Fungal mycelia are exposed to heterogenic substrates. The substrate in the central part of the colony has been (partly) degraded, whereas it is still unexplored at the periphery of the mycelium. We here assessed whether substrate heterogeneity is a main determinant of spatial gene expression in colonies of Aspergillus niger. This question was addressed by analyzing whole-genome gene expression in five concentric zones of 7-day-old maltose-and xylose-grown colonies. Expression profiles at the periphery and the center were clearly different. More than 25% of the active genes showed twofold differences in expression between the inner and outermost zones of the colony. Moreover, 9% of the genes were expressed in only one of the five concentric zones, showing that a considerable part of the genome is active in a restricted part of the colony only. Statistical analysis of expression profiles of colonies that had either been or not been transferred to fresh xylosecontaining medium showed that differential expression in a colony is due to the heterogeneity of the medium (e.g., genes involved in secretion, genes encoding proteases, and genes involved in xylose metabolism) as well as to medium-independent mechanisms (e.g., genes involved in nitrate metabolism and genes involved in cell wall synthesis and modification). Thus, we conclude that the mycelia of 7-day-old colonies of A. niger are highly differentiated. This conclusion is also indicated by the fact that distinct zones of the colony grow and secrete proteins, even after transfer to fresh medium.Filamentous fungi fulfill an essential role in global carbon cycling by degrading organic material. The organic material is colonized by a mycelium that consists of interconnected hyphae. These hyphae are exposed to a heterogenic substrate. The periphery of the colony is exposed to unexplored organic material, whereas its center is surrounded with a substrate that has been (partly) utilized. Little is known of how this heterogeneity affects cellular processes and gene expression in different parts of the mycelium. However, previous studies suggest that the vegetative colonizing mycelium is highly differentiated (11,21,24). Growth of Aspergillus niger and Phanerochaete chrysosporium mainly takes place in specific zones at the periphery and the center of the colony (11, 24). These zones also secrete proteins. For instance, glucoamylase is secreted at the periphery of an A. niger colony (24), whereas lignin peroxidase is secreted in the central growth zone of P. chrysosporium (11). Interestingly, the hyphae that release the peroxidase are morphologically distinct from the secreting hyphae at the periphery of the vegetative mycelium. The high degree of differentiation in fungal vegetative mycelia is also indicated by the fact that only a subset of growing hyphae at the periphery of an A. niger colony secrete glucoamylase (21, 24).Previous studies have shown that certain genes are differentially expressed in central and peripheral zones of the vegetative mycelium (11,18,21,25)....

Aspergillus niger is a cell factory for the production of enzymes. This fungus secretes proteins in the central part and at the periphery of the colony. The sporulating zone of the colony overlapped with the nonsecreting subperipheral zone, indicating that sporulation inhibits protein secretion. Indeed, strain ΔflbA that is affected early in the sporulation program secreted proteins throughout the colony. In contrast, the ΔbrlA strain that initiates but not completes sporulation did not show altered spatial secretion. The secretome of 5 concentric zones of xylose-grown ΔflbA colonies was assessed by quantitative proteomics. In total 138 proteins with a signal sequence for secretion were identified in the medium of ΔflbA colonies. Of these, 18 proteins had never been reported to be part of the secretome of A. niger, while 101 proteins had previously not been identified in the culture medium of xylose-grown wild type colonies. Taken together, inactivation of flbA results in spatial changes in secretion and in a more complex secretome. The latter may be explained by the fact that strain ΔflbA has a thinner cell wall compared to the wild type, enabling efficient release of proteins. These results are of interest to improve A. niger as a cell factory.

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We report the progress of a multi-disciplinary research project on solid-state fermentation (SSF) of the filamentous fungus Aspergillus oryzae. The molecular and physiological aspects of the fungus in submerged fermentation (SmF) and SSF are compared and we observe a number of differences correlated with the different growth conditions. First, the aerial hyphae which occur only in SSFs are mainly responsible for oxygen uptake. Second, SSF is characterised by gradients in temperature, water activity and nutrient concentration, and inside the hyphae different polyols are accumulating. Third, pelleted growth in SmF and mycelial growth in SSF show different gene expression and protein secretion patterns. With this approach we aim to expand our knowledge of mechanisms of fungal growth on solid substrates and to exploit the biotechnological applications.