Dataset Information

Proximity ligation scaffolding and comparison of two Trichoderma reesei strains genomes.

ABSTRACT: BACKGROUND:The presence of low complexity and repeated regions in genomes often results in difficulties to assemble sequencing data into full chromosomes. However, the availability of full genome scaffolds is essential to several investigations, regarding for instance the evolution of entire clades, the analysis of chromosome rearrangements, and is pivotal to sexual crossing studies. In non-conventional but industrially relevant model organisms, such as the ascomycete Trichoderma reesei, a complete genome assembly is seldom available. RESULTS:The chromosome scaffolds of T. reesei QM6a and Rut-C30 strains have been generated using a contact genomic/proximity ligation genomic approach. The original reference assembly, encompassing dozens of scaffolds, was reorganized into two sets of seven chromosomes. Chromosomal contact data also allowed to characterize 10-40 kb, gene-free, AT-rich (76%) regions corresponding to the T. reesei centromeres. Large chromosomal rearrangements (LCR) in Rut-C30 were then characterized, in agreement with former studies, and the position of LCR breakpoints used to assess the likely chromosome structure of other T. reesei strains [QM9414, CBS999.97 (1-1, re), and QM9978]. In agreement with published results, we predict that the numerous chromosome rearrangements found in highly mutated industrial strains may limit the efficiency of sexual reproduction for their improvement. CONCLUSIONS:The GRAAL program allowed us to generate the karyotype of the Rut-C30 strain, and from there to predict chromosome structure for most T. reesei strains for which sequence is available. This method that exploits proximity ligation sequencing approach is a fast, cheap, and straightforward way to characterize both chromosome structure and centromere sequences and is likely to represent a popular convenient alternative to expensive and work-intensive resequencing projects.

SUBMITTER: Jourdier E

PROVIDER: S-EPMC5469131 | biostudies-literature | 2017

REPOSITORIES: biostudies-literature

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Proximity ligation scaffolding and comparison of two <i>Trichoderma reesei</i> strains genomes.

Jourdier Etienne E Baudry Lyam L Poggi-Parodi Dante D Vicq Yoan Y Koszul Romain R Margeot Antoine A Marbouty Martial M Bidard Frédérique F

Biotechnology for biofuels 20170612

<h4>Background</h4>The presence of low complexity and repeated regions in genomes often results in difficulties to assemble sequencing data into full chromosomes. However, the availability of full genome scaffolds is essential to several investigations, regarding for instance the evolution of entire clades, the analysis of chromosome rearrangements, and is pivotal to sexual crossing studies. In non-conventional but industrially relevant model organisms, such as the ascomycete <i>Trichoderma rees ...[more]

PMID: 28616075

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Project description:BackgroundThe tropical ascomycete Trichoderma reesei (Hypocrea jecorina) represents one of the most efficient plant cell wall degraders. Regulation of the enzymes required for this process is affected by nutritional signals as well as other environmental signals including light.ResultsOur transcriptome analysis of strains lacking the photoreceptors BLR1 and BLR2 as well as ENV1 revealed a considerable increase in the number of genes showing significantly different transcript levels in light and darkness compared to wild-type. We show that members of all glycoside hydrolase families can be subject to light dependent regulation, hence confirming nutrient utilization including plant cell wall degradation as a major output pathway of light signalling. In contrast to N. crassa, photoreceptor mediated regulation of carbon metabolism in T. reesei occurs primarily by BLR1 and BLR2 via their positive effect on induction of env1 transcription, rather than by a presumed negative effect of ENV1 on the function of the BLR complex. Nevertheless, genes consistently regulated by photoreceptors in N. crassa and T. reesei are significantly enriched in carbon metabolic functions. Hence, different regulatory mechanisms are operative in these two fungi, while the light dependent regulation of plant cell wall degradation appears to be conserved.Analysis of growth on different carbon sources revealed that the oxidoreductive D-galactose and pentose catabolism is influenced by light and ENV1. Transcriptional regulation of the target enzymes in these pathways is enhanced by light and influenced by ENV1, BLR1 and/or BLR2. Additionally we detected an ENV1-regulated genomic cluster of 9 genes including the D-mannitol dehydrogenase gene lxr1, with two genes of this cluster showing consistent regulation in N. crassa.ConclusionsWe show that one major output pathway of light signalling in Trichoderma reesei is regulation of glycoside hydrolase genes and the degradation of hemicellulose building blocks. Targets of ENV1 and BLR1/BLR2 are for the most part distinct and indicate individual functions for ENV1 and the BLR complex besides their postulated regulatory interrelationship.

Project description:BackgroundThe ascomycete fungus Trichoderma reesei was widely used as a biotechnological workhorse for production of cellulases and recombinant proteins due to its large capacity of protein secretion. Transgenesis by random integration of a gene of interest (GOI) into the genome of T. reesei can generate series of strains that express different levels of the indicated transgene. The insertion site of the GOI plays an important role in the ultimate production of the targeted proteins. However, so far no systematic studies have been made to identify transgene integration loci for optimal expression of the GOI in T. reesei. Currently, only the locus of exocellobiohydrolases I encoding gene (cbh1) is widely used as a promising integration site to lead to high expression level of the GOI. No additional sites associated with efficient gene expression have been characterized.ResultsTo search for gene integration sites that benefit for the secreted expression of GOI, the food-and-mouth disease virus 2A protein was applied for co-expression of an Aspergillus niger lipA gene and Discosoma sp. DsRed1 gene in T. reesei, by random integration of the expression cassette into the genome. We demonstrated that the fluorescent intensity of RFP (red fluorescent protein) inside of the cell was well correlated with the secreted lipase yields, based on which, we successfully developed a high-throughput screening method to screen strains with relatively higher secreted expression of the GOI (in this study, lipase). The copy number and the insertion sites of the transgene were investigated among the selected highly expressed strains. Eventually, in addition to cbh1 gene locus, two other genome insertion loci that efficiently facilitate gene expression in T. reesei were identified.ConclusionsWe have successfully developed a high-throughput screening method to screen strains with optimal expression of the indicated secreted proteins in T. reesei. Moreover, we identified two optimal genome loci for transgene expression, which could provide new approach to modulate gene expression levels while retaining the indicated promoter and culture conditions.

Dataset Information

Proximity ligation scaffolding and comparison of two Trichoderma reesei strains genomes.

Publications

Proximity ligation scaffolding and comparison of two <i>Trichoderma reesei</i> strains genomes.

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