impact of high and low diversified duplicates in closely related species, we compared the retention rates and selection pressures on the orthologs of A. thaliana duplicates in two closely related species. Interestingly, high diversified duplicates resulting from tandem duplications tend to be retained in multiple lineages under positive selection. Low diversified duplicates by whole-genome duplications tend to be retained in multiple lineages under purifying selection. Taken together, the functional diversities determined by different duplication mechanisms had distinct effects on plant evolution.

BDSF Is a Degradation-Prone Quorum-Sensing Signal Detected by the Histidine Kinase RpfC of Xanthomonas campestris pv. campestris.

Diffusible signal factors (DSFs) are medium-chain fatty acids that induce bacterial quorum sensing. Among these compounds, BDSF is a structural analog of DSF that is commonly detected in bacterial species, and it is the predominant in planta quorum-sensing signal in Xanthomonas campestris. How BDSF is sensed in Xanthomonas spp. and the functional diversity between BDSF and DSF remain unclear. In this study, we generated genetic and biochemical evidence that BDSF is a low-active regulator of X. campestris pv. campestris quorum sensing, whereas trans-BDSF does not seem to be a signaling compound. BDSF is detected by the sensor histidine kinase RpfC. Although BDSF has relatively low physiological activities, it binds to the RpfC sensor with a high affinity and activates RpfC autophosphorylation to a level that is similar to that induced by DSF in vitro. The inconsistency in the physiological and biochemical activities of BDSF is not due to RpfC-RpfG phosphorylation or RpfG hydrolase. Neither BDSF nor DSF controls the phosphotransferase and phosphatase activities of RpfC or the ability of RpfG hydrolase activity to degrade the bacterial second messenger cyclic di-GMP. We demonstrated that BDSF is prone to degradation by RpfB, a critical fatty acyl coenzyme A ligase involved in the turnover of DSF-family signals. rpfB mutations lead to substantial increases in BDSF-induced quorum sensing. Although DSF and BDSF are similarly detected by RpfC, our data suggest that their differential degradation in cells is the major factor responsible for the diversity in their physiological effects. IMPORTANCE The diffusible signal factor (DSF) family consists of quorum-sensing signals employed by Gram-negative bacteria. These signals are a group of cis-2-unsaturated fatty acids, such as DSF, BDSF, IDSF, CDSF, and SDSF. However, the functional divergence of various DSF signals remains unclear. The present study demonstrates that though BDSF is a low active quorum-sensing signal, it binds histidine kinase RpfC with a higher affinity and activates RpfC autophosphorylation to the similar level as DSF. Rather than regulation of enzymatic activities of RpfC and its cognate response regulator RpfG encoding a c-di-GMP hydrolase, BDSF is prone to degradation in bacterial cells by RpfB, which effectively avoided the inhibition of bacterial growth by accumulating high concentrations of BDSF. Therefore, our study sheds new light on the functional differences of quorum-sensing signals and shows that bacteria balance quorum sensing and growth by fine-tuning concentrations of signaling chemicals.

Phylogeny of g6pc1 Genes and Their Functional Divergence among Sarcopterygian Vertebrates: Implications for Thermoregulatory Strategies.

Glucose-6-phosphatase catalytic subunit 1 (G6PC1) catalyzes the final rate-limiting step in endogenous glucose production and is critically important for glucose homeostasis. Although a single g6pc1 gene is present in mammals, other vertebrates possess two to five paralogs. Functional divergence between paralogs has been reported in actinopterygians and has been implicated in the acquisition of adaptive characteristics. Such reports make sarcopterygian g6pc1 an interesting research topic because unlike the aquatic habitat of actinopterygians, sarcopterygians have successfully adapted to terrestrial environments. However, little is known about the evolution of sarcopterygian g6pc1. In the present study, the evolutionary history of sarcopterygian g6pc1 was investigated using molecular phylogeny, synteny analyses, and comparison of the genomic environment. Functional divergence between paralogs was also investigated in a reptilian species, the Japanese gecko, with a focus on gene expression in the liver. Evolutionary analyses suggested that amphibians and amniotes acquired duplicated genes independently. Among the amniotes, gene duplication occurred at the root of the reptilian-avian lineage, giving rise to g6pc1-1 and g6pc1-2 classes. While the avian lineage subsequently lost the g6pc1-1, the reptiles retained both classes. This co-occurrence of gene loss and endothermy acquisition, together with the observation that mammals possess only a single gene, suggests that the duplicated g6pc1 is dispensable for endotherms. Quantitative RT-PCR analyses revealed that the two gecko genes respond differently to E2 administration, as the expression of g6pc1-1 was downregulated by E2, whereas g6pc1-2 showed no significant response. Such paralog-specific responses suggest functional divergence between paralogs, which is possibly related to reproduction.

Genome-Wide Survey Indicates Diverse Physiological Roles of Dendrobium officinale Calcium-Dependent Protein Kinase Genes.

Calcium-dependent protein kinases (CDPKs) are crucial calcium ions (Ca2+) sensors in plants with important roles in signal transduction, plant growth, development, and stress responses. Here, we identified 24 genes encoding CDPKs in Dendrobium officinale using genome-wide analysis. The phylogenetic analysis revealed that these genes formed four groups, with similar structures in the same group. The gene expression patterns following hormone treatments and yeast two-hybrid of homologous CDPK gene pairs with Rbohs showed differences, indicating functional divergence between homologous genes. In addition, the rapid accumulation of hydrogen peroxide (H2O2) and stomatal closure was observed in response to salicylic acid (SA)/jasmonic acid (JA) stress. Our data showed that CDPK9-2 and CDPK20-4 interacted with Rboh D and Rboh H, respectively, and were implicated in the generation of H2O2 and regulation of the stomatal aperture in response to salicylic acid/jasmonic acid treatment. We believe these results can provide a foundation for the functional divergence of homologous genes in D. officinale.