Dataset Information

Systemic analysis of the response of Aspergillus niger to ambient pH.

ABSTRACT:

Background

The filamentous fungus Aspergillus niger is an exceptionally efficient producer of organic acids, which is one of the reasons for its relevance to industrial processes and commercial importance. While it is known that the mechanisms regulating this production are tied to the levels of ambient pH, the reasons and mechanisms for this are poorly understood.

Methods

To cast light on the connection between extracellular pH and acid production, we integrate results from two genome-based strategies: A novel method of genome-scale modeling of the response, and transcriptome analysis across three levels of pH.

Results

With genome scale modeling with an optimization for extracellular proton-production, it was possible to reproduce the preferred pH levels for citrate and oxalate. Transcriptome analysis and clustering expanded upon these results and allowed the identification of 162 clusters with distinct transcription patterns across the different pH-levels examined. New and previously described pH-dependent cis-acting promoter elements were identified. Combining transcriptome data with genomic coordinates identified four pH-regulated secondary metabolite gene clusters. Integration of regulatory profiles with functional genomics led to the identification of candidate genes for all steps of the pal/pacC pH signalling pathway.

Conclusions

The combination of genome-scale modeling with comparative genomics and transcriptome analysis has provided systems-wide insights into the evolution of highly efficient acidification as well as production process applicable knowledge on the transcriptional regulation of pH response in the industrially important A. niger. It has also made clear that filamentous fungi have evolved to employ several offensive strategies for out-competing rival organisms.

SUBMITTER: Andersen MR

PROVIDER: S-EPMC2718513 | biostudies-literature | 2009

REPOSITORIES: biostudies-literature

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Publications

Systemic analysis of the response of Aspergillus niger to ambient pH.

Andersen Mikael R MR Lehmann Linda L Nielsen Jens J

Genome biology 20090501 5

<h4>Background</h4>The filamentous fungus Aspergillus niger is an exceptionally efficient producer of organic acids, which is one of the reasons for its relevance to industrial processes and commercial importance. While it is known that the mechanisms regulating this production are tied to the levels of ambient pH, the reasons and mechanisms for this are poorly understood.<h4>Methods</h4>To cast light on the connection between extracellular pH and acid production, we integrate results from two g ...[more]

PMID: 19409083

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Project description:Fungi utilize polysaccharide substrates through extracellular digestion catalyzed by secreted enzymes. Thus far, protein secretion by the filamentous fungus Aspergillus niger has mainly been studied at the level of individual proteins and by genome and transcriptome analyses. To extend these studies, a complementary proteomics approach was applied with the aim to investigate the changes in secretome and microsomal protein composition resulting from a shift to a high level secretion condition. During growth of A. niger on D-sorbitol, small amounts of D-maltose or D-xylose were used as inducers of the extracellular amylolytic and xylanolytic enzymes. Upon induction, protein compositions in the extracellular broth as well as in enriched secretory organelle (microsomal) fractions were analyzed using a shotgun proteomics approach. In total 102 secreted proteins and 1,126 microsomal proteins were identified in this study. Induction by D-maltose or D-xylose resulted in the increase in specific extracellular enzymes, such as glucoamylase A on D-maltose and ?-xylosidase D on D-xylose, as well as of microsomal proteins. This reflects the differential expression of selected genes coding for dedicated extracellular enzymes. As expected, the addition of extra D-sorbitol had no effect on the expression of carbohydrate-active enzymes, compared to addition of D-xylose or D-maltose. Furthermore, D-maltose induction caused an increase in microsomal proteins related to translation (e.g., Rpl15) and vesicular transport (e.g., the endosomal-cargo receptor Erv14). Millimolar amounts of the inducers D-maltose and D-xylose are sufficient to cause a direct response in specific protein expression levels. Also, after induction by D-maltose or D-xylose, the induced enzymes were found in microsomes and extracellular. In agreement with our previous findings for D-xylose induction, D-maltose induction leads to recruitment of proteins involved in proteasome-mediated degradation.

Project description:Filamentous fungi such as Aspergillus niger have a high capacity secretory system and are therefore widely exploited for the industrial production of native and heterologous proteins. However, in most cases the yields of non-fungal proteins are significantly lower than those obtained for fungal proteins. One well-studied bottleneck appears to be the result of mis-folding of heterologous proteins in the ER during early stages of secretion, with related stress responses in the host, including the unfolded protein response (UPR). This study aims at uncovering transcriptional and translational responses occurring in A. niger exposed to secretion stress.A genome-wide transcriptional analysis of protein secretion-related stress responses was determined using Affymetrix DNA GeneChips and independent verification for selected genes. Endoplasmic reticulum (ER)-associated stress was induced either by chemical treatment of the wild-type cells with dithiothreitol (DTT) or tunicamycin, or by expressing a human protein, tissue plasminogen activator (t-PA). All of these treatments triggered the UPR, as shown by the expression levels of several well-known UPR target genes. The predicted proteins encoded by most of the up-regulated genes function as part of the secretory system including chaperones, foldases, glycosylation enzymes, vesicle transport proteins, and ER-associated degradation proteins. Several genes were down-regulated under stress conditions and these included several genes that encode secreted enzymes. Moreover, translational regulation under ER stress was investigated by polysomal fractionation. This analysis confirmed the post-transcriptional control of hacA expression and highlighted that differential translation also occurs during ER stress, in particular for some genes encoding secreted proteins or proteins involved in ribosomal biogenesis and assembly.This is first genome-wide analysis of both transcriptional and translational events following protein secretion stress. Insight has been gained into the molecular basis of protein secretion and secretion-related stress in an effective protein-secreting fungus, and provides an opportunity to identify target genes for manipulation in strain improvement strategies.

Dataset Information

Systemic analysis of the response of Aspergillus niger to ambient pH.

Background

Methods

Results

Conclusions

Publications

Systemic analysis of the response of Aspergillus niger to ambient pH.

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