Bistability and hysteresis of the 'Secteur' differentiation are controlled by a two-gene locus in Nectria haematococca

BACKGROUND: Bistability and hysteresis are increasingly recognized as major properties of regulatory networks governing numerous biological phenomena, such as differentiation and cell cycle progression. The full scope of the underlying molecular mechanisms leading to bistability and hysteresis remains elusive. Nectria haemaotcocca, a saprophytic or pathogenic fungus with sexual reproduction, exhibits a bistable morphological modification characterized by a reduced growth rate and an intense pigmentation. Bistability is triggered by the presence or absence of σ, a cytoplasmic determinant. This determinant spreads in an infectious manner in the hyphae of the growing margin, insuring hysteresis of the differentiation. RESULTS: Seven mutants specifically affected in the generation of σ were selected through two different screening strategies. The s(1 )and s(2 )mutations completely abolish the generation of σ and of its morphological expression, the Secteur. The remaining five mutations promote its constitutive generation, which determines an intense pigmentation but not growth alteration. The seven mutations map at the same locus, Ses (for 'Secteur-specific'). The s(2 )mutant was obtained by an insertional mutagenesis strategy, which permitted the cloning of the Ses locus. Sequence and transcription analysis reveals that Ses is composed of two closely linked genes, SesA, mutated in the s(1 )and s(2 )mutant strains, and SesB, mutated in the s* mutant strains. SesB shares sequence similarity with animal and fungal putative proteins, with potential esterase/lipase/thioesterase activity, whereas SesA is similar to proteins of unknown function present only in the filamentous fungi Fusarium graminearum and Podospora anserina. CONCLUSIONS: The cloning of Ses provides evidence that a system encoded by two linked genes directs a bistable and hysteretic switch in a eukaryote. Atypical regulatory relations between the two proteins may account for the hysteresis of Secteur differentiation

Recovery of mutants impaired in pathogenicity after transposition of Impala in Fusarium oxysporum f. sp. melonis

The ability of transposon impala to inactivate genes involved in pathogenicity was tested in Fusarium oxysporum f. sp. melonis. Somatic excision of an impala copy inserted in the nitrate reductase-encoding niaD gene was positively selected through a phenotypic assay based on the restoration of nitrate reductase activity. Independent excision events were analyzed molecularly and shown to carry reinsertedimpala in more than 70% of the cases. Mapping of reinserted impala elements on large NotI-restriction fragments showed that impala transposes randomly. By screening 746 revertants on plants, a high proportion (3.5%) of mutants impaired in their pathogenic potential was recovered. According to the kinetics of wilt symptom development, the strains that were impaired in pathogenicity were clustered in three classes: class 1 grouped two strains that never induced Fusarium wilt symptoms on the host plant; class 2 and class 3 grouped 15 and 9 revertants which caused symptoms more than 50 and 30 days after inoculation, respectively. The first results demonstrate the efficiency of transposition in generating mutants affected in pathogenicity, which are usually difficult to obtain by classical mutagenesis, and open the possibility to clone the altered genes with impala as a tag

IMPACT DES ELEMENTS TRANSPOSABLES SUR L'EVOLUTION DU GENOME DU CHAMPIGNON PHYTOPATHOGENE FUSARIUM OXYSPORUM

LE GENOME DU CHAMPIGNON PHYTOPATHOGENE FUSARIUM OXYSPORUM CONTIENT PLUSIEURS FAMILLES D'ELEMENTS TRANSPOSABLES (ETS) REPRESENTATIVES DES CLASSES MAJEURS IDENTIFIEES CHEZ LES EUCARYOTES. CES ELEMENTS ONT LA POTENTIALITE DE JOUER UN ROLE IMPORTANT DANS LE REMODELAGE DU GENOME ET DANS LA CREATION DE LA DIVERSITE GENETIQUE, TANT PAR LES MUTATIONS ASSOCIEES A LEUR INSERTION ET LEUR EXCISION QU'A LEUR CAPACITE A ENGENDRER DES REMANIEMENTS GENETIQUES ET LEUR APTITUDE A ENVAHIR DE NOUVEAUX GENOMES. C'EST DANS CETTE PROBLEMATIQUE QUE S'INSCRIT MON TRAVAIL DE THESE. LA PREMIERE PARTIE DE MON TRAVAIL A CONSISTE A DETERMINER LES IMPACTS BIOLOGIQUES DES ETS SUR LA STRUCTURE, L'ORGANISATION ET LA PLASTICITE DU GENOME AU SEIN DE CET ORGANISME. TOUT D'ABORD J'AI RECHERCHE SI DES REMANIEMENTS CHROMOSOMIQUES POUVAIENT ETRE VISUALISES EN COMPARANT DES ELECTROCARYOTYPES DE SOUCHES DANS LESQUELLES DES EVENEMENTS DE TRANSPOSITION AVAIENT ETE SELECTIONNES. CETTE ETUDE A REVELE UNE VARIATION CARYOTYPIQUE D'UNE AMPLEUR EXCEPTIONNELLE. A L'AIDE DE MARQUEURS SPECIFIQUES DE CHACUN DES CHROMOSOMES, J'AI CARACTERISE DIFFERENTS TYPES DE REARRANGEMENTS CHROMOSOMIQUES (TRANSLOCATIONS, DELETIONS, DUPLICATIONS). AFIN D'IDENTIFIER LES MECANISMES A L'ORIGINE DE CES REARRANGEMENTS, J'AI SUIVI LE DEVENIR DE QUELQUES CHROMOSOMES, SIEGES DE TRANSPOSITION ET ANALYSE LA DISTRIBUTION DE DIFFERENTES FAMILLES D'ETS SUR LES CHROMOSOMES. J'AI AINSI PU MONTRER QUE (I) LA TRANSPOSITION PER SE NE SEMBLE PAS ENGENDRER DE REMANIEMENTS (II) CERTAINES FAMILLES D'ETS NE SONT PAS DISTRIBUEES DE FACON ALEATOIRE MAIS SONT CONCENTREES DANS CERTAINES REGIONS GENOMIQUES, (III) IL EXISTE UNE CORRELATION ENTRE LE NIVEAU ELEVE DU POLYMORPHISME DE CERTAINS CHROMOSOMES ET LA DENSITE EN ETS. CES RESULTATS SUGGERENT QUE LA VARIATION CARYOTYPIQUE OBSERVEE RESULTERAIT PLUTOT DE RECOMBINAISONS ECTOPIQUES ENTRE DES COPIES DISPERSEES D'ETS. LE SECOND VOLET DE MON TRAVAIL A PORTE SUR LA DYNAMIQUE DE L'ELEMENT FOT1 DANS LES POPULATIONS NATURELLES DE L'ESPECE F. OXYSPORUM MAIS AUSSI DANS LE GENRE FUSARIUM. PAR LA COMBINAISON DE TROIS TECHNIQUES, SOUTHERN BLOT, PCR ET SEQUENCAGE, J'AI ANALYSE LA DISTRIBUTION DE CET ELEMENT ET MESURE LE POLYMORPHISME INTRA ET INTERSPECIFIQUE. LES RENSEIGNEMENTS OBTENUS ONT REVELE QUE LA PRESENCE ET L'EVOLUTION DU NOMBRE DE COPIES DANS LES SOUCHES ETAIENT SOUS LA DEPENDANCE DE PLUSIEURS PROCESSUS : INACTIVATION EPIGENETIQUE, PERTE STOCHASTIQUE PAR DELETION ET REACQUISITION PAR TRANSFERT HORIZONTAL. ENFIN DANS LE CADRE DU DEVELOPPEMENT D'UN OUTIL DE CLONAGE BASE SUR L'UTILISATION DES TRANSPOSONS, JE ME SUIS INTERESSE A DETERMINER LE PATRON DE TRANSPOSITION. J'AI MONTRE QUE LES ELEMENTS FOT1 ET IMPALA TRANSPOSAIENT SUR DIFFERENTS CHROMOSOMES ET DANS DES FRAGMENTS GENOMIQUES DIFFERENTS SUGGERANT QU'ILS SONT CAPABLES DE S'INSERER DE FACON ALEATOIRE DANS LE GENOME. L'ENSEMBLE DES RESULTATS PRESENTES APPORTENT UNE VISION DU GENOME EXTREMEMENT FLUIDE CHEZ CET ORGANISME. LA RICHESSE EN ELEMENTS TRANSPOSABLES CONSTITUE UN PUISSANT GENERATEUR DE VARIABILITE ET PAR CONSEQUENT UN MOTEUR DANS L'EVOLUTION DE CE PATHOGENE DEPOURVU DE REPRODUCTION SEXUEE.ORSAY-PARIS 11-BU Sciences (914712101) / SudocSudocFranceF

Impala, a transposon from Fusarium oxysporum, is active in the genome of Penicillium griseoroseum

An autonomous impala transposon trapped in Fusarium oxysporum by insertion within the niaD gene encoding nitrate reductase was introduced in the genome of the fungus Penicillium griseoroseum, a producer of pectinase enzymes. Through a phenotypic assay, we demonstrate that this element is able to excise from the niaD gene and to reinsert at new genomic positions. As in the original host, impala inserts into a TA site and footprints left by impala excisions are generally 5 bp. The fact that impala is able to transpose in P. griseoroseum offers the opportunity to develop a gene-tagging system based on this element with the objective to detect and clone genes related in pectinase production

Production and characterization of pathogenicy-deficient mutants of Fusarium oxysporum f. sp. melonis

International audienc

Tagging pathogenicity genes in Fusarium graminearum using the transposon system mimp/impala

Transposon mutagenesis was applied to generate mutants in Fusarium graminearum . The mimp1/impala system originally identified in F. oxysporum proved very promising for mutagenesis as the transposon and reinserted at high frequency in (the vicinity) of genes. A collection of mutants was screened for growth, for pathogenicity and for perithecia production. Several mutants blocked in one or more functions were obtained. The wild-type phenotype of one such mutant could be restored by complementation with a non-disrupted copy of the gene. In addition reinsertions occurred on each of the four chromosomes of F. graminearum , making this system a powerful tool in the functional analyses of the > 10,000 genes predicted in the F. graminearum genome