Program Book - 27th Fungal Genetics Conference

FULL POSTER SESSION ABSTRACTSand aggressiveness, isolates were characterized for in vitro and in planta phenotypes. For the former, mycelial abundance, colony diameter, pigmentation,and sporulation were rated in replicated trials. For the latter, incubation period, primary diseased leaf area and a qualitative differential score were ratedon maize near isogenic lines with and without Ht2. Linkage mapping identified a 54.3 kb sequence of Et28A as a robust candidate region carrying S. turcicaavrHt2. In order to manage the vast amount of genotypic and phenotypic data a MySQL database was created.507. The secretome is linked to virulence in the yeast pathogen Cryptococcus. Leona Campbell 1 , Anna Simonin 1 , Janna Ferdous 1 , Matthew Padula 1 ,Elizabeth Harry 2 , Ben Herbert 3 , Dee Carter 1 . 1) School of Molecular Bioscience, University of Sydney, Sydney, N.S.W. Australia; 2) iThree Institute,University of Technology, Sydney, NSW, Australia; 3) Department of Chemistry & Biomolecular Sciences, Macquarie University, Sydney, NSW, Australia.The disease caused by pathogenic Cryptococcus spp. begins after inhalation of infectious propagules leading to infection of the lung. In some cases thepathogen disseminates to the central nervous system, resulting in meningoencephalitis, which can be fatal if left untreated. However, closely relatedstrains of Cryptococcus gattii and its sibling species C. neoformans can exhibit significantly different degrees of pathogenesis in the mammalian host. Asfungi utilize absorptive nutrition producing a range of secreted degradative enzymes, and as these may invoke a host response, the fungal secretome islikely to be very important in modulating host-pathogen interactions. To investigate this the secreteome was determined for a hypovirulent and ahypervirulent strain of C. gattii, R272 and R265 respectively, and a virulent strain of the opportunistic pathogen C. neoformans, KN99a. All strains weregrown under conditions designed to mimic those encountered in vivo. Secreted proteins were analysed using two different mass spectrometry-basedtechniques: 1D nanoLC-MS/MS and Imaging Mass Spectrometry (IMS). The three strains secreted significantly different protein cohorts. A total of 70proteins were identified with 47, 13 and 22 identified from R272, R265 and KN99a respectively. Only one protein was shared by all strains, a putativeglycosyl hydrolase. The secretomes of R265 and KN99a primarily included uncharacterized proteins, and bioinformatic analysis suggested these proteinscontained catalytic regions with roles in carbohydrate degradation. In contrast the less virulent R272 strain secreted a more diverse set of proteinsincluding canonical cytosolic proteins such as enolase and transaldolase. These proteins have been described as fungal allergens that bind IgE. Thesefindings indicate that virulence and the secretome are linked in Cryptococcus. The secretion of proteins with a putative role in nutrient scavenging byvirulent strains R265 and KN99a suggest they can source nutrients from a range of available substrates, which may allow them to exploit a wider range ofecological niches including the mammalian host. In contrast, the potentially allergenic proteins secreted by strain R272 suggest this strain triggers a moreeffective immune response, leading to clearance of the pathogen.508. Post-Transcriptional Regulation of the ER Stress Response in Cryptococcus neoformans. Virginia E. Havel, John C. Panepinto. Microbiology andImmunology, University at Buffalo, Buffalo, NY.Cryptococcus neoformans is one of a small number of fungi able to make the transition from ambient environmental temperatures to human core bodytemperature. We have previously reported that the ER stress response plays an important role during host temperature adaptation. Deletion of the RNAbinding protein, Puf4, results in temperature sensitivity and increased resistance to the ER stress inducing drug tunicamycin, leading us to hypothesize thatPuf4 post-transcriptionally regulates the ER stress response during host temperature adaptation. The ER stress response is initiated by the transcriptionfactor Hac1 (Hxl1 in C. neoformans). Hac1 translation requires unconventional splicing of the pre-spliced HAC1 mRNA to the translated HAC1 mRNA at theER surface by Ire1. Time courses measuring HXL1 mRNA splicing during a shift to 37°C demonstrate a delay in the splicing of HXL1 in puf4D when comparedto wild type. The delay in HXL1 splicing in puf4D results in a delayed and persistent induction of ER stress response transcripts in puf4D compared to wildtype as measured by northern blot. We hypothesize that Puf4 is required for localization of the HXL1 transcript to the ER outer surface where it is cleavedby Ire1, thereby promoting the induction of the ER stress response. We have also shown through EMSA analysis and RNA-immunoprecipitationexperiments that Puf4 is able to bind to ALG7 mRNA. Alg7 is involved in protein glycosylation at the ER surface and is the target of tunicamycin. Based onthe observation that puf4D has increased resistance to tunicamycin, and results demonstrating that Puf4 is able to bind ALG7 mRNA we hypothesize thatPuf4 may regulate ALG7 mRNA by repressing translation. In our model Puf4 has a bi-modal mechanism of regulating the ER stress response. ER stressresponse initiation requires Puf4-mediated localization of pre-spliced HXL1 mRNA to the ER surface. During the attenuation phase of the ER stressresponse, we hypothesize that ER transcripts are translationally repressed by Puf4, resulting in attenuation of the ER stress response and allowing the cellto return to homeostasis. Despite the well-studied mechanism of unconventional splicing by Hac1 and Hac1 homologs in yeast and other model systems,this study is the first to identify a RNA binding protein potentially involved its activation.509. A morphogenesis regulator controls cryptococcal neurotropism. Xiaorong Lin 1 , Bing Zhai 1 , Karen Wozniak 2 , Srijana Upadhyay 1 , Linqi Wang 1 , ShupingZhang 3 , Floyd Wormley 2 . 1) Biology, Texas A&M University, TAMU-3258, TX; 2) Biology, the University of Texas at San Antonio, San Antonio, Texas, USA; 3)Veterinary Medicine and Biomedical Sciences, Texas A&M University, College Station, Texas, USA.Cryptococcus neoformans is the major causative agent of cryptococcal meningitis, a disease that is responsible for more than 600,000 deaths each year.This ubiquitous environmental pathogen enters host lungs through inhalation and typically establishes asymptomatic latent infections. However,extrapulmonary dissemination often occurs in individuals with weakened immunity and Cryptococcus has a predilection to infect the brain. Braininfections are fatal and formidable to treat due to the poor penetration of most antifungals to the brain. Unfortunately, little is known about cryptococcalfactors that control its neurotropism. Here we report that a morphogenesis regulator Znf2 controls the tissue tropism of cryptococcal infection. Inparticular, activation of Znf2 abolishes Cryptococcus extrapulmonary dissemination and consequently leads to the absence of fatal brain infections in theinhalation infection model. Although Znf2 overexpression strains are avirulent in this animal model, these strains are capable of proliferating in the animallungs during the early stages of infections. Histological examinations and cytokine profiling revealed that the Znf2 overexpression strain causes enhancedmonocyte infiltration in the animal lungs. Consistently, the Znf2 overexpression strain stimulates pro-inflammatory host responses while suppressesdeleterious Th2 host responses during early stage of infection in the pulmonary infection model. Such protective host defense responses might haveprevented the extrapulmonary dissemination of Cryptococcus. In the intravenous infection model where the lung infection was bypassed and there wasuniform hematogenous dissemination, the Znf2 overexpression strain showed a specific defect in the brain infection. Taken together, our data indicatethat Znf2 helps polarize the host immunity towards protection and that it mediates cryptococcal tissue tropism during infection.510. Extracellular and intracellular signaling orchestrates morphotype-transition and virulence in human pathogen Cryptococcus neoformans. LinqiWang, Xiuyun Tian, Rachana Gyawali, Xiaorong Lin. Biology, College Station, TX.Interactions with the environment and divergent species drive the evolution of microbes. To sense and rapidly respond to these dynamic interactions,“simple” microbes developed bet-hedging social behaviors, including the construction of heterogeneous biofilm communities and transition betweendifferent morphotypes. The human fungal pathogen Cryptococcus neoformans can undergo morphotype transition between the yeast and the filamentousform. Most recently, we demonstrated that the zinc-finger regulator Znf2 bridges the bi-direction yeast-hypha transition and virulence in this pathogen.246