bioRxiv Science⌕ Search

Biology subjects

Chae, E.

Publications and source records attributed to Chae, E..

13 recordsLinked to original sources

Dynamic Filament Assembly Regulates the Prolyl Aminopeptidase Activity of Plant Immune Protein DM3

The alpha/beta hydrolase DANGEROUS MIX 3 (DM3), a proline aminopeptidase in Arabidopsis thaliana, contributes to stress resilience through both metabolic regulation and immune signaling. Its functions are partitioned within its oligomeric structure as a trimer-of-dimers, in which the immune regulatory switch resides at the dimer interface. While the enzymatic activity of DM3 is not necessary for immune response, its activity to free prolines is critical to promote tolerance to salt and drought stress. However, how this activity is regulated has remained unclear. Here, we show that DM3 undergoes dynamic and reversible assembly into higher-order filaments in a salt-sensitive manner. The cryo-electron microscopy (EM) structure of filamentous DM3 reveals that the three-stranded helical filament arises from rearrangements of a planar hexamer into a tilted hexameric configuration. Notably, within these filaments, one of the catalytic residues is flipped out to disrupt the catalytic geometry, rendering the enzyme inactive. These findings establish filament assembly as a mechanism to sequester DM3 in an inactive state and suggest that the transition between distinct oligomeric states enables DM3 to coordinate its roles in biotic and abiotic stress responses.

plant biology↗

Contrasting life history strategies explain contrasting phenology of two co-occurring bumble bee species

Across species, changing environmental conditions are altering major phenological milestones. In seasonal environments, this can involve shifts in the timing of activity onset, growth, reproduction, and senescence; however, predicting these responses can be challenging and often requires species-specific information. In this study, we investigated phenological patterns of bumble bee colony development using observations from wild nests of two common species, Bombus griseocollis and B. impatiens. Using observations of nest traffic, we documented dates of nest-searching, peak worker activity, first gyne (new queen) production, and colony senescence over three years of field data collection. We also tested whether colony growth was density dependent, because longstanding life history models have shown that colony growth patterns determine the optimal timing of reproduction in constant environments. Here, we present a novel extension of these models, using them to infer that density-independent colony growth should be associated with extended phenology in warmer years, while density-dependent should have consistent phenology. In total, we found 79 wild nests, including 34 reproductive colonies (19 B. griseocolis and 15 B. impatiens). Bombus impatiens colonies were larger, had a longer activity period, and showed density-independent growth and later reproduction in warmer years, while B. griseocollis colonies were smaller, shorter-lived, and showed density-dependent growth and similar dates of reproduction across years. Both species had higher gyne production in warmer years. These results highlight the role of life history theory for predicting interspecific variation in phenological changes, as well as a research framework for exploring possible evolutionary mismatches of existing life histories in changing environments.

ecology↗

Transcriptome Reprogramming Associated with Autoimmunity Reveals Key Points of the Growth-Defence Trade-off in Arabidopsis thaliana

Growth and defence are inversely correlated processes requiring balance for optimal fitness. While each is well studied, their regulatory trade-offs are less understood. Using DANGEROUS MIX (DM) autoimmune plants with stunted growth in Arabidopsis thaliana, we investigated the balancing act. Transcriptome analysis of three DM cases and pathogen-treated seedlings identified two major modules representing defence and growth. These core modules, comprising 4,712 genes ([~]17% of the transcriptome), were sufficient for projecting the trade-off across immune transcriptome meta-datasets. Removing all three ADR1s, the helper NLRs, reversed the expression patterns of both modules and shifted the trade-off balance. ADR1s had a stronger impact on growth-associated RLK genes than on defence genes. Autoimmune plants also showed greater chromatin accessibility changes in growth loci than in defence loci. Our findings suggest an unexpected mechanism where immune signalling, partly mediated by ADR1s, actively suppresses growth genes, providing a strategy to regulate the growth-defence trade-off.

plant biology↗

Rab2B promotes chaperonin-mediated actin folding and prevents developmental transcription reprogramming and quiescence in trypanosomes

The Rab family of small GTPases are regulators of intracellular membrane trafficking. Rab2B, a Golgi-resident Rab GTPase, has been shown to function in ER-Golgi transport and autophagosome-lysosome fusion in mammalian cells and Drosophila. In Trypanosoma brucei, the autophagy-related function of Rab2B was conserved and depletion of Rab2B blocked autophagosome-lysosome fusion. Intriguingly, depletion of Rab2B induced differentiation of T. brucei from a proliferative slender form to a quiescent stumpy form, a step crucial for parasite transmission. Chaperonin CCT/TRiC was identified as an effector of Rab2B. Depletion of CCT/TRiC subunits or actin (a known CCT/TRiC substrate), or blocking actin polymerisation with Latrunculin A all triggered differentiation, suggesting a role of Rab2B on CCT/TRiC-mediated actin folding. Using an actin chromobody, we identified a pool of nuclear actin that is crucial for transcription reprogramming and life cycle differentiation in T. brucei. The study revealed a function of Rab2B and actin in T. brucei transcription reprogramming leading to cell differentiation, and an unexpected role of Rab2B and CCT/TRiC mediated actin folding.

cell biology↗

Structure determinants of DANGEROUS MIX 3, an alpha/beta hydrolase, for triggering NLR-mediated genetic incompatibility in plants

Genetic incompatibility occurs when a mismatched pair of plant immune components mounts autoimmune responses in hybrids. Highly diversified NLR receptors are main culprits of the genetic conflict, recognizing host proteins from different origin as immune trigger. Here, we report the molecular mechanism underlying a DANGEROUS MIX (DM) autoimmunity, comprising DM2h/RPP1 NLR and its incompatible partner DM3, an alpha/beta hydrolase. Cryo-electron microscopy reveals the oligomeric nature of two natural DM3 variants in a trimer of dimer configuration. The polymorphism triggering autoimmunity is located at the dimer interface, resulting in drastic structural differences such that dimerizing helix and loop reinforcing the interface is lost and disordered. Structure-function analysis shows that integrity of the dimer interface, but neither maintenance of hexamer nor its enzymatic activity, is the key factor contributing to autoimmunity. Our finding pinpoints checkpoints embedded in the oligomeric configuration of a host enzyme that controls the switching mechanism of NLR activity.

plant biology↗

Generation of inheritable A-to-G transitions using adenine base editing and NG-PAM Cas9 in Arabidopsis thaliana

CRISPR/Cas9 technology is an important tool for functional genomics and crop improvement. It can be used to generate mutations at precise positions in the genome. Base editors consist of deaminase components and Cas9 to specify the type of mutation, such as C-to-T (cytosine base editors) or A-to-G (adenine base editors) transition. Available adenine base editor vectors usually make use of canonical Cas9, which limits their use to 5-NGG-3 containing targets. We combined a relaxed variant of SpCas9 that uses 5-NG-3 containing targets with the adenine base editor containing TadA7.10 or TadA8e to make a set of vectors. By using a phenotype-based screen, we found that our vectors efficiently induce A-to-G somatic mutations in a 5-NGG-3 PAM context in Arabidopsis thaliana up to 81% efficiency. Such mutations are inheritable at the homozygous stage in T2. Among tested vectors, pECNUS4 (Addgene #184887), which carries TadA8e, showed highest efficiency at generating a stable A-to-G transition in a 5-NGH-3 PAM context in the gene DM3. Using this vector, we were able to recreate a naturally occurring allele of DM3 in two generations without the transgene. pECNUS4 is a new component of the CRISPR toolbox to be used for introducing desired adenine base transitions with an expanded target window for functional genomic research and trait improvement.

plant biology↗

Generating minimum set of gRNA to cover multiple targets in multiple genomes with MINORg

MINORg is an offline gRNA design tool that generates the smallest possible combination of gRNA capable of covering all desired targets in multiple non-reference genomes. As interest in pangenomic research grows, so does the workload required for large screens in multiple individuals. MINORg aims to lessen this workload by capitalising on sequence homology to favour multi-target gRNA while simultaneously screening multiple genetic backgrounds in order to generate reusable gRNA panels. We demonstrated the practical application of MINORg by knocking out a 11 homologous genes tandemly arrayed in a multigene cluster in two Arabidopsis thaliana lineages using three gRNA output by MINORg. Source code is freely available at https://github.com/rlrq/MINORg.

bioinformatics↗

Genotyping-by-sequencing-based identification of Arabidopsis pattern recognition receptor RLP32 recognizing proteobacterial translation initiation factor IF1

Pattern-triggered immunity (PTI) is a central component of plant immunity. Activation of PTI relies on the recognition of microbe-derived structures, termed patterns, through plant encoded surface-resident pattern recognition receptors (PRRs). We have identified proteobacterial translation initiation factor 1 (IF1) as an immunogenic pattern that triggers PTI in Arabidopsis thaliana and some related Brassicaceae species. Unlike most other immunogenic patterns identified, IF1 elicitor activity cannot be assigned to a small peptide epitope, suggesting that tertiary fold features are required for IF1 receptor activation. We have deployed natural variation in IF1 sensitivity to identify leucine-rich repeat (LRR) receptor-like protein 32 (RLP32) as the corresponding Arabidopsis receptor using a restriction site-associated DNA sequencing (RAD-seq) approach. Transgenic expression of RLP32 confers IF1 sensitivity to rlp32 mutants, IF1-insensitive Arabidopsis accessions and IF1-insensitive Nicotiana benthamiana. RLP32 binds IF1 specifically and forms complexes with LRR receptor kinases SOBIR1 and BAK1 to mediate signaling. Similar to previously identified PRRs RLP32 confers resistance to Pseudomonas syringae infection, highlighting an unexpectedly complex array of bacterial pattern sensors within a single plant species.

plant biology↗

A singleton NLR of recent origin causes hybrid necrosis in Arabidopsis thaliana

Hybrid necrosis in plants arises from conflict between divergent alleles of immunity genes contributed by different parents, resulting in autoimmunity. We investigate a severe hybrid necrosis case in Arabidopsis thaliana, where the hybrid does not develop past the cotyledon stage and dies three weeks after sowing. Massive transcriptional changes take place in the hybrid, including the upregulation of most NLR disease resistance genes. This is due to an incompatible interaction between the singleton TIR-NLR gene DANGEROUS MIX 10 (DM10), which was recently relocated from a larger NLR cluster, and an unlinked locus, DANGEROUS MIX 11 (DM11). There are multiple DM10 allelic variants in the global A. thaliana population, several of which have premature stop codons. One of these, which has a truncated LRR domain, corresponds to the DM10 risk allele. The DM10 locus and the adjacent genomic region in the risk allele carriers are highly differentiated from those in the non-risk carriers in the global A. thaliana population, suggesting that this allele became geographically widespread only relatively recently. The DM11 risk allele is much rarer and found only in two accessions from southwestern Spain - a region from which the DM10 risk haplotype is absent - indicating that the ranges of DM10 and DM11 risk alleles may be non-overlapping.

evolutionary biology↗

RPW8/HR Repeats Predict NLR-dependent Hybrid Performance

Hybrid offspring can look very different from their parents, including having greatly increased or decreased fitness. In many plant species, conflicts between divergent elements of the immune system can cause hybrids to express autoimmunity, a generally deleterious syndrome known as hybrid necrosis. We are investigating multiple hybrid necrosis cases in Arabidopsis thaliana that are caused by allele-specific interactions between different variants at two unlinked resistance (R) gene clusters. One is the RESISTANCE TO PERONOSPORA PARASITICA 7 (RPP7) cluster, which encodes an intracellular nucleotide binding site-leucine rich repeat (NLR) immune receptors that confer strain-specific resistance to oomycetes. The other is the RESISTANCE TO POWDERY MILDEW 8 (RPW8)/HOMOLOG OF RPW8 (HR) locus, which encodes atypical resistance proteins that can confer broad-spectrum resistance to filamentous pathogens. There is extensive structural variation in the RPW8/HR cluster, both at the level of gene copy number and at the level of C-terminal protein repeats of unknown function. We demonstrate that the number of RPW8/HR repeats correlate, albeit in a complex manner, with the severity of hybrid necrosis when these alleles are combined with specific RPP7 variants. This observation suggests that gross structural differences, rather than individual amino acid polymorphisms, guide the genetic interaction between RPW8/HR and RPP7 alleles. We discuss these findings in light of the similarity of RPW8/HR proteins with pore-forming toxins, MLKL and HET-S, from mammals and fungi.

genetics↗

Modulation of ACD6 dependent hyperimmunity by natural alleles of an Arabidopsis thaliana NLR resistance gene

Plants defend themselves against pathogens by activating an array of immune responses. Unfortunately, immunity programs may also cause unintended collateral damage to the plant itself. The quantitative disease resistance gene ACCELERATED CELL DEATH 6 (ACD6) serves as a nexus for the trade-off between growth and pathogen resistance in wild populations of Arabidopsis thaliana. An autoimmune allele, ACD6-Est, first identified in the natural accession Est-1, is found in over 10% of wild strains, even though it causes a clear fitness penalty under optimal growth conditions. There is, however, extensive variation in the strength of the autoimmune phenotype expressed by strains with an ACD6-Est allele, indicative of genetic modifiers. Quantitative genetic analysis suggests that the population genetic basis of ACD6 modulation is complex, with different strains often carrying different large-effect modifiers. One modifier is SUPPRESSOR OF NPR1-1, CONSTITUTIVE 1 (SNC1), located in a highly polymorphic cluster of nucleotide-binding domain and leucine-rich repeat (NLR) immune receptor genes, which are prototypes for qualitative disease resistance genes. Allelic variation at SNC1 correlates with ACD6-Est activity in multiple accessions, and a common structural variant affecting the NL linker sequence can explain differences in SNC1 activity. Taken together, we find that an NLR gene can mask the activity of an ACD6 autoimmune allele in natural A. thaliana populations, thereby linking different arms of the plant immune system.\n\nAuthor summaryPlants defend themselves against pathogens by activating immune responses. Unfortunately, these can cause unintended collateral damage to the plant itself. Nevertheless, some wild plants have genetic variants that confer a low threshold for the activation of immunity. While these enable a plant to respond particularly quickly to pathogen attack, such variants might be potentially dangerous. We are investigating one such variant of the immune gene ACCELERATED CELL DEATH 6 (ACD6) in the plant Arabidopsis thaliana. We discovered that there are variants at other genetic loci that can mask the effects of an overly active ACD6 gene. One of these genes, SUPPRESSOR OF NPR1-1, CONSTITUTIVE 1 (SNC1), codes for a known immune receptor. The SNC1 variant that attenuates ACD6 activity is rather common in A. thaliana populations, suggesting that new combinations of the hyperactive ACD6 variant and this antagonistic SNC1 variant will often arise by natural crosses. Similarly, because the two genes are unlinked, outcrossing will often lead to the hyperactive ACD6 variants being unmasked again. We propose that allelic diversity at SNC1 contributes to the maintenance of the hyperactive ACD6 variant in natural A. thaliana populations.

plant biology↗

A rainfall-manipulation experiment with 517 Arabidopsis thaliana accessions

The gold standard for studying natural selection and adaptation in the wild is to quantify lifetime fitness of individuals from natural populations that have been grown together in a common garden, or that have been reciprocally transplanted. By combining fitness values with species traits and genome sequences, one can infer selection coefficients at the genetic level. Here we present a rainfall-manipulation experiment with 517 whole-genome sequenced natural accessions of the plant Arabidopsis thaliana spanning the global distribution of the species. The experiments were conducted in two field stations in contrasting climates, in the Mediterranean and in Central Europe, where we built rainout shelters and simulated high and low rainfall. Using custom image analysis we quantified fitness- and phenology-related traits for 23,154 pots, which contained about 14,500 plants growing independently, and over 310,000 plants growing in small populations (max. 30 plants). This large field experiment dataset, which associates fitness and ecologically-relevant traits with genomes, will provide an important resource to test eco-evolutionary genetic theories and to understand the potential evolutionary impacts of future climates on an important plant model species.

evolutionary biology↗

The genetic architecture of recurrent segregation distortion in Arabidopsis thaliana

The equal probability of transmission of alleles from either parent during sexual reproduction is a central tenet of genetics and evolutionary biology. Yet, there are many cases where this rule is violated. Such violations limit intraspecific gene flow and can facilitate the formation of genetic barriers, a first step in speciation. Biased transmission of alleles, or segregation distortion, can result from a number of biological processes including epistatic interactions between incompatible loci, gametic selection, and meiotic drive. Examples of these phenomena have been identified in many species, implying that they are universal, but comprehensive species-wide studies of segregation distortion are lacking. We have performed a species-wide screen for distorted allele frequencies in over 500 segregating populations of Arabidopsis thaliana using reduced-representation genome sequencing. Biased transmission of alleles was evident in up to a quarter of surveyed populations. Most populations exhibited distortion at only one genomic region, with some regions being repeatedly affected in multiple populations. Our results begin to elucidate the species-level architecture of biased transmission of genetic material in A. thaliana, and serve as a springboard for future studies into the basis of intraspecific genetic barriers.

plant biology↗