Imperial College London

DR MICHALIS BARKOULAS

Faculty of Natural SciencesDepartment of Life Sciences

Reader in Molecular Genetics
 
 
 
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Contact

 

+44 (0)20 7594 5227m.barkoulas

 
 
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Location

 

506Sir Alexander Fleming BuildingSouth Kensington Campus

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Summary

 

Publications

Publication Type
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41 results found

Grover M, Gang SS, Troemel ER, Barkoulas Met al., 2024, Proteasome inhibition triggers tissue-specific immune responses against different pathogens in C. elegans, PLOS Biology, Vol: 22, Pages: e3002543-e3002543

<jats:p>Protein quality control pathways play important roles in resistance against pathogen infection. For example, the conserved transcription factor SKN-1/NRF up-regulates proteostasis capacity after blockade of the proteasome and also promotes resistance against bacterial infection in the nematode <jats:italic>Caenorhabditis elegans</jats:italic>. SKN-1/NRF has 3 isoforms, and the SKN-1A/NRF1 isoform, in particular, regulates proteasomal gene expression upon proteasome dysfunction as part of a conserved bounce-back response. We report here that, in contrast to the previously reported role of SKN-1 in promoting resistance against bacterial infection, loss-of-function mutants in <jats:italic>skn-1a</jats:italic> and its activating enzymes <jats:italic>ddi-1</jats:italic> and <jats:italic>png-1</jats:italic> show constitutive expression of immune response programs against natural eukaryotic pathogens of <jats:italic>C</jats:italic>. <jats:italic>elegans</jats:italic>. These programs are the oomycete recognition response (ORR), which promotes resistance against oomycetes that infect through the epidermis, and the intracellular pathogen response (IPR), which promotes resistance against intestine-infecting microsporidia. Consequently, <jats:italic>skn-1a</jats:italic> mutants show increased resistance to both oomycete and microsporidia infections. We also report that almost all ORR/IPR genes induced in common between these programs are regulated by the proteasome and interestingly, specific ORR/IPR genes can be induced in distinct tissues depending on the exact trigger. Furthermore, we show that increasing proteasome function significantly reduces oomycete-mediated induction of multiple ORR markers. Altogether, our findings demonstrate that proteasome regulation keeps innate immune responses in check in a tissue-specific manner against natural eukaryotic pathogens of the <jats

Journal article

Drury F, Grover M, Hintze M, Saunders J, Fasseas MK, Constantinou C, Barkoulas Met al., 2023, A PAX6-regulated receptor tyrosine kinase pairs with a pseudokinase to activate immune defense upon oomycete recognition in Caenorhabditis elegans., Proc Natl Acad Sci U S A, Vol: 120

Oomycetes were recently discovered as natural pathogens of Caenorhabditis elegans, and pathogen recognition alone was shown to be sufficient to activate a protective transcriptional program characterized by the expression of multiple chitinase-like (chil) genes. However, the molecular mechanisms underlying oomycete recognition in animals remain fully unknown. We performed here a forward genetic screen to uncover regulators of chil gene induction and found several independent loss-of-function alleles of old-1 and flor-1, which encode receptor tyrosine kinases belonging to the C. elegans-specific KIN-16 family. We report that OLD-1 and FLOR-1 are both necessary for mounting the immune response and act in the epidermis. FLOR-1 is a pseudokinase that acts downstream of the active kinase OLD-1 and regulates OLD-1 levels at the plasma membrane. Interestingly, the old-1 locus is adjacent to the chil genes in the C. elegans genome, thereby revealing a genetic cluster important for oomycete resistance. Furthermore, we demonstrate that old-1 expression at the anterior side of the epidermis is regulated by the VAB-3/PAX6 transcription factor, well known for its role in visual system development in other animals. Taken together, our study reveals both conserved and species-specific factors shaping the activation and spatial characteristics of the immune response to oomycete recognition.

Journal article

Drury F, Grover M, Hintze M, Saunders J, Fasseas MK, Constantinou C, Barkoulas Met al., 2022, A receptor tyrosine kinase regulated by the transcription factor VAB-3/PAX6 pairs with a pseudokinase to trigger immune signalling upon oomycete recognition in<i>C. elegans</i>

<jats:title>Summary</jats:title><jats:p>Oomycetes were recently discovered as natural pathogens of<jats:italic>Caenorhabditis elegans</jats:italic>and pathogen recognition alone was shown to be sufficient to activate a protective transcriptional program in the host characterised by the expression of multiple<jats:italic>chitinase-like</jats:italic>(<jats:italic>chil</jats:italic>) genes. However, the molecular mechanisms underlying oomycete recognition in animals remain fully unknown. We performed here a forward genetic screen to uncover regulators of<jats:italic>chil</jats:italic>gene induction and found several independent loss-of-function alleles of<jats:italic>old-1</jats:italic>and<jats:italic>flor-1,</jats:italic>which encode receptor tyrosine kinases belonging to the<jats:italic>C. elegans</jats:italic>-specific KIN-16 family. We present evidence that OLD-1 is an active kinase mounting the immune response, and FLOR-1 a pseudokinase that is also required for the response and regulates the distribution of OLD-1 at the epidermal membrane. Interestingly, the<jats:italic>old-1</jats:italic>locus is adjacent to the<jats:italic>chil</jats:italic>genes in the nematode genome, thereby revealing a genetic cluster important for oomycete-resistance. Furthermore, we identify the VAB-3/PAX-6 transcription factor known for its role in visual system development to regulate<jats:italic>old-1</jats:italic>expression, and consequently the spatial pattern of the response to oomycete recognition. Taken together, our study reveals both conserved and species-specific factors shaping the response to oomycete recognition.</jats:p>

Journal article

Gang SS, Grover M, Reddy KC, Raman D, Chang Y-T, Ekiert DC, Barkoulas M, Troemel ERet al., 2022, A pals-25 gain-of-function allele triggers systemic resistance against natural pathogens of C. elegans, PLoS Genetics, Vol: 18, ISSN: 1553-7390

Regulation of immunity throughout an organism is critical for host defense. Previous studies in the nematode Caenorhabditis elegans have described an “ON/OFF” immune switch comprised of the antagonistic paralogs PALS-25 and PALS-22, which regulate resistance against intestinal and epidermal pathogens. Here, we identify and characterize a PALS-25 gain-of-function mutant protein with a premature stop (Q293*), which we find is freed from physical repression by its negative regulator, the PALS-22 protein. PALS-25(Q293*) activates two related gene expression programs, the Oomycete Recognition Response (ORR) against natural pathogens of the epidermis, and the Intracellular Pathogen Response (IPR) against natural intracellular pathogens of the intestine. A subset of ORR/IPR genes is upregulated in pals-25(Q293*) mutants, and they are resistant to oomycete infection in the epidermis, and microsporidia and virus infection in the intestine, but without compromising growth. Surprisingly, we find that activation of PALS-25 seems to primarily stimulate the downstream bZIP transcription factor ZIP-1 in the epidermis, with upregulation of gene expression in both the epidermis and in the intestine. Interestingly, we find that PALS-22/25-regulated epidermal-to-intestinal signaling promotes resistance to the N. parisii intestinal pathogen, demonstrating cross-tissue protective immune induction from one epithelial tissue to another in C. elegans.

Journal article

Katsanos D, Barkoulas M, 2022, Targeted DamID in C. elegans reveals a direct role for LIN-22 and NHR-25 in antagonising the epidermal stem cell fate, Science Advances, Vol: 8, Pages: 1-15, ISSN: 2375-2548

Transcription factors are key players in gene networks controlling cell fate specification during development. In multicellular organisms, they often display complex patterns of expression and binding to their targets, hence tissue-specificity is required in the characterisation of transcription factor-target interactions. We introduce here Targeted DamID (TaDa) as a method for tissue-specific transcription factor target identification in intact C. elegans animals. We employ TaDa to recover targets in the epidermis for two key transcription factors, the HES1 homologue LIN-22 and the NR5A1/2 nuclear hormone receptor NHR-25. We demonstrate a direct link between LIN-22 and the Wnt signalling pathway through repression of the Frizzled receptor lin-17. We also find a direct role for NHR-25 in promoting cell differentiation via repressing the expression of stem cell-promoting GATA factors. Our results expand our understanding of the epidermal gene network and highlight the potential of TaDa to dissect the architecture of tissue-specific gene regulatory networks.

Journal article

Kalita R, Flanagan W, Lightley J, Kumar S, Alexandrov Y, Garcia E, Hintze M, Barkoulas M, Dunsby C, French PMWet al., 2021, Single-shot phase contrast microscopy using polarisation-resolved differential phase contrast, JOURNAL OF BIOPHOTONICS, Vol: 14, ISSN: 1864-063X

Journal article

Grover M, Fasseas M, Essmann C, Kenneth L, Braendle C, Felix M-A, Glockling S, Barkoulas Met al., 2021, Infection of C. elegans by Haptoglossa species reveals shared features in the host response to oomycete detection, Frontiers in Cellular and Infection Microbiology, Vol: 11, ISSN: 2235-2988

Oomycetes are a group of eukaryotic organisms that includes many important pathogens of animals and plants. Within this group, the Haptoglossa genus is characterised by the presence of specialised gun cells carrying a harpoon-like infection apparatus. While several Haptoglossa pathogens have been morphologically described, there are currently no host systems developed to study the infection process or host responses in the lab. In this study, we report that Haptoglossa species are potent natural pathogens of Caenorhabditis nematodes. Using electron microscopy, we characterise the infection process in C. elegans and demonstrate that the oomycete causes excessive tissue degradation upon entry in the body cavity, whilst leaving the host cuticle intact. We also report that the host transcriptional response to Haptoglossa infection shares similarities with the response against the oomycete Myzocytiopsis humicola, a key example of which is the induction of chitinase-like (chil) genes in the hypodermis. We demonstrate that this shared feature of the host response can be mounted by pathogen detection without any infection, as previously shown for M. humicola. These results highlight similarities in the nematode immune response to natural infection by phylogenetically distinct oomycetes.

Journal article

Katsanos D, Ferrando-Marco M, Razzaq I, Aughey G, Southall T, Barkoulas Met al., 2021, Gene expression profiling of epidermal cell types in C. elegans using Targeted-DamID., Development, Vol: 148, Pages: 1-16

The epidermis of Caenorhabditis elegans is an essential tissue for survival as it contributes to the formation of the cuticle barrier, as well as facilitates developmental progression and animal growth. Most of the epidermis consists of the hyp7 hypodermal syncytium, the nuclei of which are largely generated by the seam cells that exhibit stem cell-like behaviour during development. How the seam cell progenitors differ transcriptionally from the differentiated hypodermis is poorly understood. Here, we introduce Targeted DamID (TaDa) in C. elegans as a method for identifying genes expressed within a tissue of interest without cell isolation. We show that TaDa signal enrichment profiles can be used to identify genes transcribed in the epidermis and use this method to resolve differences in gene expression between the seam cells and the hypodermis. We finally predict and functionally validate new transcription and chromatin factors acting in seam cell development. These findings provide insights into cell-type-specific gene expression profiles likely associated with epidermal cell fate patterning.

Journal article

Koneru SL, Hintze M, Katsanos D, Barkoulas Met al., 2021, Cryptic genetic variation in a heat shock protein modifies the outcome of a mutation affecting epidermal stem cell development in <i>C. elegans</i>, NATURE COMMUNICATIONS, Vol: 12, ISSN: 2041-1723

Journal article

Hintze M, Katsanos D, Shahrezaei V, Barkoulas Met al., 2021, Phenotypic robustness of epidermal stem cell number in C. elegans Is modulated by the activity of the conserved N-acetyltransferase nath-10/NAT10, Frontiers in Cell and Developmental Biology, Vol: 9, Pages: 1-14, ISSN: 2296-634X

Individual cells and organisms experience perturbations from internal and external sources, yet manage to buffer these to produce consistent phenotypes, a property known as robustness. While phenotypic robustness has often been examined in unicellular organisms, it has not been sufficiently studied in multicellular animals. Here, we investigate phenotypic robustness in Caenorhabditis elegans seam cells. Seam cells are stem cell-like epithelial cells along the lateral edges of the animal, which go through asymmetric and symmetric divisions contributing cells to the hypodermis and neurons, while replenishing the stem cell reservoir. The terminal number of seam cells is almost invariant in the wild-type population, allowing the investigation of how developmental precision is achieved. We report here that a loss-of-function mutation in the highly conserved N-acetyltransferase nath-10/NAT10 increases seam cell number variance in the isogenic population. RNA-seq analysis revealed increased levels of mRNA transcript variability in nath-10 mutant populations, which may have an impact on the phenotypic variability observed. Furthermore, we found disruption of Wnt signaling upon perturbing nath-10 function, as evidenced by changes in POP-1/TCF nuclear distribution and ectopic activation of its GATA transcription factor target egl-18. These results highlight that NATH-10/NAT-10 can influence phenotypic variability partly through modulation of the Wnt signaling pathway.

Journal article

Koneru SL, Quah FX, Ghose R, Hintze M, Gritti N, van Zon JS, Barkoulas Met al., 2021, A role for the fusogen <i>eff-1</i> in epidermal stem cell number robustness in <i>Caenorhabditis elegans</i>, SCIENTIFIC REPORTS, Vol: 11, ISSN: 2045-2322

Journal article

Kalita R, Flanagan W, Lightley J, Kumar S, Alexandrov Y, Garcia E, Hintze M, Barkoulas M, Dunsby C, French PMWet al., 2021, Single-shot phase contrast microscopy using polarisation-resolved differential phase contrast

<jats:title>Abstract</jats:title><jats:p>We present a robust, low-cost single-shot implementation of differential phase microscopy utilising a polarisation-sensitive camera to simultaneously acquire 4 images from which the phase gradients and quantitative phase image can be calculated. This polarisation-resolved differential phase contrast (pDPC) microscopy technique can be interleaved with single-shot imaging polarimetry.</jats:p>

Journal article

Grover M, Barkoulas M, 2021, C. elegans as a new tractable host to study infections by animal pathogenic oomycetes, PLoS Pathogens, Vol: 17, Pages: 1-7, ISSN: 1553-7366

Journal article

Fasseas M, Grover M, Drury F, Essmann C, Kaulich E, Schafer W, Barkoulas Met al., 2021, Chemosensory neurons modulate the response to oomycete recognition in Caenorhabditis elegans, Cell Reports, Vol: 34, ISSN: 2211-1247

Understanding how animals detect and respond to pathogen threats is central to dissecting mechanisms of host immunity. The oomycetes represent a diverse eukaryotic group infecting various hosts from nematodes to humans. We have previously shown that Caenorhabditis elegans mounts a defense response consisting of the induction of chitinase-like (chil) genes in the epidermis to combat infection by its natural oomycete pathogen Myzocytiopsis humicola. We provide here evidence that C. elegans can sense the oomycete by detecting an innocuous extract derived from animals infected with M. humicola. The oomycete recognition response (ORR) leads to changes in the cuticle and reduction in pathogen attachment, thereby increasing animal survival. We also show that TAX-2/TAX-4 function in chemosensory neurons is required for the induction of chil-27 in the epidermis in response to extract exposure. Our findings highlight that neuron-to-epidermis communication may shape responses to oomycete recognition in animal hosts.

Journal article

Adikes RC, Kohrman AQ, Martinez MAQ, Palmisano NJ, Smith JJ, Medwig-Kinney TN, Min M, Sallee MD, Ahmed OB, Kim N, Liu S, Morabito RD, Weeks N, Zhao Q, Zhang W, Feldman JL, Barkoulas M, Pani AM, Spencer SL, Martin BL, Matus DQet al., 2020, Visualizing the metazoan proliferation-quiescence decision <i>in vivo</i>, ELIFE, Vol: 9, ISSN: 2050-084X

Journal article

Katsanos D, Barkoulas M, 2020, Targeted DamID in <i>C. elegans</i> reveals a role for LIN-22 and NHR-25 in epidermal cell differentiation

<jats:title>Abstract</jats:title><jats:p>Transcription factors are key players in gene networks controlling cell fate specification during development. In multicellular organisms, they can display complex patterns of expression and binding to their targets, which necessitates tissue-specific characterisation of transcription factor-target interactions. Here, we focus on <jats:italic>C. elegans</jats:italic> seam cell development, which is used as a model of robust epidermal stem cell patterning. Despite our knowledge of multiple transcription factors playing a role in epidermal development, the composition of the gene network underlying cell fate patterning remains largely unknown. We introduce Targeted DamID (TaDa) that allows tissue-specific transcription factor target identification in intact <jats:italic>C. elegans</jats:italic> animals without cell isolation. We employ this method to recover putative targets in the epidermis for two transcription factors, the HES1 homologue LIN-22 and the NR5A1/2 nuclear hormone receptor NHR-25. Using single-molecule FISH (smFISH), we validate TaDa predictions and reveal a role for these transcription factors in promoting cell differentiation, as well as an unusual link between a HES factor and the Wnt signalling pathway.</jats:p><jats:p>Our results expand our understanding of the epidermal gene network and highlight the power of TaDa to dissect the architecture of tissue-specific gene regulatory networks.</jats:p>

Journal article

Hintze M, Koneru SL, Gilbert SPR, Katsanos D, Lambert J, Barkoulas Met al., 2020, A Cell Fate Switch in the <i>Caenorhabditis elegans</i> Seam Cell Lineage Occurs Through Modulation of the Wnt Asymmetry Pathway in Response to Temperature Increase, GENETICS, Vol: 214, Pages: 927-939, ISSN: 0016-6731

Journal article

Reddy KC, Dror T, Underwood RS, Osman GA, Elder CR, Desjardins CA, Cuomo CA, Barkoulas M, Troemel ERet al., 2019, Antagonistic paralogs control a switch between growth and pathogen resistance in C. elegans, PLoS Pathogens, Vol: 15, Pages: 1-28, ISSN: 1553-7366

Immune genes are under intense, pathogen-induced pressure, which causes these genes to diversify over evolutionary time and become species-specific. Through a forward genetic screen we recently described a C. elegans-specific gene called pals-22 to be a repressor of “Intracellular Pathogen Response” or IPR genes. Here we describe pals-25, which, like pals-22, is a species-specific gene of unknown biochemical function. We identified pals-25 in a screen for suppression of pals-22 mutant phenotypes and found that mutations in pals-25 suppress all known phenotypes caused by mutations in pals-22. These phenotypes include increased IPR gene expression, thermotolerance, and immunity against natural pathogens, including Nematocida parisii microsporidia and the Orsay virus. Mutations in pals-25 also reverse the reduced lifespan and slowed growth of pals-22 mutants. Transcriptome analysis indicates that pals-22 and pals-25 control expression of genes induced not only by natural pathogens of the intestine, but also by natural pathogens of the epidermis. Indeed, in an independent forward genetic screen we identified pals-22 as a repressor and pals-25 as an activator of epidermal defense gene expression. In summary, the species-specific pals-22 and pals-25 genes act as a switch to regulate a program of gene expression, growth, and defense against diverse natural pathogens in C. elegans.

Journal article

Barkoulas M, Osman G, Fasseas M, Koneru S, Kyrou Ket al., 2018, Natural infection of C. elegans by an oomycete reveals a new pathogen-specific immune response, Current Biology, Vol: 28, Pages: 640-648.e5, ISSN: 1879-0445

In its natural habitat, the nematode Caenorhabditis elegans encounters a plethora of other organisms, including many that are pathogenic [ 1 ; 2]. The study of interactions between C. elegans and various pathogens has contributed to characterizing key mechanisms of innate immunity [ 2; 3 ; 4]. However, how C. elegans recognizes different pathogens to mount pathogen-specific immune responses remains still largely unknown [ 3; 5; 6; 7 ; 8]. Expanding the range of known C. elegans-infecting pathogens and characterizing novel pathogen-specific immune responses are key steps toward answering this question. We report here that the oomycete Myzocytiopsis humicola is a natural pathogen of C. elegans, and we describe its infection strategy. We identify a new host immune response to pathogen exposure that involves induction of members of a previously uncharacterized gene family encoding chitinase-like (CHIL) proteins. We demonstrate that this response is highly specific against M. humicola and antagonizes the infection. We propose that CHIL proteins may diminish the ability of the oomycete to infect by hindering pathogen attachment to the host cuticle. This work expands our knowledge of natural eukaryotic pathogens of C. elegans and introduces a new pathosystem to address how animal hosts recognize and respond to oomycete infections.

Journal article

Katsanos D, Koneru SL, Mestek Boukhibar L, Gritti N, Ghose R, Appleford PJ, Doitsidou M, Woollard A, van Zon JS, Poole RJ, Barkoulas Met al., 2017, Stochastic loss and gain of symmetric divisions in the C. elegans epidermis perturbs robustness of stem cell number., PLoS Biology, Vol: 15, ISSN: 1544-9173

Biological systems are subject to inherent stochasticity. Nevertheless, development is remarkably robust, ensuring the consistency of key phenotypic traits such as correct cell numbers in a certain tissue. It is currently unclear which genes modulate phenotypic variability, what their relationship is to core components of developmental gene networks, and what is the developmental basis of variable phenotypes. Here, we start addressing these questions using the robust number of Caenorhabditis elegans epidermal stem cells, known as seam cells, as a readout. We employ genetics, cell lineage tracing, and single molecule imaging to show that mutations in lin-22, a Hes-related basic helix-loop-helix (bHLH) transcription factor, increase seam cell number variability. We show that the increase in phenotypic variability is due to stochastic conversion of normally symmetric cell divisions to asymmetric and vice versa during development, which affect the terminal seam cell number in opposing directions. We demonstrate that LIN-22 acts within the epidermal gene network to antagonise the Wnt signalling pathway. However, lin-22 mutants exhibit cell-to-cell variability in Wnt pathway activation, which correlates with and may drive phenotypic variability. Our study demonstrates the feasibility to study phenotypic trait variance in tractable model organisms using unbiased mutagenesis screens.

Journal article

Barkoulas M, Vargas Velazquez AM, Peluffo AE, Félix MAet al., 2016, Evolution of New cis-Regulatory Motifs Required for Cell-Specific Gene Expression in Caenorhabditis, PLOS Genetics, Vol: 12, ISSN: 1553-7390

Patterning of C. elegans vulval cell fates relies on inductive signaling. In this induction event, a single cell, the gonadal anchor cell, secretes LIN-3/EGF and induces three out of six competent precursor cells to acquire a vulval fate. We previously showed that this developmental system is robust to a four-fold variation in lin-3/EGF genetic dose. Here using single-molecule FISH, we find that the mean level of expression of lin-3 in the anchor cell is remarkably conserved. No change in lin-3 expression level could be detected among C. elegans wild isolates and only a low level of change-less than 30%-in the Caenorhabditis genus and in Oscheius tipulae. In C. elegans, lin-3 expression in the anchor cell is known to require three transcription factor binding sites, specifically two E-boxes and a nuclear-hormone-receptor (NHR) binding site. Mutation of any of these three elements in C. elegans results in a dramatic decrease in lin-3 expression. Yet only a single E-box is found in the Drosophilae supergroup of Caenorhabditis species, including C. angaria, while the NHR-binding site likely only evolved at the base of the Elegans group. We find that a transgene from C. angaria bearing a single E-box is sufficient for normal expression in C. elegans. Even a short 58 bp cis-regulatory fragment from C. angaria with this single E-box is able to replace the three transcription factor binding sites at the endogenous C. elegans lin-3 locus, resulting in the wild-type expression level. Thus, regulatory evolution occurring in cis within a 58 bp lin-3 fragment, results in a strict requirement for the NHR binding site and a second E-box in C. elegans. This single-cell, single-molecule, quantitative and functional evo-devo study demonstrates that conserved expression levels can hide extensive change in cis-regulatory site requirements and highlights the evolution of new cis-regulatory elements required for cell-specific gene expression.

Journal article

Grimbert S, Tietze K, Barkoulas M, Sternberg PW, Félix MA, Braendle Cet al., 2016, Anchor cell signaling and vulval precursor cell positioning establish a reproducible spatial context during C. elegans vulval induction, Developmental Biology, Vol: 416, Pages: 123-135, ISSN: 1095-564X

How cells coordinate their spatial positioning through intercellular signaling events is poorly understood. Here we address this topic using Caenorhabditis elegans vulval patterning during which hypodermal vulval precursor cells (VPCs) adopt distinct cell fates determined by their relative positions to the gonadal anchor cell (AC). LIN-3/EGF signaling by the AC induces the central VPC, P6.p, to adopt a 1° vulval fate. Exact alignment of AC and VPCs is thus critical for correct fate patterning, yet, as we show here, the initial AC-VPC positioning is both highly variable and asymmetric among individuals, with AC and P6.p only becoming aligned at the early L3 stage. Cell ablations and mutant analysis indicate that VPCs, most prominently 1° cells, move towards the AC. We identify AC-released LIN-3/EGF as a major attractive signal, which therefore plays a dual role in vulval patterning (cell alignment and fate induction). Additionally, compromising Wnt pathway components also induces AC-VPC alignment errors, with loss of posterior Wnt signaling increasing stochastic vulval centering on P5.p. Our results illustrate how intercellular signaling reduces initial spatial variability in cell positioning to generate reproducible interactions across tissues.

Journal article

Matus DQ, Lohmer LL, Kelley LC, Schindler AJ, Kohrman AQ, Barkoulas M, Zhang W, Chi Q, Sherwood DRet al., 2015, Invasive cell fate requires G1 Cell-cycle arrest and histone deacetylase-mediated changes in gene expression, Developmental Cell, Vol: 35, Pages: 162-174, ISSN: 1534-5807

Despite critical roles in development and cancer, the mechanisms that specify invasive cellular behavior are poorly understood. Through a screen of transcription factors in Caenorhabditis elegans, we identified G1 cell-cycle arrest as a precisely regulated requirement of the anchor cell (AC) invasion program. We show that the nuclear receptor nhr-67/tlx directs the AC into G1 arrest in part through regulation of the cyclin-dependent kinase inhibitor cki-1. Loss of nhr-67 resulted in non-invasive, mitotic ACs that failed to express matrix metalloproteinases or actin regulators and lack invadopodia, F-actin-rich membrane protrusions that facilitate invasion. We further show that G1 arrest is necessary for the histone deacetylase HDA-1, a key regulator of differentiation, to promote pro-invasive gene expression and invadopodia formation. Together, these results suggest that invasive cell fate requires G1 arrest and that strategies targeting both G1-arrested and actively cycling cells may be needed to halt metastatic cancer.

Journal article

Boukhibar LM, Barkoulas M, 2015, The developmental genetics of biological robustness, Annals of Botany, Vol: 117, Pages: 699-707, ISSN: 1095-8290

Background Living organisms are continuously confronted with perturbations, such as environmental changes that include fluctuations in temperature and nutrient availability, or genetic changes such as mutations. While some developmental systems are affected by such challenges and display variation in phenotypic traits, others continue consistently to produce invariable phenotypes despite perturbation. This ability of a living system to maintain an invariable phenotype in the face of perturbations is termed developmental robustness. Biological robustness is a phenomenon observed across phyla, and studying its mechanisms is central to deciphering the genotype–phenotype relationship. Recent work in yeast, animals and plants has shown that robustness is genetically controlled and has started to reveal the underlying mechinisms behind it.Scope and Conclusions Studying biological robustness involves focusing on an important property of developmental traits, which is the phenotypic distribution within a population. This is often neglected because the vast majority of developmental biology studies instead focus on population aggregates, such as trait averages. By drawing on findings in animals and yeast, this Viewpoint considers how studies on plant developmental robustness may benefit from strict definitions of what is the developmental system of choice and what is the relevant perturbation, and also from clear distinctions between gene effects on the trait mean and the trait variance. Recent advances in quantitative developmental biology and high-throughput phenotyping now allow the design of targeted genetic screens to identify genes that amplify or restrict developmental trait variance and to study how variation propagates across different phenotypic levels in biological systems. The molecular characterization of more quantitative trait loci affecting trait variance will provide further insights into the evolution of genes modulating developmental robustness. The

Journal article

Felix M-A, Barkoulas M, 2015, Pervasive robustness in biological systems, NATURE REVIEWS GENETICS, Vol: 16, Pages: 483-496, ISSN: 1471-0056

Journal article

van Zon JS, Kienle S, Huelsz-Prince G, Barkoulas M, van Oudenaarden Aet al., 2015, Cells change their sensitivity to an EGF morphogen gradient to control EGF-induced gene expression., Nature Communications, Vol: 6, Pages: 7053-7053, ISSN: 2041-1723

How cells in developing organisms interpret the quantitative information contained in morphogen gradients is an open question. Here we address this question using a novel integrative approach that combines quantitative measurements of morphogen-induced gene expression at single-mRNA resolution with mathematical modelling of the induction process. We focus on the induction of Notch ligands by the LIN-3/EGF morphogen gradient during vulva induction in Caenorhabditis elegans. We show that LIN-3/EGF-induced Notch ligand expression is highly dynamic, exhibiting an abrupt transition from low to high expression. Similar transitions in Notch ligand expression are observed in two highly divergent wild C. elegans isolates. Mathematical modelling and experiments show that this transition is driven by a dynamic increase in the sensitivity of the induced cells to external LIN-3/EGF. Furthermore, this increase in sensitivity is independent of the presence of LIN-3/EGF. Our integrative approach might be useful to study induction by morphogen gradients in other systems.

Journal article

Hay AS, Pieper B, Cooke E, Mandakova T, Cartolano M, Tattersall AD, Ioio RD, McGowan SJ, Barkoulas M, Galinha C, Rast MI, Hofhuis H, Then C, Plieske J, Ganal M, Mott R, Martinez-Garcia JF, Carine MA, Scotland RW, Gan X, Filatov DA, Lysak MA, Tsiantis Met al., 2014, <i>Cardamine hirsuta</i>: a versatile genetic system for comparative studies, PLANT JOURNAL, Vol: 78, Pages: 1-15, ISSN: 0960-7412

Journal article

Barkoulas M, van Zon JS, Milloz J, van Oudenaarden A, Felix M-Aet al., 2013, Robustness and Epistasis in the <i>C-elegans</i> Vulval Signaling Network Revealed by Pathway Dosage Modulation, DEVELOPMENTAL CELL, Vol: 24, Pages: 64-75, ISSN: 1534-5807

Journal article

Felix M-A, Barkoulas M, 2012, Robustness and flexibility in nematode vulva development, TRENDS IN GENETICS, Vol: 28, Pages: 185-195, ISSN: 0168-9525

Journal article

Prasad K, Grigg SP, Barkoulas M, Yadav RK, Sanchez-Perez GF, Pinon V, Blilou I, Hofhuis H, Dhonukshe P, Galinha C, Mahonen AP, Muller WH, Raman S, Verkleij AJ, Snel B, Reddy GV, Tsiantis M, Scheres Bet al., 2011, <i>Arabidopsis</i> PLETHORA Transcription Factors Control Phyllotaxis, CURRENT BIOLOGY, Vol: 21, Pages: 1123-1128, ISSN: 0960-9822

Journal article

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