Historical Routes for Diversification of Domesticated Chickpea Inferred from Landrace Genomics.

According to archaeological records, chickpea (Cicer arietinum) was first domesticated in the Fertile Crescent about 10,000 years BP. Its subsequent diversification in Middle East, South Asia, Ethiopia, and the Western Mediterranean, however, remains obscure and cannot be resolved using only archeological and historical evidence. Moreover, chickpea has two market types: "desi" and "kabuli," for which the geographic origin is a matter of debate. To decipher chickpea history, we took the genetic data from 421 chickpea landraces unaffected by the green revolution and tested complex historical hypotheses of chickpea migration and admixture on two hierarchical spatial levels: within and between major regions of cultivation. For chickpea migration within regions, we developed popdisp, a Bayesian model of population dispersal from a regional representative center toward the sampling sites that considers geographical proximities between sites. This method confirmed that chickpea spreads within each geographical region along optimal geographical routes rather than by simple diffusion and estimated representative allele frequencies for each region. For chickpea migration between regions, we developed another model, migadmi, that takes allele frequencies of populations and evaluates multiple and nested admixture events. Applying this model to desi populations, we found both Indian and Middle Eastern traces in Ethiopian chickpea, suggesting the presence of a seaway from South Asia to Ethiopia. As for the origin of kabuli chickpeas, we found significant evidence for its origin from Turkey rather than Central Asia.

Transcriptomic Analysis of Flowering Time Genes in Cultivated Chickpea and Wild Cicer.

Chickpea (Cicer arietinum L.) is a major grain legume and a good source of plant-based protein. However, comprehensive knowledge of flowering time control in Cicer is lacking. In this study, we acquire high-throughput transcriptome sequencing data and analyze changes in gene expression during floral transition in the early flowering cultivar ICCV 96029, later flowering C. arietinum accessions, and two wild species, C. reticulatum and C. echinospermum. We identify Cicer orthologs of A. thaliana flowering time genes and analyze differential expression of 278 genes between four species/accessions, three tissue types, and two conditions. Our results show that the differences in gene expression between ICCV 96029 and other cultivated chickpea accessions are vernalization-dependent. In addition, we highlight the role of FTa3, an ortholog of FLOWERING LOCUS T in Arabidopsis, in the vernalization response of cultivated chickpea. A common set of differentially expressed genes was found for all comparisons between wild species and cultivars. The direction of expression change for different copies of the FT-INTERACTING PROTEIN 1 gene was variable in different comparisons, which suggests complex mechanisms of FT protein transport. Our study makes a contribution to the understanding of flowering time control in Cicer, and can provide genetic strategies to further improve this important agronomic trait.

Interplay between sex determination cascade and major signaling pathways during Drosophila eye development: Perspectives for future research.

Understanding molecular mechanisms of sexually dimorphic organ growth is a fundamental problem of developmental biology. Recent quantitative studies showed that the Drosophila compound eye is a convenient model to study the determination of the final organ size. In Drosophila, females have larger eyes than males and this is evident even after correction for the larger body size. Moreover, female eyes include more ommatidia (photosensitive units) than male eyes and this difference is specified at the third larval instar in the eye primordia called eye imaginal discs. This may result in different visual capabilities between the two sexes and have behavioral consequences. Despite growing evidence on the genetic bases of eye size variation between different Drosophila species and strains, mechanisms responsible for within-species sexual dimorphism still remain elusive. Here, we discuss a presumptive crosstalk between the sex determination cascade and major signaling pathways during dimorphic eye development. Male- and female-specific isoforms of Doublesex (Dsx) protein are known to control sex-specific differentiation in the somatic tissues. However, no data on Dsx function during eye disc growth and patterning are currently available. Remarkably, Sex lethal (Sxl), the sex determination switch protein, was shown to directly affect Hedgehog (Hh) and Notch (N) signaling in the Drosophila wing disc. The similarity of signaling pathways involved in the wing and eye disc growth suggests that Sxl might be integrated into regulation of eye development. Dsx role in the eye disc requires further investigation. We discuss currently available data on sex-biased gene expression in the Drosophila eye and highlight perspectives for future studies.

The role of AUX1 during lateral root development in the domestication of the model C4 grass Setaria italica.

C4 photosynthesis increases the efficiency of carbon fixation by spatially separating high concentrations of molecular oxygen from Rubisco. The specialized leaf anatomy required for this separation evolved independently many times. The morphology of C4 root systems is also distinctive and adapted to support high rates of photosynthesis; however, little is known about the molecular mechanisms that have driven the evolution of C4 root system architecture. Using a mutant screen in the C4 model plant Setaria italica, we identify Siaux1-1 and Siaux1-2 as root system architecture mutants. Unlike in S. viridis, AUX1 promotes lateral root development in S. italica. A cell by cell analysis of the Siaux1-1 root apical meristem revealed changes in the distribution of cell volumes in all cell layers and a dependence of the frequency of protophloem and protoxylem strands on SiAUX1. We explore the molecular basis of the role of SiAUX1 in seedling development using an RNAseq analysis of wild-type and Siaux1-1 plants and present novel targets for SiAUX1-dependent gene regulation. Using a selection sweep and haplotype analysis of SiAUX1, we show that Hap-2412TT in the promoter region of SiAUX1 is an allele which is associated with lateral root number and has been strongly selected for during Setaria domestication.

Mechanisms of Vernalization-Induced Flowering in Legumes.

Vernalization is the requirement for exposure to low temperatures to trigger flowering. The best knowledge about the mechanisms of vernalization response has been accumulated for Arabidopsis and cereals. In Arabidopsis thaliana, vernalization involves an epigenetic silencing of the MADS-box gene FLOWERING LOCUS C (FLC), which is a flowering repressor. FLC silencing releases the expression of the main flowering inductor FLOWERING LOCUS T (FT), resulting in a floral transition. Remarkably, no FLC homologues have been identified in the vernalization-responsive legumes, and the mechanisms of cold-mediated transition to flowering in these species remain elusive. Nevertheless, legume FT genes have been shown to retain the function of the main vernalization signal integrators. Unlike Arabidopsis, legumes have three subclades of FT genes, which demonstrate distinct patterns of regulation with respect to environmental cues and tissue specificity. This implies complex mechanisms of vernalization signal propagation in the flowering network, that remain largely elusive. Here, for the first time, we summarize the available information on the genetic basis of cold-induced flowering in legumes with a special focus on the role of FT genes.

Quality Control of Human Pluripotent Stem Cell Colonies by Computational Image Analysis Using Convolutional Neural Networks.

Human pluripotent stem cells are promising for a wide range of research and therapeutic purposes. Their maintenance in culture requires the deep control of their pluripotent and clonal status. A non-invasive method for such control involves day-to-day observation of the morphological changes, along with imaging colonies, with the subsequent automatic assessment of colony phenotype using image analysis by machine learning methods. We developed a classifier using a convolutional neural network and applied it to discriminate between images of human embryonic stem cell (hESC) colonies with "good" and "bad" morphological phenotypes associated with a high and low potential for pluripotency and clonality maintenance, respectively. The training dataset included the phase-contrast images of hESC line H9, in which the morphological phenotype of each colony was assessed through visual analysis. The classifier showed a high level of accuracy (89%) in phenotype prediction. By training the classifier on cropped images of various sizes, we showed that the spatial scale of ~144 µm was the most informative in terms of classification quality, which was an intermediate size between the characteristic diameters of a single cell (~15 µm) and the entire colony (~540 µm). We additionally performed a proteomic analysis of several H9 cell samples used in the computational analysis and showed that cells of different phenotypes differentiated at the molecular level. Our results indicated that the proposed approach could be used as an effective method of non-invasive automated analysis to identify undesirable developmental anomalies during the propagation of pluripotent stem cells.

Genetic Determinants of Fiber-Associated Traits in Flax Identified by Omics Data Integration.

In this paper, we explore potential genetic factors in control of flax phenotypes associated with fiber by mining a collection of 306 flax accessions from the Federal Research Centre of the Bast Fiber Crops, Torzhok, Russia. In total, 11 traits were assessed in the course of 3 successive years. A genome-wide association study was performed for each phenotype independently using six different single-locus models implemented in the GAPIT3 R package. Moreover, we applied a multivariate linear mixed model implemented in the GEMMA package to account for trait correlations and potential pleiotropic effects of polymorphisms. The analyses revealed a number of genomic variants associated with different fiber traits, implying the complex and polygenic control. All stable variants demonstrate a statistically significant allelic effect across all 3 years of the experiment. We tested the validity of the predicted variants using gene expression data available for the flax fiber studies. The results shed new light on the processes and pathways associated with the complex fiber traits, while the pinpointed candidate genes may be further used for marker-assisted selection.

Genomic Regions Associated with Fusarium Wilt Resistance in Flax.

Modern flax cultivars are susceptible to many diseases; arguably, the most economically damaging of these is the Fusarium wilt fungal disease. Over the past decades international flax breeding initiatives resulted in the development of resistant cultivars. However, much remains to be learned about the mechanisms of resistance to Fusarium infection in flax. As a first step to uncover the genetic factors associated with resistance to Fusarium wilt disease, we performed a genome-wide association study (GWAS) using 297 accessions from the collection of the Federal Research Centre of the Bast Fiber Crops, Torzhok, Russia. These genotypes were infected with a highly pathogenic Fusarium oxysporum f.sp. lini MI39 strain; the wilt symptoms were documented in the course of three successive years. Six different single-locus models implemented in GAPIT3 R package were applied to a selected subset of 72,526 SNPs. A total of 15 QTNs (Quantitative Trait Nucleotides) were detected during at least two years of observation, while eight QTNs were found during all three years of the experiment. Of these, ten QTNs occupied a region of 640 Kb at the start of chromosome 1, while the remaining QTNs mapped to chromosomes 8, 11 and 13. All stable QTNs demonstrate a statistically significant allelic effect across 3 years of the experiment. Importantly, several QTNs spanned regions that harbored genes involved in the pathogen recognition and plant immunity response, including the KIP1-like protein (Lus10025717) and NBS-LRR protein (Lus10025852). Our results provide novel insights into the genetic architecture of flax resistance to Fusarium wilt and pinpoint potential candidate genes for further in-depth studies.

A Genomic Blueprint of Flax Fungal Parasite Fusarium oxysporum f. sp. lini.

Fusarium wilt of flax is an aggressive disease caused by the soil-borne fungal pathogen Fusarium oxysporum f. sp. lini. It is a challenging pathogen presenting a constant threat to flax production industry worldwide. Previously, we reported chromosome-level assemblies of 5 highly pathogenic F. oxysporum f. sp. lini strains. We sought to characterize the genomic architecture of the fungus and outline evolutionary mechanisms shaping the pathogen genome. Here, we reveal the complex multi-compartmentalized genome organization and uncover its diverse evolutionary dynamics, which boosts genetic diversity and facilitates host adaptation. In addition, our results suggest that host of functions implicated in the life cycle of mobile genetic elements are main contributors to dissimilarity between proteomes of different Fusaria. Finally, our experiments demonstrate that mobile genetics elements are expressed in planta upon infection, alluding to their role in pathogenicity. On the whole, these results pave the way for further in-depth studies of evolutionary forces shaping the host-pathogen interaction.

Quantitative analysis reveals genotype- and domain- specific differences between mRNA and protein expression of segmentation genes in Drosophila.

In many biological systems gene expression at mRNA and protein levels is not identical. Rigorous comparison of such differences on a spatio-temporal scale is still not feasible by high-throughput transcriptomic and proteomic analyses of early embryo development. Here, we characterize differences between mRNA and protein expression of Drosophila segmentation genes at the level of individual gene expression domains. We obtained quantitative imaging data on expression of gap genes gt and hb and pair-rule gene eve for Drosophila wild type embryos, Kr null mutants and Kr+/Kr- heterozygotes. To compare mRNA and protein expression we use several criteria including difference in amplitude and positions of expression domains, pattern shape and positional variability. For a number of gene expression domains we show examples where protein expression does not repeat mRNA expression even after a temporal delay. We calculated time delays between eve pattern formation at the level of mRNA and protein for wild type embryos, Kr mutants and Kr+/Kr- heterozygotes. We detect that in wild type embryos, the amplitudes of eve stripes 3 and 7 do not differ significantly at the level of mRNA, however, stripe 3 is higher than stripe 7 at the protein level. We further show that hb mRNA and protein expression in both anterior and posterior domains significantly differs at specific time points. The formation of hb PS4 stripe at the mRNA level proceeds five times faster than at the level of protein. With regard to spatial expression, we show that the offset between posterior gt mRNA and protein domains is much larger in Kr mutants than in wild type embryos and heterozygotes. Finally, we analyze differences in positional variability of eve stripe 7 expression in Kr mutants and Kr+/Kr- heterozygotes at the level of mRNA and protein. These results enable further perspectives to uncover mechanisms underlying discrepancies between mRNA and protein expression in early embryo.

The Genome Sequence of Five Highly Pathogenic Isolates of Fusarium oxysporum f. sp. lini.

Fusarium wilt is the most destructive fungal disease in flax, limiting flax cultivation in all the main flax and linseed growing countries. The causative agent is seedborne and soilborne fungus F. oxysporum f. sp. lini. Here, we report, for the first time, genome assemblies of five highly pathogenic isolates of Fusarium oxysporum f. sp. lini, namely monoisolate 39 and strains F329, F324, F282, F287. In addition, syntenic analysis provided a powerful approach to distinguish between core and lineage-specific parts of the genome. These results lay a solid foundation for comparative genomics studies of plant fungal pathogens, evolution of pathogenicity, and virulence factors underlying the dynamics of host-pathogen interactions, thus eventually offering solutions to Fusarium disease control.

Multi-trait multi-locus SEM model discriminates SNPs of different effects.

BACKGROUND: There is a plethora of methods for genome-wide association studies. However, only a few of them may be classified as multi-trait and multi-locus, i.e. consider the influence of multiple genetic variants to several correlated phenotypes. RESULTS: We propose a multi-trait multi-locus model which employs structural equation modeling (SEM) to describe complex associations between SNPs and traits - multi-trait multi-locus SEM (mtmlSEM). The structure of our model makes it possible to discriminate pleiotropic and single-trait SNPs of direct and indirect effect. We also propose an automatic procedure to construct the model using factor analysis and the maximum likelihood method. For estimating a large number of parameters in the model, we performed Bayesian inference and implemented Gibbs sampling. An important feature of the model is that it correctly copes with non-normally distributed variables, such as some traits and variants. CONCLUSIONS: We applied the model to Vavilov's collection of 404 chickpea (Cicer arietinum L.) accessions with 20-fold cross-validation. We analyzed 16 phenotypic traits which we organized into five groups and found around 230 SNPs associated with traits, 60 of which were of pleiotropic effect. The model demonstrated high accuracy in predicting trait values.

Dynamical climatic model for time to flowering in Vigna radiata.

BACKGROUND: Phenology data collected recently for about 300 accessions of Vigna radiata (mungbean) is an invaluable resource for investigation of impacts of climatic factors on plant development. RESULTS: We developed a new mathematical model that describes the dynamic control of time to flowering by daily values of maximal and minimal temperature, precipitation, day length and solar radiation. We obtained model parameters by adaptation to the available experimental data. The models were validated by cross-validation and used to demonstrate that the phenology of adaptive traits, like flowering time, is strongly predicted not only by local environmental factors but also by plant geographic origin and genotype. CONCLUSIONS: Of local environmental factors maximal temperature appeared to be the most critical factor determining how faithfully the model describes the data. The models were applied to forecast time to flowering of accessions grown in Taiwan in future years 2020-2030.

Genome-wide association study in accessions of the mini-core collection of mungbean (Vigna radiata) from the World Vegetable Gene Bank (Taiwan).

BACKGROUND: Mungbean (Vigna radiata (L.) R. Wilczek, or green gram) is important tropical and sub-tropical legume and a rich source of dietary protein and micronutrients. In this study we employ GWAS to examine the genetic basis of variation in several important traits in mungbean, using the mini-core collection established by the World Vegetable Center, which includes 296 accessions that represent the major market classes. This collection has been grown in a common field plot in southern European part of Russia in 2018. RESULTS: We used 5041 SNPs in 293 accessions that passed strict filtering for genetic diversity, linkage disequilibrium, population structure and GWAS analysis. Polymorphisms were distributed among all chromosomes, but with variable density. Linkage disequilibrium decayed in approximately 105 kb. Four distinct subgroups were identified within 293 accessions with 70% of accessions attributed to one of the four populations. By performing GWAS on the mini-core collection we have found several loci significantly associated with two important agronomical traits. Four SNPs associated with possibility of maturation in Kuban territory of Southern Russia in 2018 were identified within a region of strong linkage which contains genes encoding zinc finger A20 and an AN1 domain stress-associated protein. CONCLUSIONS: The core collection of mungbean established by the World Vegetable Center is a valuable resource for mungbean breeding. The collection has been grown in southern European part of Russia in 2018 under incidental stresses caused by abnormally hot weather and different photoperiod. We have found several loci significantly associated with color of hypocotyl and possibility of maturation under these stressful conditions. SNPs associated with possibility of maturation localize to a region on chromosome 2 with strong linkage, in which genes encoding zinc finger A20 and AN1 domain stress associated protein (SAP) are located. Phenotyping of WorldVeg collection for maturation traits in temperate climatic locations is important as phenology remains a critical breeding target for mungbean. As demand rises for mungbean, production in temperate regions with shorter growing seasons becomes crucial to keep up with needs. Uncovering SNPs for phenology traits will speed breeding efforts.

Genomic Analysis of Vavilov's Historic Chickpea Landraces Reveals Footprints of Environmental and Human Selection.

A defining challenge of the 21st century is meeting the nutritional demands of the growing human population, under a scenario of limited land and water resources and under the specter of climate change. The Vavilov seed bank contains numerous landraces collected nearly a hundred years ago, and thus may contain 'genetic gems' with the potential to enhance modern breeding efforts. Here, we analyze 407 landraces, sampled from major historic centers of chickpea cultivation and secondary diversification. Genome-Wide Association Studies (GWAS) conducted on both phenotypic traits and bioclimatic variables at landraces sampling sites as extended phenotypes resulted in 84 GWAS hits associated to various regions. The novel haploblock-based test identified haploblocks enriched for single nucleotide polymorphisms (SNPs) associated with phenotypes and bioclimatic variables. Subsequent bi-clustering of traits sharing enriched haploblocks underscored both non-random distribution of SNPs among several haploblocks and their association with multiple traits. We hypothesize that these clusters of pleiotropic SNPs represent co-adapted genetic complexes to a range of environmental conditions that chickpea experienced during domestication and subsequent geographic radiation. Linking genetic variation to phenotypic data and a wealth of historic information preserved in historic seed banks are the keys for genome-based and environment-informed breeding intensification.

Non-linear regression models for time to flowering in wild chickpea combine genetic and climatic factors.

BACKGROUND: Accurate prediction of crop flowering time is required for reaching maximal farm efficiency. Several models developed to accomplish this goal are based on deep knowledge of plant phenology, requiring large investment for every individual crop or new variety. Mathematical modeling can be used to make better use of more shallow data and to extract information from it with higher efficiency. Cultivars of chickpea, Cicer arietanum, are currently being improved by introgressing wild C. reticulatum biodiversity with very different flowering time requirements. More understanding is required for how flowering time will depend on environmental conditions in these cultivars developed by introgression of wild alleles. RESULTS: We built a novel model for flowering time of wild chickpeas collected at 21 different sites in Turkey and grown in 4 distinct environmental conditions over several different years and seasons. We propose a general approach, in which the analytic forms of dependence of flowering time on climatic parameters, their regression coefficients, and a set of predictors are inferred automatically by stochastic minimization of the deviation of the model output from data. By using a combination of Grammatical Evolution and Differential Evolution Entirely Parallel method, we have identified a model that reflects the influence of effects of day length, temperature, humidity and precipitation and has a coefficient of determination of R2=0.97. CONCLUSIONS: We used our model to test two important hypotheses. We propose that chickpea phenology may be strongly predicted by accession geographic origin, as well as local environmental conditions at the site of growth. Indeed, the site of origin-by-growth environment interaction accounts for about 14.7% of variation in time period from sowing to flowering. Secondly, as the adaptation to specific environments is blueprinted in genomes, the effects of genes on flowering time may be conditioned on environmental factors. Genotype-by-environment interaction accounts for about 17.2% of overall variation in flowering time. We also identified several genomic markers associated with different reactions to climatic factor changes. Our methodology is general and can be further applied to extend existing crop models, especially when phenological information is limited.

In silico evolution of the Drosophila gap gene regulatory sequence under elevated mutational pressure.

BACKGROUND: Cis-regulatory sequences are often composed of many low-affinity transcription factor binding sites (TFBSs). Determining the evolutionary and functional importance of regulatory sequence composition is impeded without a detailed knowledge of the genotype-phenotype map. RESULTS: We simulate the evolution of regulatory sequences involved in Drosophila melanogaster embryo segmentation during early development. Natural selection evaluates gene expression dynamics produced by a computational model of the developmental network. We observe a dramatic decrease in the total number of transcription factor binding sites through the course of evolution. Despite a decrease in average sequence binding energies through time, the regulatory sequences tend towards organisations containing increased high affinity transcription factor binding sites. Additionally, the binding energies of separate sequence segments demonstrate ubiquitous mutual correlations through time. Fewer than 10% of initial TFBSs are maintained throughout the entire simulation, deemed 'core' sites. These sites have increased functional importance as assessed under wild-type conditions and their binding energy distributions are highly conserved. Furthermore, TFBSs within close proximity of core sites exhibit increased longevity, reflecting functional regulatory interactions with core sites. CONCLUSION: In response to elevated mutational pressure, evolution tends to sample regulatory sequence organisations with fewer, albeit on average, stronger functional transcription factor binding sites. These organisations are also shaped by the regulatory interactions among core binding sites with sites in their local vicinity.

Analysis of functional importance of binding sites in the Drosophila gap gene network model.

BACKGROUND: The statistical thermodynamics based approach provides a promising framework for construction of the genotype-phenotype map in many biological systems. Among important aspects of a good model connecting the DNA sequence information with that of a molecular phenotype (gene expression) is the selection of regulatory interactions and relevant transcription factor bindings sites. As the model may predict different levels of the functional importance of specific binding sites in different genomic and regulatory contexts, it is essential to formulate and study such models under different modeling assumptions. RESULTS: We elaborate a two-layer model for the Drosophila gap gene network and include in the model a combined set of transcription factor binding sites and concentration dependent regulatory interaction between gap genes hunchback and Kruppel. We show that the new variants of the model are more consistent in terms of gene expression predictions for various genetic constructs in comparison to previous work. We quantify the functional importance of binding sites by calculating their impact on gene expression in the model and calculate how these impacts correlate across all sites under different modeling assumptions. CONCLUSIONS: The assumption about the dual interaction between hb and Kr leads to the most consistent modeling results, but, on the other hand, may obscure existence of indirect interactions between binding sites in regulatory regions of distinct genes. The analysis confirms the previously formulated regulation concept of many weak binding sites working in concert. The model predicts a more or less uniform distribution of functionally important binding sites over the sets of experimentally characterized regulatory modules and other open chromatin domains.

Quantitative dynamics and increased variability of segmentation gene expression in the Drosophila Krüppel and knirps mutants.

Here we characterize the response of the Drosophila segmentation system to mutations in two gap genes, Kr and kni, in the form of single or double homozygotes and single heterozygotes. Segmentation gene expression in these genotypes was quantitatively monitored with cellular resolution in space and 6.5 to 13min resolution in time. As is the case with wild type, we found that gene expression domains in the posterior portion of the embryo shift to the anterior over time. In certain cases, such as the gt posterior domain in Kr mutants, the shifts are significantly larger than is seen in wild type embryos. We also investigated the effects of Kr and kni on the variability of gene expression. Mutations often produce variable phenotypes, and it is well known that the cuticular phenotype of Kr mutants is variable. We sought to understand the molecular basis of this effect. We find that throughout cycle 14A the relative levels of eve and ftz expression in stripes 2 and 3 are variable among individual embryos. Moreover, in Kr and kni mutants, unlike wild type, the variability in positioning of the posterior Hb domain and eve stripe 7 is not decreased or filtered with time. The posterior Gt domain in Kr mutants is highly variable at early times, but this variability decreases when this domain shifts in the anterior direction to the position of the neighboring Kni domain. In contrast to these findings, positional variability throughout the embryo does not decrease over time in double Kr;kni mutants. In heterozygotes the early expression patterns of segmentation genes resemble patterns seen in homozygous mutants but by the onset of gastrulation they become similar to the wild type patterns. Finally, we note that gene expression levels are reduced in Kr and kni mutant embryos and have a tendency to decrease over time. This is a surprising result in view of the role that mutual repression is thought to play in the gap gene system.

Lack of tailless leads to an increase in expression variability in Drosophila embryos.

Developmental processes are robust, or canalised: dynamic patterns of gene expression across space and time are regulated reliably and precisely in the presence of genetic and environmental perturbations. It remains unclear whether canalisation relies on specific regulatory factors (such as heat-shock proteins), or whether it is based on more general redundancy and distributed robustness at the network level. The latter explanation implies that mutations in many regulatory factors should exhibit loss of canalisation. Here, we present a quantitative characterisation of segmentation gene expression patterns in mutants of the terminal gap gene tailless (tll) in Drosophila melanogaster. Our analysis provides new insights into the dynamic mechanisms underlying gap gene regulation, and reveals significantly increased variability of gene expression in the mutant compared to the wild-type background. We show that both position and timing of posterior segmentation gene expression domains vary strongly from embryo-to-embryo in tll mutants. This variability must be caused by a vulnerability in the regulatory system which is hidden or buffered in the wild-type, but becomes uncovered by the deletion of tll. Our analysis provides evidence that loss of canalisation in mutants could be more widespread than previously thought.