Context-dependent antioxidant defense system (ADS)-based stress memory in response to recurrent environmental challenges in congeneric invasive species.

Marine ecosystems are facing escalating environmental fluctuations owing to climate change and human activities, imposing pressures on marine species. To withstand recurring environmental challenges, marine organisms, especially benthic species lacking behavioral choices to select optimal habitats, have to utilize well-established strategies such as the antioxidant defense system (ADS) to ensure their survival. Therefore, understanding of the mechanisms governing the ADS-based response is essential for gaining insights into adaptive strategies for managing environmental challenges. Here we conducted a comparative analysis of the physiological and transcriptional responses based on the ADS during two rounds of 'hypersalinity-recovery' challenges in two model congeneric invasive ascidians, Ciona robusta and C. savignyi. Our results demonstrated that C. savignyi exhibited higher tolerance and resistance to salinity stresses at the physiological level, while C. robusta demonstrated heightened responses at the transcriptional level. We observed distinct transcriptional responses, particularly in the utilization of two superoxide dismutase (SOD) isoforms. Both Ciona species developed physiological stress memory with elevated total SOD (T-SOD) and glutathione (GSH) responses, while only C. robusta demonstrated transcriptional stress memory. The regulatory distinctions within the Nrf2-Keap1 signalling pathway likely explain the formation disparity of transcriptional stress memory between both Ciona species. These findings support the 'context-dependent stress memory hypothesis', emphasizing the emergence of species-specific stress memory at diverse regulatory levels in response to recurrent environmental challenges. Our results enhance our understanding of the mechanisms of environmental challenge management in marine species, particularly those related to the ADS. Supplementary Information: The online version contains supplementary material available at 10.1007/s42995-024-00228-y.

Interplays between cis- and trans-Acting Factors for Alternative Splicing in Response to Environmental Changes during Biological Invasions of Ascidians.

Alternative splicing (AS), a pivotal biological process contributing to phenotypic plasticity, creates a bridge linking genotypes with phenotypes. Despite its importance, the AS mechanisms underlying environmental response and adaptation have not been well studied, and more importantly, the cis- and trans-acting factors influencing AS variation remain unclear. Using the model invasive congeneric ascidians, Ciona robusta, and Ciona savignyi, we compared their AS responses to environmental changes and explored the potential determinants. Our findings unveiled swift and dynamic AS changes in response to environmental challenges, and differentially alternative spliced genes (DASGs) were functionally enriched in transmembrane transport processes. Interestingly, both the prevalence and level of AS in C. robusta were lower than those observed in C. savignyi. Furthermore, these two indices were higher under temperature stresses compared to salinity stresses in C. savignyi. All the observed patterns underscore the species-specific and environmental context-dependent AS responses to environmental challenges. The dissimilarities in genomic structure and exon/intron size distributions between these two species likely contributed to the observed AS variation. Moreover, we identified a total of 11 and 9 serine/arginine-rich splicing factors (SRSFs) with conserved domains and gene structures in the genomes of C. robusta and C. savignyi, respectively. Intriguingly, our analysis revealed that all detected SRSFs did not exhibit prevalent AS regulations. Instead, we observed AS control over a set of genes related to splicing factors and spliceosome components. Altogether, our results elucidate species-specific and environmental challenge-dependent AS response patterns in closely related invasive ascidians. The identified splicing factors and spliceosome components under AS control offer promising candidates for further investigations into AS-mediated rapid responses to environmental challenges complementary to SRSFs.

Multidimensional plasticity jointly contributes to rapid acclimation to environmental challenges during biological invasions.

Rapid plastic response to environmental changes, which involves extremely complex underlying mechanisms, is crucial for organismal survival during many ecological and evolutionary processes such as those in global change and biological invasions. Gene expression is among the most studied molecular plasticity, while co- or posttranscriptional mechanisms are still largely unexplored. Using a model invasive ascidian Ciona savignyi, we studied multidimensional short-term plasticity in response to hyper- and hyposalinity stresses, covering the physiological adjustment, gene expression, alternative splicing (AS), and alternative polyadenylation (APA) regulations. Our results demonstrated that rapid plastic response varied with environmental context, timescales, and molecular regulatory levels. Gene expression, AS, and APA regulations independently acted on different gene sets and corresponding biological functions, highlighting their nonredundant roles in rapid environmental adaptation. Stress-induced gene expression changes illustrated the use of a strategy of accumulating free amino acids under high salinity and losing/reducing them during low salinity to maintain the osmotic homoeostasis. Genes with more exons were inclined to use AS regulations, and isoform switches in functional genes such as SLC2a5 and Cyb5r3 resulted in enhanced transporting activities by up-regulating the isoforms with more transmembrane regions. The extensive 3'-untranslated region (3'UTR) shortening through APA was induced by both salinity stresses, and APA regulation predominated transcriptomic changes at some stages of stress response. The findings here provide evidence for complex plastic mechanisms to environmental changes, and thereby highlight the importance of systemically integrating different levels of regulatory mechanisms in studying initial plasticity in evolutionary trajectories.

(Epi)genomic adaptation driven by fine geographical scale environmental heterogeneity after recent biological invasions.

Elucidating processes and mechanisms involved in rapid local adaptation to varied environments is a poorly understood but crucial component in management of invasive species. Recent studies have proposed that genetic and epigenetic variation could both contribute to ecological adaptation, yet it remains unclear on the interplay between these two components underpinning rapid adaptation in wild animal populations. To assess their respective contributions to local adaptation, we explored epigenomic and genomic responses to environmental heterogeneity in eight recently colonized ascidian (Ciona intestinalis) populations at a relatively fine geographical scale. Based on MethylRADseq data, we detected strong patterns of local environment-driven DNA methylation divergence among populations, significant epigenetic isolation by environment (IBE), and a large number of local environment-associated epigenetic loci. Meanwhile, multiple genetic analyses based on single nucleotide polymorphisms (SNPs) showed genomic footprints of divergent selection. In addition, for five genetically similar populations, we detected significant methylation divergence and local environment-driven methylation patterns, indicating the strong effects of local environments on epigenetic variation. From a functional perspective, a majority of functional genes, Gene Ontology (GO) terms, and biological pathways were largely specific to one of these two types of variation, suggesting partial independence between epigenetic and genetic adaptation. The methylation quantitative trait loci (mQTL) analysis showed that the genetic variation explained only 18.67% of methylation variation, further confirming the autonomous relationship between these two types of variation. Altogether, we highlight the complementary interplay of genetic and epigenetic variation involved in local adaptation, which may jointly promote populations' rapid adaptive capacity and successful invasions in different environments. The findings here provide valuable insights into interactions between invaders and local environments to allow invasive species to rapidly spread, thus contributing to better prediction of invasion success and development of management strategies.

Complementary genomic and epigenomic adaptation to environmental heterogeneity.

While adaptation is commonly thought to result from selection on DNA sequence-based variation, recent studies have highlighted an analogous epigenetic component as well. However, the relative roles of these mechanisms in facilitating population persistence under environmental heterogeneity remain unclear. To address the underlying genetic and epigenetic mechanisms and their relationship during environmental adaptation, we screened the genomes and epigenomes of nine global populations of a predominately sessile marine invasive tunicate, Botryllus schlosseri. We detected clear population differentiation at the genetic and epigenetic levels. Patterns of genetic and epigenetic structure were significantly influenced by local environmental variables. Among these variables, minimum annual sea surface temperature was identified as the top explanatory variable for both genetic and epigenetic variation. However, patterns of population structure driven by genetic and epigenetic variation were somewhat distinct, suggesting possible autonomy of epigenetic variation. We found both shared and specific genes and biological pathways among genetic and epigenetic loci associated with environmental factors, consistent with complementary and independent contributions of genetic and epigenetic variation to environmental adaptation in this system. Collectively, these mechanisms may facilitate population persistence under environmental change and sustain successful invasions across novel environments.

Neighbours matter: Effects of genomic organization on gene expression plasticity in response to environmental stresses during biological invasions.

Gene expression regulation has been widely recognized as an important molecular mechanism underlying phenotypic plasticity in environmental adaptation. However, it remains largely unexplored on the effects of genomic organization on gene expression plasticity under environmental stresses during biological invasions. Here, we use an invasive model ascidian, Ciona robusta, to investigate how genomic organization affects gene expression in response to salinity stresses during range expansions. Our study showed that neighboring genes were co-expressed and approximately 30% of stress responsive genes were physically clustered on chromosomes. Such coordinated expression was substantially affected by the physical distance and orientation of genes. Interestingly, the overall expression correlation of neighboring genes was significantly decreased under high salinity stresses, illustrating that the co-expression regulation could be disrupted by salinity challenges. Furthermore, the clustering of genes was associated with their function constraints and expression patterns - operon genes enriched in gene expression machinery had the highest transcriptional activity and expression stability. Notably, our analyses showed that the tail-to-tail genes, mainly involved in biological functions related to phosphorylation, homeostatic process, and ion transport, exhibited higher intrinsic expression variability and greater response to salinity challenges. Altogether, the results obtained here provide new insights into the effects of gene organization on gene expression plasticity under environmental challenges, hence improving our knowledge on mechanisms of rapid environmental adaptation during biological invasions.

Promoter analysis of TLR5M and TLR5S revealed NF-κB might be a conserved cis-element in TLR5S promoter of large yellow croaker.

Toll like receptor 5 (TLR5) plays a crucial role in the innate immune response by recognizing bacterial flagellin proteins. In the present study, the genomic and 5'-flanking sequences of LcTLR5M (membrane) and LcTLR5S (soluble) were cloned from the large yellow croaker, Larimichthys crocea. Then, their promoter activities were determined in human embryonic kidney (HEK293T) cells. LcTLR5M contained 4 exons and 3 introns, and LcTLR5S contained 2 exons and 1 intron. Bioinformatic prediction of transcription factor binding sites (TFBSs) indicated that the promoter structures were different between LcTLR5M and LcTLR5S. A dual luciferase assay showed that the deletion mutant -471 to +189 of LcTLR5M promoter possessed the greatest activity with 36 times activity of the control (P < 0.05). For LcTLR5S, two deletion mutants, -1687 to +106 and - 720 to +106, showed high promoter activity. Furthermore, site-directed mutation demonstrated that a -392 to -391 AT/GG substitution in Oct-1 binding site, and a -104 to -103 GG/TT and a -98 to -97 CC/AA substitution in the NF-κB binding site of TLR5S caused a significant decline of luciferase activity (P < 0.05). Moreover, the co-transfection of an NF-κB/p65 over-expression plasmid with wild type TLR5S (-720 to +106) resulted in an extremely significant increase of promoter activity by more than 9 times compared with the NF-kB mutant (P < 0.01). Our findings suggest that the genomic organization and promoter structure may differ between LcTLR5M and LcTLR5S. Oct1 and NF-κB binding sites might be cis-regulatory elements in the LcTLR5S promoter. NF-κB/p65 plays an important role in LcTLR5S promoter activation through binding with the NF-κB binding site.

Protein-mediated bioadhesion in marine organisms: A review.

Protein-mediated bioadhesion is one of the crucial physiological processes in marine organisms, by which they can firmly adhere to underwater substrates. Most marine adhesive organisms are biofoulers, causing negative effects on marine ecosystems and huge economic losses to aquaculture and maritime industries. Furthermore, adhesive proteins in these organisms are promising bionic candidates for high-performance artificial materials with great application value. In-depth understanding of the bioadhesion in marine ecosystems is of dual significance for resolving biofouling issue and developing marine bionic products. Here, we review the research progress of protein-mediated bioadhesion in marine organisms. The adhesion processes such as protein biosynthesis and secretion are similar among organisms, but the detailed features such as compositions, structures, and molecular functions of adhesive proteins are distinct. Hydroxylation, glycosylation, and phosphorylation are important post-translational modifications during the processes of adhesion. The contents of some amino acids such as glycine, tyrosine and cysteine involved in underwater adhesion are significantly higher, which is a sequence feature of barnacle cement and mussel foot proteins. The amyloid structures and conserved domains/motifs such as EGF and vWFA distributed in adhesive proteins are involved in the underwater adhesion. In addition, the oxidative cross-linking also plays an important role in marine bioadhesion. Overall, the unique and common features identified for the protein-mediated bioadhesion in diverse marine organisms here provide background information and essential reference for characterizing marine adhesive proteins and associated functional domains, formulating antifouling strategies, and developing novel biomimetic adhesives.

Local environment-driven adaptive evolution in a marine invasive ascidian (Molgula manhattensis).

Elucidating molecular mechanisms of environment-driven adaptive evolution in marine invaders is crucial for understanding invasion success and further predicting their future invasions. Although increasing evidence suggests that adaptive evolution could contribute to organisms' adaptation to varied environments, there remain knowledge gaps regarding how environments influence genomic variation in invaded habitats and genetic bases underlying local adaptation for most marine invaders. Here, we performed restriction-site-associated DNA sequencing (RADseq) to assess population genetic diversity and further investigate genomic signatures of local adaptation in the marine invasive ascidian, Molgula manhattensis. We revealed that most invasive populations exhibited significant genetic differentiation, low recent gene flow, and no signal of significant population bottleneck. Based on three genome scan approaches, we identified 109 candidate loci potentially under environmental selection. Redundancy analysis and variance partitioning analysis suggest that local environmental factors, particularly the salinity-related variables, represent crucial evolutionary forces in driving adaptive divergence. Using the newly developed transcriptome as a reference, 14 functional genes were finally obtained with potential roles in salinity adaptation, including SLC5A1 and SLC9C1 genes from the solute carrier gene (SLC) superfamily. Our findings confirm that differed local environments could rapidly drive adaptive divergence among invasive populations and leave detectable genomic signatures in marine invaders.

Identification of DNA (de)methylation-related genes and their transcriptional response to environmental challenges in an invasive model ascidian.

Marine invasive species are constantly challenged by acute or recurring environmental stresses during their range expansions. DNA methylation-mediated stress memory has been proposed to effectively affect species' response and enhance their overall performance in recurring environmental challenges. In order to further test this proposal in marine invasive species, we identified genes in the DNA methylation and demethylation processes in the highly invasive model species, Ciona robusta, and subsequently investigated the expression patterns of these genes under recurring salinity stresses. After a genome-wide comprehensive survey, we found a total of six genes, including two genes of DNA methyltransferase 3a (DNMT3a1 and DNMT3a2), and one gene of DNA methyltransferase 1 (DNMT1), methyl-CpG-binding domain protein 2 (MBD2), methyl-CpG-binding domain protein 4 (MBD4) and ten-eleven-translocation protein 1 (TET1). Phylogenetic reconstruction and domain arrangement analyses showed that the deduced proteins of the identified genes were evolutionarily conserved and functionally similar with their orthologs. All genes were constitutively expressed in all four tested tissues. Interestingly, we found time-dependent and stress-specific gene expression patterns under high and low salinity stresses. Under the recurring high salinity stresses, DNMT3a1 and TET1 conformed to the definition of memory genes, while under the recurring low salinity stresses, two DNMT3a paralogues were identified as the memory genes. Altogether, our results clearly showed that the transcriptional patterns of (de)methylation-related genes were significantly influenced by environmental stresses, and the transcriptional memory of some (de)methylation-related genes should play crucial roles in DNA methylation-mediated stress memory during the process of biological invasions.

Highly dynamic transcriptional reprogramming and shorter isoform shifts under acute stresses during biological invasions.

Phenotypic plasticity has been increasingly recognized for its importance in adaptation to novel environments, and initial rapid plastic response to acute stresses usually serves as the stepping stone for future adaptation. Differential gene expression and alternative splicing have been proposed as two underlying mechanisms for rapid plastic response to environmental stresses. Here, we used an invasive model species, Ciona savignyi, to investigate the temporary plastic changes under temperature stresses on gene expression and alternative splicing. Our results revealed rapid and highly dynamic gene expression reprogramming and alternative splicing switch under acute stresses. Distinct transcriptional response profiles were triggered by two types of temperature stresses, showing resilience recovery and increasing divergence under heat and cold challenges, respectively. Interestingly, alternative exons were more inclined to be skipped under both heat and cold stresses, leading to shorter isoforms but with maintained Open Reading Frames (ORFs). Although similar response patterns were observed between differential gene expression and alternative splicing, low overlap between Differentially Expressed Genes (DEGs) and Differentially Alternative Spliced Genes (DASGs) suggests that distinct gene sets and associated functions should be involved in temperature challenges. Thus, alternative splicing should offer an additional layer of plastic response to environmental challenges. Finally, we identified key plastic genes involved in both gene expression regulation and alternative splicing. The results obtained here shed light on adaptation and accommodation mechanisms during biological invasions, particularly for acute environmental changes at early stages of biological invasions such as transport and introduction.

Stress Memory of Recurrent Environmental Challenges in Marine Invasive Species: Ciona robusta as a Case Study.

Fluctuating environmental changes impose tremendous stresses on sessile organisms in marine ecosystems, in turn, organisms develop complex response mechanisms to keep adaptive homeostasis for survival. Physiological plasticity is one of the primary lines of defense against environmental challenges, and such defense often relies on the antioxidant defense system (ADS). Hence, it is imperative to understand response mechanisms of ADS to fluctuating environments. Invasive species provide excellent models to study how species cope with environmental stresses, as invasive species encounter sudden, and often recurrent, extensive environmental challenges during the whole invasion process. Here, we studied the roles of ADS on rapid response to recurrent cold challenges in a highly invasive tunicate (Ciona robusta) by simulating cold stresses during its invasion process. We assessed antioxidative indicators, including malondialdehyde (MDA), total antioxidant capacity (T-AOC), superoxide dismutase (SOD), catalase (CAT), and glutathione (GSH), as well as transcriptional changes of ADS-related genes to reveal the physiological plasticity under recurring cold stresses. Our results demonstrated that physiological homeostasis relied on the resilience of ADS, which further accordingly tuned antioxidant activity and gene expression to changing environments. The initial cold stress remodeled baselines of ADS to promote the development of stress memory, and subsequent stress memory largely decreased the physiological response to recurrent environmental challenges. All results here suggest that C. robusta could develop stress memory to maintain physiological homeostasis in changing or harsh environments. The results obtained in this study provide new insights into the mechanism of rapid physiological adaption during biological invasions.

Formation mechanism of organo-chromium (III) complexes from bioreduction of chromium (VI) by Aeromonas hydrophila.

Chromium is a common heavy metal widely present in aquatic environments. Cost-effective remediation of chromium-contaminated environment can be realized by microbial reduction of Cr(VI) to Cr(III). The genus Aeromonas species is one of such Cr(VI) reducers, whose reduction mechanism remains unrevealed and the main factors governing the Cr(VI) reduction pathways are unknown yet. In this work, the performances and mechanisms of Cr(VI) anaerobic reduction by Aeromonas hydrophila ATCC 7966 were investigated. This strain exhibited excellent Cr(VI) resistance and could utilize a suite of electron donors to support Cr(VI) bioreduction. The Cr(VI) bioreduction processes involved both extracellular (the metal-reducing and respiratory pathway) and intracellular reaction pathways. Adding anthraquinone-2,6-disulfonate or humic acid as a mediator substantially enhanced the Cr(VI) bioreduction. The forms and distribution of the Cr(VI) bioreduction products were affected by the medium composition. Soluble organo-Cr(III) complexes were identified as the main Cr(VI) reduction products when basal salts medium was adopted. Given the environmental ubiquity of the genus Aeromonas, the findings in this work may facilitate a better understanding about the transformation behaviors and fates of Cr(VI) in environments and provide useful clues to tune the bioremediation of chromium-contaminated environments.

Genome-Wide Identification and Evaluation of New Reference Genes for Gene Expression Analysis Under Temperature and Salinity Stresses in Ciona savignyi.

Rapid adaptation/accommodation to changing environments largely contributes to maximal survival of invaders during biological invasions, usually leading to success in crossing multiple barriers and finally in varied environments in recipient habitats. Gene expression is one of the most important and rapid ways during responses to environmental stresses. Selection of proper reference genes is the crucial prerequisite for gene expression analysis using the common approach, real-time quantitative PCR (RT-qPCR). Here we identified eight candidate novel reference genes from the RNA-Seq data in an invasive model ascidian Ciona savignyi under temperature and salinity stresses. Subsequently, the expression stability of these eight novel reference genes, as well as other six traditionally used reference genes, was evaluated using RT-qPCR and comprehensive tool RefFinder. Under the temperature stress, two traditional reference genes, ribosomal proteins S15 and L17 (RPS15, RPL17), and one novel gene Ras homolog A (RhoA), were recommended as the top three stable genes, which can be used to normalize target genes with a high and moderate expression level, respectively. Under the salinity stress, transmembrane 9 superfamily member (TMN), MOB kinase activator 1A-like gene (MOB) and ubiquitin-conjugating enzyme (UBQ2) were suggested as the top three stable genes. On the other hand, several commonly used reference genes such as α-tubulin (TubA), ß-tubulin (TubB) and glyceraldehyde-3-phosphate dehydrogenase (GAPDH) showed unstable expressions, thus these genes should not be used as internal controls for gene expression analysis. We also tested the expression level of an important stress response gene, large proline-rich protein bag6-like gene (BAG) using different reference genes. As expected, we observed different results and conclusions when using different normalization methods, thus suggesting the importance of selection of proper reference genes and associated normalization methods. Our results provide a valuable reference gene resource for the normalization of gene expression in the study of environmental adaptation/accommodation during biological invasions using C. savignyi as a model.

Identification and characterization of proteins involved in stolon adhesion in the highly invasive fouling ascidian Ciona robusta.

Adhesive ascidians have caused serious biofouling problems and huge economic losses in marine ecosystems. However, adhesion mechanisms, particularly on functional proteins involved in ascidian adhesion, remain largely unexplored. Here, we identified 26 representative stolon proteins from the highly invasive fouling ascidian Ciona robusta using the proteomics approach. The uncharacterized stolon proteins were rich in adhesion-related conserved domains. Real-time quantitative PCR further revealed specific expressions of these uncharacterized protein genes in stolon tissue, suggesting their potential roles in stolon adhesion.> A recombinant vWFA domain-containing uncharacterized protein, ascidian stolon protein 1 (ASP-1), was successfully expressed in a baculovirus-insect cell system and purified in vitro. Coating experiment showed that tyrosinase-modified ASP-1 could absorb to glass and organic glass stronger than unmodified ASP-1, while only modified ASP-1 could absorb to aluminum foil. Quartz crystal microbalance analysis also showed the increase in absorption ability of ASP-1 after modification. In addition, abundant 3,4-l-dihydroxyphenylalanine (DOPA) in modified protein was detected by nitroblue tetrazolium staining. These results suggest that ASP-1 be involved in ascidian DOPA-dependent and material-selective adhesion. Overall, this study provides insight into molecular mechanisms of C. robusta stolon adhesion, and findings here are expected to be conductive to develop strategies against biofouling caused by ascidians.

Adaptive shifts of bacterioplankton communities in response to nitrogen enrichment in a highly polluted river.

Anthropogenic activity-mediated nutrient pollution, especially nitrogen enrichment, poses one of the major threats to river ecosystems. However, it remains unclear how and to which extent it affects aquatic microbial communities, especially in heavily polluted rivers. In this study, a significant environmental gradient, particularly nitrogen gradient, was observed along a wastewater receiving river, the North Canal River (NCR). The pollution level was highest, moderate, and lowest in the up-, middle, and down-streams, respectively. The community composition of bacterioplankton transitioned from being Betaproteobacteria-dominated upstream to Gammaproteobacteria-dominated downstream. Copiotrophic groups, such as Polynucleobacter (Betaproteobacteria) and Hydrogenophaga (Betaproteobacteria), were dominant in the upstream. Multiple statistical analyses indicated that total nitrogen (TN) was the most important factor driving the adaptive shifts of community structure. Analyses of co-occurrence networks showed that the complexity of networks was disrupted in the up- and middle streams, while enhanced in the downstream. Our findings here suggested that microbial interactions were reduced in response to the aggravation of nutrient pollution. Similar to these changes, we observed significant dissimilarity of composition of functional groups, with highest abundance of nitrogen metabolism members under the highest level of nitrogen enrichment. Further analyses indicated that most of these functional groups belonged to Betaproteobacteria, suggesting the potential coupling of community composition and function diversity. In summary, adaptive shifts of bacterioplankton community composition, as well as species interactions, occurred in response to nutrient pollution in highly polluted water bodies.

Methylation divergence of invasive Ciona ascidians: Significant population structure and local environmental influence.

The geographical expansion of invasive species usually leads to temporary and/or permanent changes at multiple levels (genetics, epigenetics, gene expression, etc.) to acclimatize to abiotic and/or biotic stresses in novel environments. Epigenetic variation such as DNA methylation is often involved in response to diverse local environments, thus representing one crucial mechanism to promote invasion success. However, evidence is scant on the potential role of DNA methylation variation in rapid environmental response and invasion success during biological invasions. In particular, DNA methylation patterns and possible contributions of varied environmental factors to methylation differentiation have been largely unknown in many invaders, especially for invasive species in marine systems where extremely complex interactions exist between species and surrounding environments. Using the methylation-sensitive amplification polymorphism (MSAP) technique, here we investigated population methylation structure at the genome level in two highly invasive model ascidians, Ciona robusta and C. intestinalis, collected from habitats with varied environmental factors such as temperature and salinity. We found high intrapopulation methylation diversity and significant population methylation differentiation in both species. Multiple analyses, such as variation partitioning analysis, showed that both genetic variation and environmental factors contributed to the observed DNA methylation variation. Further analyses found that 24 and 20 subepiloci were associated with temperature and/or salinity in C. robusta and C. intestinalis, respectively. All these results clearly showed significant methylation divergence among populations of both invasive ascidians, and varied local environmental factors, as well as genetic variation, were responsible for the observed DNA methylation patterns. The consistent findings in both species here suggest that DNA methylation, coupled with genetic variation, may facilitate local environmental adaptation during biological invasions, and DNA methylation variation molded by local environments may contribute to invasion success.

Genome-Wide Identification, Characterization and Expression Analyses of Heat Shock Protein-Related Genes in a Highly Invasive Ascidian Ciona savignyi.

Biological response to rapid changing environments is an outstanding research question in ecology and evolution. Biological invasions provide excellent "natural" experiments to study such a complex response process, as invaders often encounter rapidly changing environments during biological invasions. The regulation of heat shock proteins (Hsp) is a common pathway responsible for various environmental stresses; however, the comprehensive study on Hsp system across the whole genome and potential roles in determining invasion success are still largely unexplored. Here, we used a marine invasive model ascidian, Ciona savignyi, to investigate transcriptional response of Hsp-related genes to harsh environments. We identified 32 genes, including three Hsp20, six Hsp40, ten Hsp60, eight Hsp70, three Hsp90, one Hsp100, and one heat shock transcription factor (Hsf), across the whole genome of C. savignyi. We further characterized gene structure and protein motifs, and identified potential heat shock elements (HSEs) in promoters of Hsp genes. The expression analysis showed that most Hsp genes, but not all, were involved in transcriptional response to temperature and salinity challenges in a duration- and stress-specific pattern, and the maximum amplitude of induction occurred in Hsp70-4 after 1-h of high temperature treatment. However, the Hsf gene was scarcely induced and limited interactions were predicted between Hsp and Hsf genes. Our study provide the first systematic genome-wide analysis of Hsp and Hsf family in the marine invasive model ascidian, and our results are expected to dissect Hsp-based molecular mechanisms responsible for extreme environmental adaptation using Ciona as a model system.

Continuous degradation of ciprofloxacin in a manganese redox cycling system driven by Pseudomonas putida MnB-1.

Ciprofloxacin (CIP), as an extensively used antibiotic, has been widely detected at a high level in the environment and has raised environmental pollution concerns. Thus, efficient and cost-effective methods for CIP degradation are highly desired. Biologically produced manganese oxides (BioMnOx) offer a promising perspective for CIP degradation because of their catalytic reactivity and cost-effectiveness. However, the release of Mn(II) from BioMnOx prevents the further oxidation of pollutants. As a consequence, continuous CIP degradation by BioMnOx is not feasible. In this work, a manganese redox cycling system driven by Pseudomonas putida MnB-1 was constructed for continuous degradation of CIP. In such a system CIP was oxidized continuously and rapidly by re-oxidizing the formed Mn(II) to regenerate reactive BioMnOx, which also protected the strain from CIP toxicity. CIP was degraded through N-dealkylation passway. No significant loss of BioMnOx reactivity was observed in three-cycle CIP degradation process, suggesting the stability of this system. An overlooked intracellular BioMnOx, which was involved in CIP degradation, was discovered in P. putida MnB-1. Moreover, the important role of Mn(III) in facilitating CIP removal in this system was also identified. This work provides useful information to better understand the degradation of antibiotic compounds mediated by microbes in environments.

Rapid response to changing environments during biological invasions: DNA methylation perspectives.

Dissecting complex interactions between species and their environments has long been a research hot spot in the fields of ecology and evolutionary biology. The well-recognized Darwinian evolution has well-explained long-term adaptation scenarios; however, "rapid" processes of biological responses to environmental changes remain largely unexplored, particularly molecular mechanisms such as DNA methylation that have recently been proposed to play crucial roles in rapid environmental adaptation. Invasive species, which have capacities to successfully survive rapidly changing environments during biological invasions, provide great opportunities to study molecular mechanisms of rapid environmental adaptation. Here, we used the methylation-sensitive amplified polymorphism (MSAP) technique in an invasive model ascidian, Ciona savignyi, to investigate how species interact with rapidly changing environments at the whole-genome level. We detected quite rapid DNA methylation response: significant changes of DNA methylation frequency and epigenetic differentiation between treatment and control groups occurred only after 1 hr of high-temperature exposure or after 3 hr of low-salinity challenge. In addition, we detected time-dependent hemimethylation changes and increased intragroup epigenetic divergence induced by environmental stresses. Interestingly, we found evidence of DNA methylation resilience, as most stress-induced DNA methylation variation maintained shortly (~48 hr) and quickly returned back to the control levels. Our findings clearly showed that invasive species could rapidly respond to acute environmental changes through DNA methylation modifications, and rapid environmental changes left significant epigenetic signatures at the whole-genome level. All these results provide fundamental background to deeply investigate the contribution of DNA methylation mechanisms to rapid contemporary environmental adaptation.