A systems biology approach to dissect the morphogenetic network of Aspergillus niger that links growth with product formation

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Filamentous fungi such as Aspergillus niger are harnessed as cell factories for the production of a diverse range of organic acids, proteins, and secondary metabolites. However, the adequate macroscopic fungal morphology for a given target product varies, cannot currently be generalized, and the advantages and disadvantages of mycelial and pellet cultivation have to be carefully adjusted for a given process. Hence, a rational genetic approach to predict and engineer the macroscopic morphology of filamentous fungi for bioprocesses is so far impossible.The overall objective of the proposal is to gain mechanistic insights into the link between citric acid production, protein secretion, growth and morphology of A. niger. Our comprehensive analysis of A. niger transcriptomic data uncovered that a sub-network related to citric acid biosynthesis (“citA network” contains 282 genes, 57 of which can be predicted to be important for polar growth, organelle functioning, vesicle transport, signalling, and cell wall and cell membrane integrity). Notably, 14 of these genes have a predicted function in the Golgi complex, thus allocating a specific function of this vesicle transport machinery and signalling hub in the natural capability of A. niger to overproduce and secrete citric acid. In view of the importance of the Golgi complex in vesicle transport for protein overproduction and protein secretion in A. niger, we propose that a systematic analysis of the predicted 57 “citA morphogenes” will reveal new fundamental insights into the link between citric acid biosynthesis and protein secretion with morphological decisions. Such a proposed link is highly speculative and has not been studied yet in filamentous fungi. We will exploit state-of-the-art tools which have been established in our labs for A. niger including systems biology tools (i.e. genomics, transcriptomics, metabolomics and bioinformatics), synthetic biology tools (i.e. inducible expression allowing growth- and metabolism-independent expression based on the Tet-on system), controlled bioreactor cultivations for physiological characterizations, and confocal fluorescence microscopy to study organelle localisation during misexpression of the citA morphogenes. Using such a holistic approach, which is embedded in four work packages, the role of the “citA morphogenes” for A. niger will be studied in detail to achieve the following four objectives:• Identification of morphology modulators from 57 “citA morphogenes” • Unravelling the importance of 57 “citA morphogenes” for citric acid and protein production• Unravelling the importance of selected “citA morphogenes” for vesicle trafficking, hyphal growth and pellet formation • Deduction of lead genes to rationally mould A. niger with respect to high productivityThe ultimate goal is to understand the mechanisms behind the citA sub-network that balance hyphal growth with citric acid production and protein secretion.