**11. Induction of cellulases**

*chaete chrysosporium*, *Ceriporiopsis subvermispora*, *Phlebia subserialis*, and *Pleurotus ostreatus*, which are able to metabolize the lignin in a variety of lignocellulosic biomass [62, 67, 68]. In addition, other species of fungi, such as *Postia placenta* (a brown-rot fungus), and some bacteria (such as *Azospirillum lipoferum* and *Marinomonas mediterranea*), are able to metabolize lignin. The saprotrophic homobasidiomycete *Pycnoporus cinnabarinus* is recog‐ nized by its high lignocellulolytic potential [69] overproducing high redox potential lac‐ case, and a variety of studies have been performed in order to increase the ability of this specie to produce laccases for biotechnological applications, including heterologous ex‐ pression in other species such as *A. niger* [70, 71, 72]. In addition, white-rot fungi such as *Cyathus cinnabarinus* and *Cyathus bulleri* demonstrated potential to degrade lignin [73, 74].

220 Sustainable Degradation of Lignocellulosic Biomass - Techniques, Applications and Commercialization

In summary, microbial degradation of lignocellulosic material requires a concerted action of a variety of enzymes arranged in an enzymatic complex, depending on the biomass to be degraded. The gene expression, production and secretion of plant cell wall-degrading en‐ zymes demand energy from the microbial cells and therefore the overall process is highly regulated. There is an intense cross-talk in induction of expression of the genes encoding dif‐ ferent classes of enzymes. The control of the regulation of CWDEs production could be the key for the development of new microbial strains that efficiently produce and secrete CWDEs. The regulation of genes encoding polysaccharide-degrading enzymes will be the

**10. Regulation of cell-wall degrading enzymes production in fungi**

The production of CWDEs by fungi is an energy-consuming process. The fine-tuned regula‐ tion of genes encoding CWDEs ensures that these enzymes will be produced only under conditions in which the fungus requires plant polymers as carbon source. Readily metabo‐ lizable carbohydrates repress the synthesis of enzymes related to catabolism of alternative carbon sources such as plant cell wall polysaccharides. In this way, preferential utilization of the most favored carbon source prevails, and one of the regulatory mechanisms involved in this adaptation is carbon catabolite repression (CCR). The CCR is activated by many carbon sources, depending on the lifestyle of the microorganism, but usually glucose is the most re‐ pressive molecule [75]. Nowadays, the search for microorganisms able to efficiently degrade lignocellulosic biomass is pivotal for the establishment of sustainable production of bio‐ mass-derived ethanol and other biocompounds. In this context, CCR appears as a major challenge to overcome, once this mechanism is responsible for enzymatic exclusion of less preferred carbon source such as lignocellulose-derived sugars. Hence, the comprehension of molecular mechanisms behind CCR, as well as the transcriptional control of cell wall de‐ rived enzymes are prerequisite in order to develop new microbial strains for lignocellulose degradation. In this section, the induction of expression of cellulases and hemicellulases, the transcriptional control of genes encoding CWDEs and the overall mechanism behind CCR

subject of the next section of this chapter.

will be discussed.

Although the biochemistry of the process behind lignocellulosic degradation has been stud‐ ied in detail, the mechanism by which filamentous fungi sense the substrate and initiate the overall process of hydrolases production is still unsolved. Some researchers have been pro‐ posed that a low constitutive level of cellulase expression is responsible for the formation of an inducer from cellulose, amplifying the signal. Another group of scientists suggest that the fungus initiates a starvation process, which could in turn activates cellulase/hemicellulase expression. Also, it is possible that an inducing sugar derived from carbohydrates released somehow from the fungal cell wall could be the derepressing molecules for hydrolase in‐ duction. Despite of the fact that the true mechanism behind natural cellulase/hemicellulase induction is still lacking, some individual molecules are known to induce these hydrolases.

The fungus *Trichoderma ressei* is an impressive producer of cellulases and most of studies concerning the regulation of cellulase genes have been performed in this specie. The most powerful inducer of cellulases in *T. reesei* is sophorose, a disaccharide composed of β-1,2 linked glucose units. Sophorose appears to be formed from cellobiose through transglycosy‐ lation activity of β-glucosidase [76 - 78]. In addition to *T. reesei*, sophorose is known to induce cellulase expression in *A. terreus* and *P. purpurogenum* [79 - 80].

Cellobiose (two β-1,4-linked glucose units) appears to induce cellulase expression in many species of fungi. Cellobiose is formed as the end product of cellobiohydrolases activity, and it has been show to induce cellulase expression in *T. reesei*, *Volvariella volvacea*, *P. janthinellum* and *A. nidulans* [81 - 84]. However, studies concerning the inducing effect of cellobiose on cellulase expression are controversial [6]. For instance, cellobiose can be transglycosylated by β-glucosidases, producing sophorose, which could be the true inducer of cellulases. Be‐ sides, β-glucosidases are able to cleave the cellobiose into glucose, which may cause repres‐ sion by CCR. Therefore, the outcome in cellobiose cultures appears to be dependent on the balance between hydrolysis and transglycosylation, as well as the subsequent uptake of the generated sugars and the intracellular signals they initiate.

Lactose (1,4-*O*- β-D-galactopyranosyl-D-glucose) is a disaccharide that has been shown eco‐ nomically viable to induce cellulase expression in *T. reesei*. Interestingly, lactose is not a com‐ ponent of plant cell wall polymers and the mechanism through which this sugar induces cellulase expression appears to be complex. In filamentous fungi, lactose is cleaved by ex‐ tracellular β-galactosidase into glucose and galactose. Lactose induction of cellulase genes requires the β-anomer of D-galactose, which can be converted to fructose by an alternative pathway in addition to the Leloir pathway [85]. In this alternative pathway, D-xylose reduc‐ tase (encoded by *xyl1*) is the enzyme catalyzing the first step [86].

Moreover, induction of cellulase genes could be achieved in *T. reesei* cultures after addition of various other oligosaccharides such as laminaribiose, gentiobiose, xylobiose, L-sorbose and δ-cellobiono-1,5-lactone. L-arabitol and different xylans also have been show to induce expression of cellobiohydrolase 1 (*cbh1*) in *T. reesei* (reviewed in reference [6].
