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Genome-wide expression analysis upon constitutive activation of the HacA bZIP transcription factor in Aspergillus niger reveals a coordinated cellular response to counteract ER stress.

Carvalho ND, Jørgensen TR, Arentshorst M, Nitsche BM, van den Hondel CA, Archer DB, Ram AF - BMC Genomics (2012)

Bottom Line: In addition to the well known HacA targets such as the ER resident foldases and chaperones, GO enrichment analysis revealed up-regulation of genes involved in protein glycosylation, phospholipid biosynthesis, intracellular protein transport, exocytosis and protein complex assembly in the HacACA mutant.Biological processes over-represented in the down-regulated genes include those belonging to central metabolic pathways, translation and transcription.The results indicate that the constitutive activation of the HacA leads to a coordinated regulation of the folding and secretion capacity of the cell, but with consequences on growth and fungal physiology to reduce secretion stress.

View Article: PubMed Central - HTML - PubMed

Affiliation: Institute of Biology Leiden, Leiden University, Molecular Microbiology and Biotechnology, BE Leiden, The Netherlands.

ABSTRACT

Background: HacA/Xbp1 is a conserved bZIP transcription factor in eukaryotic cells which regulates gene expression in response to various forms of secretion stress and as part of secretory cell differentiation. In the present study, we replaced the endogenous hacA gene of an Aspergillus niger strain with a gene encoding a constitutively active form of the HacA transcription factor (HacACA). The impact of constitutive HacA activity during exponential growth was explored in bioreactor controlled cultures using transcriptomic analysis to identify affected genes and processes.

Results: Transcription profiles for the wild-type strain (HacAWT) and the HacACA strain were obtained using Affymetrix GeneChip analysis of three replicate batch cultures of each strain. In addition to the well known HacA targets such as the ER resident foldases and chaperones, GO enrichment analysis revealed up-regulation of genes involved in protein glycosylation, phospholipid biosynthesis, intracellular protein transport, exocytosis and protein complex assembly in the HacACA mutant. Biological processes over-represented in the down-regulated genes include those belonging to central metabolic pathways, translation and transcription. A remarkable transcriptional response in the HacACA strain was the down-regulation of the AmyR transcription factor and its target genes.

Conclusions: The results indicate that the constitutive activation of the HacA leads to a coordinated regulation of the folding and secretion capacity of the cell, but with consequences on growth and fungal physiology to reduce secretion stress.

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Related in: MedlinePlus

Analysis of differentially expressed genes in HacACA (this study) and Guillemette’ study[[37]]. Venn diagrams of the number of overlapping and non-overlapping induced or repressed genes of A. niger  HacACA-1/HacAWT(A) and genes from Guillemette et al. (2007) induced (or repressed) at least in two conditions (B) or induced in all conditions (C).
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Figure 5: Analysis of differentially expressed genes in HacACA (this study) and Guillemette’ study[[37]]. Venn diagrams of the number of overlapping and non-overlapping induced or repressed genes of A. niger HacACA-1/HacAWT(A) and genes from Guillemette et al. (2007) induced (or repressed) at least in two conditions (B) or induced in all conditions (C).

Mentions: To gain a broader overview of the impact of a constitutive activation of HacA on A. niger we compared our data set (HacACA-1/HacAWT) with the data of Guillemette and co-workers ([37]; [Additional file 3 and Additional file 16) in which the genome-wide transcriptional protein secretion-related stress responses was analyzed. In this study [37], transcriptional targets of the UPR pathway were identified by treatment of A. niger with the ER-disturbing chemical agents tunicamycin and dithiothreitol (DTT) and using a strain producing the recombinant tissue plasminogen activator (t-PA) as a model for heterologous protein production. As shown in Figure 5, in the induced set of genes, 13 genes are commonly unregulated in both studies (all conditions) and 81 genes are differentially expressed in HacACA-1/HacAWT in at least two of the three conditions performed by Guillemette et al. [37]. These 94 commonly induced genes include all the genes identified in the Guillemette et al.’ study related to protein folding, translocation/signal peptidase complex and glycosylation and most of the genes that belong to the categories of vesicle trafficking and lipid metabolism [Additional file 16. However, more genes belonging to each of these categories have been identified in the HacACA-1/HacAWT comparison (Figure 4 and Additional file 10, Additional file 11, Additional file 12, Additional file 13). Unique genes found in at least two of the conditions tested (56) and not in our data set relate mainly to the categories of cellular transport, stress related, amino acid metabolism, carbohydrate metabolism and unclassified genes.


Genome-wide expression analysis upon constitutive activation of the HacA bZIP transcription factor in Aspergillus niger reveals a coordinated cellular response to counteract ER stress.

Carvalho ND, Jørgensen TR, Arentshorst M, Nitsche BM, van den Hondel CA, Archer DB, Ram AF - BMC Genomics (2012)

Analysis of differentially expressed genes in HacACA (this study) and Guillemette’ study[[37]]. Venn diagrams of the number of overlapping and non-overlapping induced or repressed genes of A. niger  HacACA-1/HacAWT(A) and genes from Guillemette et al. (2007) induced (or repressed) at least in two conditions (B) or induced in all conditions (C).
© Copyright Policy - open-access
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC3472299&req=5

Figure 5: Analysis of differentially expressed genes in HacACA (this study) and Guillemette’ study[[37]]. Venn diagrams of the number of overlapping and non-overlapping induced or repressed genes of A. niger HacACA-1/HacAWT(A) and genes from Guillemette et al. (2007) induced (or repressed) at least in two conditions (B) or induced in all conditions (C).
Mentions: To gain a broader overview of the impact of a constitutive activation of HacA on A. niger we compared our data set (HacACA-1/HacAWT) with the data of Guillemette and co-workers ([37]; [Additional file 3 and Additional file 16) in which the genome-wide transcriptional protein secretion-related stress responses was analyzed. In this study [37], transcriptional targets of the UPR pathway were identified by treatment of A. niger with the ER-disturbing chemical agents tunicamycin and dithiothreitol (DTT) and using a strain producing the recombinant tissue plasminogen activator (t-PA) as a model for heterologous protein production. As shown in Figure 5, in the induced set of genes, 13 genes are commonly unregulated in both studies (all conditions) and 81 genes are differentially expressed in HacACA-1/HacAWT in at least two of the three conditions performed by Guillemette et al. [37]. These 94 commonly induced genes include all the genes identified in the Guillemette et al.’ study related to protein folding, translocation/signal peptidase complex and glycosylation and most of the genes that belong to the categories of vesicle trafficking and lipid metabolism [Additional file 16. However, more genes belonging to each of these categories have been identified in the HacACA-1/HacAWT comparison (Figure 4 and Additional file 10, Additional file 11, Additional file 12, Additional file 13). Unique genes found in at least two of the conditions tested (56) and not in our data set relate mainly to the categories of cellular transport, stress related, amino acid metabolism, carbohydrate metabolism and unclassified genes.

Bottom Line: In addition to the well known HacA targets such as the ER resident foldases and chaperones, GO enrichment analysis revealed up-regulation of genes involved in protein glycosylation, phospholipid biosynthesis, intracellular protein transport, exocytosis and protein complex assembly in the HacACA mutant.Biological processes over-represented in the down-regulated genes include those belonging to central metabolic pathways, translation and transcription.The results indicate that the constitutive activation of the HacA leads to a coordinated regulation of the folding and secretion capacity of the cell, but with consequences on growth and fungal physiology to reduce secretion stress.

View Article: PubMed Central - HTML - PubMed

Affiliation: Institute of Biology Leiden, Leiden University, Molecular Microbiology and Biotechnology, BE Leiden, The Netherlands.

ABSTRACT

Background: HacA/Xbp1 is a conserved bZIP transcription factor in eukaryotic cells which regulates gene expression in response to various forms of secretion stress and as part of secretory cell differentiation. In the present study, we replaced the endogenous hacA gene of an Aspergillus niger strain with a gene encoding a constitutively active form of the HacA transcription factor (HacACA). The impact of constitutive HacA activity during exponential growth was explored in bioreactor controlled cultures using transcriptomic analysis to identify affected genes and processes.

Results: Transcription profiles for the wild-type strain (HacAWT) and the HacACA strain were obtained using Affymetrix GeneChip analysis of three replicate batch cultures of each strain. In addition to the well known HacA targets such as the ER resident foldases and chaperones, GO enrichment analysis revealed up-regulation of genes involved in protein glycosylation, phospholipid biosynthesis, intracellular protein transport, exocytosis and protein complex assembly in the HacACA mutant. Biological processes over-represented in the down-regulated genes include those belonging to central metabolic pathways, translation and transcription. A remarkable transcriptional response in the HacACA strain was the down-regulation of the AmyR transcription factor and its target genes.

Conclusions: The results indicate that the constitutive activation of the HacA leads to a coordinated regulation of the folding and secretion capacity of the cell, but with consequences on growth and fungal physiology to reduce secretion stress.

Show MeSH
Related in: MedlinePlus