Limits...
Kinetic transcriptome analysis reveals an essentially intact induction system in a cellulase hyper-producer Trichoderma reesei strain.

Poggi-Parodi D, Bidard F, Pirayre A, Portnoy T, Blugeon C, Seiboth B, Kubicek CP, Le Crom S, Margeot A - Biotechnol Biofuels (2014)

Bottom Line: Cross-comparison of our transcriptome data with previously identified mutations revealed that most genes from our dataset have not been mutated.The fact that few regulated genes have been affected by mutagenesis suggests that the induction mechanism is essentially intact compared to that for the wild-type isolate QM6a and might be engineered for further improvement of T. reesei.Genes from two specific clusters might be potential targets for such genetic engineering.

View Article: PubMed Central - PubMed

Affiliation: IFP Energies nouvelles, 1-4 avenue de Bois-Préau, 92852 Rueil-Malmaison, France ; Sorbonne Universités, UPMC Univ Paris 06, Institut de Biologie Paris-Seine (IBPS), F-75005 Paris, France.

ABSTRACT

Background: The filamentous fungus Trichoderma reesei is the main industrial cellulolytic enzyme producer. Several strains have been developed in the past using random mutagenesis, and despite impressive performance enhancements, the pressure for low-cost cellulases has stimulated continuous research in the field. In this context, comparative study of the lower and higher producer strains obtained through random mutagenesis using systems biology tools (genome and transcriptome sequencing) can shed light on the mechanisms of cellulase production and help identify genes linked to performance. Previously, our group published comparative genome sequencing of the lower and higher producer strains NG 14 and RUT C30. In this follow-up work, we examine how these mutations affect phenotype as regards the transcriptome and cultivation behaviour.

Results: We performed kinetic transcriptome analysis of the NG 14 and RUT C30 strains of early enzyme production induced by lactose using bioreactor cultivations close to an industrial cultivation regime. RUT C30 exhibited both earlier onset of protein production (3 h) and higher steady-state productivity. A rather small number of genes compared to previous studies were regulated (568), most of them being specific to the NG 14 strain (319). Clustering analysis highlighted similar behaviour for some functional categories and allowed us to distinguish between induction-related genes and productivity-related genes. Cross-comparison of our transcriptome data with previously identified mutations revealed that most genes from our dataset have not been mutated. Interestingly, the few mutated genes belong to the same clusters, suggesting that these clusters contain genes playing a role in strain performance.

Conclusions: This is the first kinetic analysis of a transcriptomic study carried out under conditions approaching industrial ones with two related strains of T. reesei showing distinctive cultivation behaviour. Our study sheds some light on some of the events occurring in these strains following induction by lactose. The fact that few regulated genes have been affected by mutagenesis suggests that the induction mechanism is essentially intact compared to that for the wild-type isolate QM6a and might be engineered for further improvement of T. reesei. Genes from two specific clusters might be potential targets for such genetic engineering.

No MeSH data available.


Related in: MedlinePlus

Schematic view of gene expression kinetics in NG 14 and RUT C30. Approximate regulation timing for each cluster is shown, and the number of each cluster is indicated inside each arrow. Red up-arrows and green down-arrows represent up- and down-regulated clusters respectively. Black bold line indicates the start time of protein induction and production in each strain. Main functional categories found in each cluster are indicated as Neo (gluconeogenesis), Rib (ribosome biogenesis), AA (amino acid metabolism), Lip (lipid metabolism), Caz (CAZymes), TP (transporter), Met (metabolism), Pro (extracellular protein degradation), Sec (secretion) and Ctrl (transcriptional control).
© Copyright Policy - open-access
Related In: Results  -  Collection

License 1 - License 2
getmorefigures.php?uid=PMC4279801&req=5

Fig5: Schematic view of gene expression kinetics in NG 14 and RUT C30. Approximate regulation timing for each cluster is shown, and the number of each cluster is indicated inside each arrow. Red up-arrows and green down-arrows represent up- and down-regulated clusters respectively. Black bold line indicates the start time of protein induction and production in each strain. Main functional categories found in each cluster are indicated as Neo (gluconeogenesis), Rib (ribosome biogenesis), AA (amino acid metabolism), Lip (lipid metabolism), Caz (CAZymes), TP (transporter), Met (metabolism), Pro (extracellular protein degradation), Sec (secretion) and Ctrl (transcriptional control).

Mentions: NG 14 and RUT C30 have seldom been compared for their respective performances [3,44,45]. Our data showed that RUT C30 has a specific productivity twice as high as that of NG 14 during the first 50 h of the fed-batch phase. Moreover, two main differences are reflected by the production curves (Figure 1); RUT C30 starts production at least 3 h earlier, and steady-state productivity during fed-batch is higher than for NG 14. A comparison of differentially expressed clusters found for each strain and sorted by time point is shown in Figure 5. It is interesting to note that no cluster is temporally correlated with the onset of protein production.Figure 5


Kinetic transcriptome analysis reveals an essentially intact induction system in a cellulase hyper-producer Trichoderma reesei strain.

Poggi-Parodi D, Bidard F, Pirayre A, Portnoy T, Blugeon C, Seiboth B, Kubicek CP, Le Crom S, Margeot A - Biotechnol Biofuels (2014)

Schematic view of gene expression kinetics in NG 14 and RUT C30. Approximate regulation timing for each cluster is shown, and the number of each cluster is indicated inside each arrow. Red up-arrows and green down-arrows represent up- and down-regulated clusters respectively. Black bold line indicates the start time of protein induction and production in each strain. Main functional categories found in each cluster are indicated as Neo (gluconeogenesis), Rib (ribosome biogenesis), AA (amino acid metabolism), Lip (lipid metabolism), Caz (CAZymes), TP (transporter), Met (metabolism), Pro (extracellular protein degradation), Sec (secretion) and Ctrl (transcriptional control).
© Copyright Policy - open-access
Related In: Results  -  Collection

License 1 - License 2
Show All Figures
getmorefigures.php?uid=PMC4279801&req=5

Fig5: Schematic view of gene expression kinetics in NG 14 and RUT C30. Approximate regulation timing for each cluster is shown, and the number of each cluster is indicated inside each arrow. Red up-arrows and green down-arrows represent up- and down-regulated clusters respectively. Black bold line indicates the start time of protein induction and production in each strain. Main functional categories found in each cluster are indicated as Neo (gluconeogenesis), Rib (ribosome biogenesis), AA (amino acid metabolism), Lip (lipid metabolism), Caz (CAZymes), TP (transporter), Met (metabolism), Pro (extracellular protein degradation), Sec (secretion) and Ctrl (transcriptional control).
Mentions: NG 14 and RUT C30 have seldom been compared for their respective performances [3,44,45]. Our data showed that RUT C30 has a specific productivity twice as high as that of NG 14 during the first 50 h of the fed-batch phase. Moreover, two main differences are reflected by the production curves (Figure 1); RUT C30 starts production at least 3 h earlier, and steady-state productivity during fed-batch is higher than for NG 14. A comparison of differentially expressed clusters found for each strain and sorted by time point is shown in Figure 5. It is interesting to note that no cluster is temporally correlated with the onset of protein production.Figure 5

Bottom Line: Cross-comparison of our transcriptome data with previously identified mutations revealed that most genes from our dataset have not been mutated.The fact that few regulated genes have been affected by mutagenesis suggests that the induction mechanism is essentially intact compared to that for the wild-type isolate QM6a and might be engineered for further improvement of T. reesei.Genes from two specific clusters might be potential targets for such genetic engineering.

View Article: PubMed Central - PubMed

Affiliation: IFP Energies nouvelles, 1-4 avenue de Bois-Préau, 92852 Rueil-Malmaison, France ; Sorbonne Universités, UPMC Univ Paris 06, Institut de Biologie Paris-Seine (IBPS), F-75005 Paris, France.

ABSTRACT

Background: The filamentous fungus Trichoderma reesei is the main industrial cellulolytic enzyme producer. Several strains have been developed in the past using random mutagenesis, and despite impressive performance enhancements, the pressure for low-cost cellulases has stimulated continuous research in the field. In this context, comparative study of the lower and higher producer strains obtained through random mutagenesis using systems biology tools (genome and transcriptome sequencing) can shed light on the mechanisms of cellulase production and help identify genes linked to performance. Previously, our group published comparative genome sequencing of the lower and higher producer strains NG 14 and RUT C30. In this follow-up work, we examine how these mutations affect phenotype as regards the transcriptome and cultivation behaviour.

Results: We performed kinetic transcriptome analysis of the NG 14 and RUT C30 strains of early enzyme production induced by lactose using bioreactor cultivations close to an industrial cultivation regime. RUT C30 exhibited both earlier onset of protein production (3 h) and higher steady-state productivity. A rather small number of genes compared to previous studies were regulated (568), most of them being specific to the NG 14 strain (319). Clustering analysis highlighted similar behaviour for some functional categories and allowed us to distinguish between induction-related genes and productivity-related genes. Cross-comparison of our transcriptome data with previously identified mutations revealed that most genes from our dataset have not been mutated. Interestingly, the few mutated genes belong to the same clusters, suggesting that these clusters contain genes playing a role in strain performance.

Conclusions: This is the first kinetic analysis of a transcriptomic study carried out under conditions approaching industrial ones with two related strains of T. reesei showing distinctive cultivation behaviour. Our study sheds some light on some of the events occurring in these strains following induction by lactose. The fact that few regulated genes have been affected by mutagenesis suggests that the induction mechanism is essentially intact compared to that for the wild-type isolate QM6a and might be engineered for further improvement of T. reesei. Genes from two specific clusters might be potential targets for such genetic engineering.

No MeSH data available.


Related in: MedlinePlus