Background:The molecular mechanism underlying the regulation of cellulase production by T. reesei is unclear. Results: The absence of sugar transporter Stp1 enhanced cellulase gene induction whereas the absence of Crt1 abolished cellulase gene expression. Conclusion: Crt1 is essential in cellulase gene induction independent of intracellular sugar delivery. Significance: These data shed light on the mechanism by which T. reesei senses and transmits cellulose signal.

Cryptococcus neoformans is a major fungal pathogen that disseminates to the central nervous system (CNS) to cause fatal meningoencephalitis, but little is known about immune responses within this immune-privileged site. CD4 ϩ T cells have demonstrated roles in anticryptococcal defenses, but increasing evidence suggests that they may contribute to clinical deterioration and pathology in both HIVpositive (HIVϩ) and non-HIV patients who develop immune reconstitution inflammatory syndrome (IRIS) and post-infectious inflammatory response syndrome (PIIRS), respectively. Here we report a novel murine model of cryptococcal meningoencephalitis and a potential damaging role of T cells in disseminated cryptococcal CNS infection. In this model, fungal burdens plateaued in the infected brain by day 7 postinfection, but activation of microglia and accumulation of CD45 hi leukocytes was significantly delayed relative to fungal growth and did not peak until day 21. The inflammatory leukocyte infiltrate consisted predominantly of gamma interferon (IFN-␥)-producing CD4 ϩ T cells, conventionally believed to promote fungal clearance and recovery. However, more than 50% of mice succumbed to infection and neurological dysfunction between days 21 and 35 despite a 100-fold reduction in fungal burdens. Depletion of CD4 ϩ cells significantly impaired IFN-␥ production, CD8 ϩ T cell and myeloid cell accumulation, and fungal clearance from the CNS but prevented the development of clinical symptoms and mortality. These findings conclusively demonstrate that although CD4 ϩ T cells are necessary to control fungal growth, they can also promote significant immunopathology and mortality during CNS infection. The results from this model may provide important guidance for development and use of anti-inflammatory therapies to minimize CNS injury in patients with severe cryptococcal infections.IMPORTANCE CNS infection with the fungal pathogen Cryptococcus neoformans often results in debilitating brain injury and has a high mortality rate despite antifungal treatment. Treatment is complicated by the fact that immune responses needed to eliminate infection are also thought to drive CNS damage in a subset of both HIVϩ and non-HIV patients. Thus, physicians need to balance efforts to enhance patients' immune responses and promote microbiological control with antiinflammatory therapy to protect the CNS. Here we report a novel model of cryptococcal meningoencephalitis demonstrating that fungal growth within the CNS does not immediately cause symptomatic disease. Rather, accumulation of antifungal immune cells critically mediates CNS injury and mortality. This model demonstrates that antifungal immune responses in the CNS can cause detrimental pathology and

Appropriate perception of cellulose outside the cell by transforming it into an intracellular signal ensures the rapid production of cellulases by cellulolytic Hypocrea jecorina. The major extracellular ␤-glucosidase BglI (CEL3a) has been shown to contribute to the efficient induction of cellulase genes. Multiple ␤-glucosidases belonging to glycosyl hydrolase (GH) family 3 and 1, however, exist in H. jecorina. Here we demonstrated that CEL1b, like CEL1a, was an intracellular ␤-glucosidase displaying in vitro transglycosylation activity. We then found evidence that these two major intracellular ␤-glucosidases were involved in the rapid induction of cellulase genes by insoluble cellulose. Deletion of cel1a and cel1b significantly compromised the efficient gene expression of the major cellulase gene, cbh1. Simultaneous absence of BglI, CEL1a, and CEL1b caused the induction of the cellulase gene by cellulose to further deteriorate. The induction defect, however, was not observed with cellobiose. The absence of the three ␤-glucosidases, rather, facilitated the induced synthesis of cellulase on cellobiose. Furthermore, addition of cellobiose restored the productive induction on cellulose in the deletion strains. The results indicate that the three ␤-glucosidases may not participate in transforming cellobiose beyond hydrolysis to provoke cellulase formation in H. jecorina. They may otherwise contribute to the accumulation of cellobiose from cellulose as inducing signals.

bLactose (1,4-O-␤-D-galacto-pyranosyl-D-glucose) induces cellulolytic enzymes in Trichoderma reesei and is in fact one of the most important soluble carbon sources used to produce cellulases on an industrial level. The mechanism underlying the induction is, however, not fully understood. In this study, we investigated the cellular functions of the intracellular ␤-glucosidases CEL1a and CEL1b in the induction of cellulase genes by lactose in T. reesei. We demonstrated that while CEL1a and CEL1b were functionally equivalent in mediating the induction, the simultaneous absence of these intracellular ␤-glucosidases abolished cbh1 gene expression on lactose. D-Galactose restored the efficient cellulase gene induction in the ⌬cel1a strain independently of its reductive metabolism, but not in the ⌬cel1a ⌬cel1b strain. A further comparison of the transcriptional responses of the ⌬cel1a ⌬cel1b strain complemented with wild-type CEL1a or a catalytically inactive CEL1a version and the ⌬cel1a strain constitutively expressing CEL1a or the Kluyveromyces lactis ␤-galactosidase LAC4 showed that both the CEL1a protein and its glycoside hydrolytic activity were indispensable for cellulase induction by lactose. We also present evidence that intracellular ␤-glucosidase-mediated lactose induction is further conveyed to XYR1 to ensure the efficiently induced expression of cellulase genes.C ost-effective conversion of plant cell wall-derived polysaccharides holds the potential for production of an environmentally clean and renewable source of energy and platform chemicals (1). Trichoderma reesei (teleomorph Hypocrea jecorina) is well known for its high capacity to secrete large amounts of lignocellulosic enzymes that release fermentable sugars and has thus been developed into one of the most prolific industrial cellulase producers. High-yield production of the bulk of the plant cell walldegrading machinery in T. reesei is, however, dependent on induction by insoluble substrates that include cellulose, hemicellulose, and mixtures of plant polymers. Considering the ease of manipulation and the complication of separating enzymes from insoluble plant cell wall materials, soluble inducing substrates are usually preferred or required (2). Among others, the disaccharide lactose (1,4-O-␤-D-galacto-pyranosyl-D-glucose) is an important and economic soluble carbon source for cellulase production by T. reesei. However, the induced cellulase yields on lactose are usually lower than those on cellulose (3, 4). Understanding the differences in the inducing efficiency and the mode by which lactose triggers cellulase formation would be helpful for improving the performance of industrial strains.In fungi, catabolism of lactose is thought to proceed either by extracellular hydrolysis and subsequent uptake of the resulting sugar monomers or by uptake of the disaccharide followed by intracellular hydrolysis (4). For T. reesei, it has been assumed that lactose metabolism relies on the first strategy, based on several findings, including the absence of appar...