Assessment of the pectinolytic network of Aspergillus niger by functional genomics : insights from the transcriptome

More than a century ago, in 1889, A. Fernbach presented a detailed report about the invertase of Aspergillus niger in the third edition of “Annales De L'institut Pasteur”. Since then, many of the enzymes secreted by A. niger have found a broad range of applications, and today they are produced on an industrial scale. This filamentous fungus is also used as a primary source for the production of organic acids and other economically important metabolites. Although, many of these fermentation processes are well established, the underlying genetics are still not well understood. The recent determination of the genome sequence of A. niger illustrated the versatile metabolic capacities of this fungus and created the opportunity for challenging research aiming to unravel the fine tuned metabolic network and the full enzymatic potential of A. niger. The work described in this thesis is focused on the genome mining and transcriptional profiling of the enzymatic network of A. niger involved in pectin degradation and utilization. Pectin was chosen because of the broad application of it and its derivates in food, feed, medicines, and cosmetics and the economical importance of this polymer in several technological processes. This study addresses several issues concerning pectin degradation by A. niger: i) the complete inventory of the known and potential pectinolytic activities encoded in the genome of this fungus; ii) the understanding of dynamics of their gene-expression on (complex) carbon sources in order to unravel underling metabolic networks; iii) the evolutionary aspects of the pectinolytic system of A. niger and other filamentous fungi. Chapter 1 of this thesis discusses the importance of fungi and A. niger for the biotech industry, and gives a brief introduction to the structural elements of pectin, the types of enzymatic activities involved in its degradation, and the present knowledge of the metabolism of its major constituent – galacturonic acid. The current advances in fungal ‘omics research are pointed out. Chapter 2 presents the construction of pectin specific cDNA libraries from A. niger, and the annotation of more than 200 of the obtained expressed sequence tags. Chapter 3 focuses on the roles of and the interactions between the 21 genes within glycoside hydrolase family 28 - the largest group of A. niger pectinases. A special emphasis is given to a new, previously unanticipated, group of exo-acting enzymes. Chapter 4 describes the survey for all genes encoding pectinolytic activities in A. niger and their transcriptional profiles. In this chapter, a hypothetical model of the pectinolytic network of A. niger is proposed. Chapter 5 focuses on the identification of the genes involved in galacturonic acid catabolism in A. niger, the verification of some of the enzymatic activities encoded by them, and elaborates on the presence of an evolutionary conserved galacturonic acid pathway operating only in filamentous fungi capable of plant cell wall degradation. In Chapter 6, the evolutionary conservation of transcriptional response of the pectinolytic system of A. niger is compared to that of the model organism A. nidulans. This chapter further describes the identification of the evolutionary conserved, regulatory core of functional orthologs involved in galacturonic acid utilization and metabolism. In Chapter 7, the results of the work described in this thesis are summarized and discussed.