Coprinopsis cinerea, 亦稀灰蓋鬼傘，是研究擔子菌發育過程的模範生物。它的生命週期短，容易培養，亦有已建立的遺傳和分子生物研究技術。最近，它的完整基因組序列亦被發表。C. cinerea 的子實體萌生和發展是快速而複雜的過程。它受多種因素影響，如交配基因，營養消耗，光照和溫度。然而，我們對於C. cinerea 出菇的基本機制和涉及的分子途徑仍不清楚。在這項研究中，我採用NimbleGen 微陣列，以了解13,320 個C. cinerea 基因模型的表達。此微陣列覆蓋不同發育階段，包括雙核菌絲體，子實體初體，第2階段原基體，年青和成熟子實體。11,815個預測，在至少一個發育階段表達。707個基因模型在出菇的萌生過程有差異表達。我發現一些可能參與子實體的萌生和發展的份子。它們可能在檢測養分、形態、信號轉導和應激反應方面，擔當重要角色。此外，我亦分析了轉錄因子 、蛋白激酶組和細胞色素P450的基因表達模式。 / C. cinerea 的子實體發展是與光暗週期同步。當子實體初體在沒有光的環境培植，使會形成dark stipe。101個基因在dark stipe 表達差異。它們可能參與原基體成熟的過程。此微陣列基因表達數據，對了解菇機制有價格的信息。 / 胞吞作用是真核細胞透過質膜內陷將外來物質攝取的過程。Rab5 和Rab7 分別控制早期和晚期的胞吞作用。C. cinerea 的胞吞作用是組織由110個基因組成。FM4-64 螢光顯示在C. cinerea 菌絲體的胞吞作用是依賴肌動蛋白和能源。從菌絲體到年青子實體，Cc. Rab5的表達維持相同水平，而Cc.Rab7 的表達則不斷增加，兩者在成熟子實體的表達都是最高，原位雜交體技術顯示 Cc.Rab5 和Cc.Rab7 的 mRNA在年青子實體的子實層以及整個子實層上菌摺表達。我在第2階段原基體進行RNA 乾擾，致使Cc.Rab5 和Cc.Rab7的基因表達敲落。這導致原基體的生長遲緩。及後形成的成熟子實體亦有異常形態。因此，我推測Cc. Rab5 和Cc.Rab7參與出菇過程，並影響擔孢子的形成。這些結果表明，胞吞作用在C.Cinerea 子實體發育過程中發揮定一定的作用。 / Coprinopsis cinerea, is a model organism for studying developmental processes in basidiomycetous fungi. It has a short life cycle, easy to be cultivated in laboratory and can be accessed by various genetic and molecular techniques. Recently, its complete genome sequence was released. The fruiting body development in C. cinerea is a rapid yet complicated process. It is under the regulation of various factors such as mating type genes, nutrients depletion, light and temperature. However, the underlying mechanism and molecular events involved during fruiting body initiation and development remains unclear. / In this study, fruiting body developmental stages including mycelium, fruiting initials, stage 2 primordium, young and mature fruiting body, were analyzed with a comprehensive NimbleGen microarray. 11,815 out of 13,320 predicted gene models were expressed in at least one of the stages. 707 genes were differentially expressed during fruiting body initiation. Potential players involved in nutrients sensing, morphogenesis, signaling pathways and stress response were identified. In particular, expression patterns of all transcription factors, kinome and cytochrome P450s were analyzed. / The fruiting body development of C. cinerea is synchronized with the light/dark cycle. Differentially expressed genes were found in dark stipe produced by keeping fruiting initials in complete darkness. 101 genes, which are likely to be involved in maturation of primordium were identified. / Endocytosis is an essential process in eukaryotes through which cells take up extracellular substrates by membrane invaginations. Rab5 and Rab7 control the early and late stage of endocytosis respectively. The C. cinerea endocytic machinery composed of 110 genes models. The endocytic pathway was traced by FM4-64 and was found to be actin- and energy-dependent. Temporal and spatial expressions of Cc.Rab5 and Cc.Rab7 during fruiting body development were studied. Cc.Rab5 expressed constitutively from mycelium to young fruiting body stage, and reached the highest in the mature fruiting body. The expression of Cc.Rab7 increased continually from mycelium to mature fruiting body stage. From the in situ RNA-RNA hybridization results, both transcripts were localized at the hymenium layer in the young fruiting body and throughout the gill tissue of the mature cap. Knock-down of Cc.Rab5 and Cc.Rab7 by siRNA resulted in retarded growth of the stage 2 primordium and abnormal mature fruiting body. Cc.Rab5 and Cc.Rab7 may be involved in the formation of basidiospores. Endocytosis may play some roles during fruiting body development in C. cinerea. / Detailed summary in vernacular field only. / Detailed summary in vernacular field only. / Detailed summary in vernacular field only. / Lee, Yung Yung. / Thesis (Ph.D.)--Chinese University of Hong Kong, 2012. / Includes bibliographical references (leaves 156-179). / Electronic reproduction. Hong Kong : Chinese University of Hong Kong, [2012] System requirements: Adobe Acrobat Reader. Available via World Wide Web. / Abstract also in Chinese. / Abstract --- p.i / 論文摘要 --- p.iii / Abbreviations --- p.iv / Acknowledgements --- p.v / Table of Contents --- p.vi / List of Figures --- p.x / List of Tables --- p.xiii / Chapter Chapter 1 --- Literature Review --- p.1 / Chapter 1.1 --- Importance of fruiting body (mushrooms) production --- p.1 / Chapter 1.2 --- Introduction on Coprinopsis cinerea --- p.1 / Chapter 1.2.1 --- General introduction --- p.1 / Chapter 1.2.2 --- Life cycle and morphology --- p.2 / Chapter 1.2.3 --- Growth conditions for C. cinerea --- p.4 / Chapter 1.3 --- Regulation of fruiting development in C. cinerea --- p.5 / Chapter 1.3.1 --- Regulation by mating types --- p.5 / Chapter 1.3.2 --- Regulation by light-dark cycle --- p.6 / Chapter 1.3.3 --- Regulation by physiological factors --- p.9 / Chapter 1.4 --- Fruiting-specific genes --- p.9 / Chapter 1.5 --- Genome project of C. cinerea --- p.10 / Chapter 1.6 --- Transformation and gene silencing in C. cinerea --- p.11 / Chapter 1.7 --- DNA microarray --- p.12 / Chapter 1.8 --- Endocytosis --- p.13 / Chapter 1.8.1 --- The endocytic pathway --- p.14 / Chapter 1.8.2 --- Rab GTPase --- p.17 / Chapter 1.8.2.1 --- Control of the active and inactive state of Rab proteins --- p.18 / Chapter 1.8.2.2 --- Functions of Rab GTPases in vesicular transport --- p.19 / Chapter 1.8.2.3 --- Rab5 and Rab7 GTPase --- p.20 / Chapter 1.8.3 --- Endocytosis in fungi --- p.21 / Chapter 1.9 --- Aims of project --- p.22 / Chapter Chapter 2 --- Whole genome expression analysis during fruiting body development --- p.25 / Chapter 2.1 --- Introduction --- p.25 / Chapter 2.2 --- Materials and Methods --- p.26 / Chapter 2.2.1 --- NimbleGen 12x135K gene expression microarray --- p.26 / Chapter 2.2.1.1 --- Strains and culture conditions --- p.26 / Chapter 2.2.1.2 --- RNA extraction --- p.27 / Chapter 2.2.1.3 --- Overall design of the NimbleGen custom Microarray --- p.28 / Chapter 2.2.1.4 --- Microarray hybridization, data extraction and normalization --- p.28 / Chapter 2.2.2 --- Microarray data analysis, clustering and GO assignment --- p.29 / Chapter 2.2.3 --- Validation of expression patterns of NimbleGen microarray and analysis of differentially expressed genes by quantitative real-time PCR --- p.29 / Chapter 2.2.3.1 --- cDNA synthesis --- p.29 / Chapter 2.2.3.2 --- Primer design and verification --- p.30 / Chapter 2.2.3.3 --- Real time PCR and data analysis --- p.32 / Chapter 2.3 --- Results --- p.33 / Chapter 2.3.1 --- Whole-genome expression during fruiting body development --- p.33 / Chapter 2.3.2 --- Differentially expressed genes during fruiting body initiation --- p.48 / Chapter 2.3.2.1 --- Fruiting body initiation-specific genes --- p.52 / Chapter 2.3.3 --- Gene expression analysis during fruiting body development --- p.53 / Chapter 2.3.4 --- The C. cinerea kinome --- p.55 / Chapter 2.3.5 --- Transcription factors in C. cinerea --- p.60 / Chapter 2.3.6 --- The cytochrome P450 family in C. cinerea --- p.65 / Chapter 2.3.7 --- Validation of NimbleGen microarray data by quantitative real-time PCR --- p.68 / Chapter 2.4 --- Discussion --- p.77 / Chapter Chapter 3 --- Effect of light on gene expression of fruiting body development --- p.92 / Chapter 3.1 --- Introduction --- p.92 / Chapter 3.2 --- Materials and Methods --- p.93 / Chapter 3.2.1 --- NimbleGen 12x135K gene expression microarray --- p.93 / Chapter 3.2.2 --- Validation of expression patterns of NimbleGen microarray and analysis of differentially expressed genes by quantitative real-time PCR --- p.93 / Chapter 3.2.2.1 --- cDNA synthesis --- p.94 / Chapter 3.2.2.2 --- Primer design and verification --- p.94 / Chapter 3.3 --- Results --- p.95 / Chapter 3.3.1 --- Differentially expressed genes in dark stipes --- p.95 / Chapter 3.3.2 --- Validation of expression patterns of NimbleGen microarray by real-time PCR --- p.102 / Chapter 3.4 --- Discussion --- p.107 / Chapter Chapter 4 --- Endocytosis in C. cinerea and its role in fruiting body development --- p.111 / Chapter 4.1 --- Introduction --- p.112 / Chapter 4.2 --- Materials and Methods --- p.112 / Chapter 4.2.1 --- The endosomal machinery of C. cinerea --- p.112 / Chapter 4.2.2 --- Tracing the endocytic pathway sing FM-64 --- p.113 / Chapter 4.2.2.1 --- Strains and culture conditions --- p.113 / Chapter 4.2.2.2 --- FM4-64 internalization in mycelium of C. cinerea --- p.113 / Chapter 4.2.2.3 --- Drug treatment effect on the internalization of FM4-64 dye --- p.114 / Chapter 4.2.3 --- Temporal and spatial expression of Cc.Rab5 and Cc.Rab7 --- p.114 / Chapter 4.2.3.1 --- Cloning of Cc.Rab5 and Cc.Rab7 --- p.114 / Chapter 4.2.3.1.1 --- RNA extraction and cDNA synthesis --- p.114 / Chapter 4.2.3.1.2 --- TA cloning of amplification products and bacterial transformation --- p.115 / Chapter 4.2.3.1.3 --- PCR screening for positive transformants and sequencing --- p.115 / Chapter 4.2.3.2 --- Quantitative real-time PCR --- p.116 / Chapter 4.2.3.2.1 --- RNA extraction and cDNA synthesis --- p.116 / Chapter 4.2.3.2.2 --- Primer design and verification --- p.116 / Chapter 4.2.3.2.3 --- Real time PCR and data analysis --- p.117 / Chapter 4.2.3.3 --- In situ RNA-RNA hybridization --- p.117 / Chapter 4.2.3.3.1 --- Tissue preparation --- p.117 / Chapter 4.2.3.3.2 --- RNA probe synthesis --- p.117 / Chapter 4.2.3.3.3 --- Hybridization, signal development and image viewing --- p.118 / Chapter 4.2.4 --- Knock-down of endogenous Cc.Rab5 and Cc.Rab7 by siRNA --- p.119 / Chapter 4.2.4.1 --- Strains and culture conditions --- p.119 / Chapter 4.2.4.2 --- Production of dsRNA of Cc.Rab5 and Cc.Rab7 --- p.119 / Chapter 4.2.4.3 --- Digestion of dsRNA to give siRNA --- p.120 / Chapter 4.2.4.4 --- Effects of Cc.Rab5 and Cc.Rab7 siRNA on fruiting body development --- p.120 / Chapter 4.2.4.4.1 --- Application of siRNA to C. cinerea culture --- p.120 / Chapter 4.2.4.4.2 --- Validation of the knock-down efficacy by real-time PCR --- p.121 / Chapter 4.3 --- Results --- p.122 / Chapter 4.3.1 --- The endosomal machinery of C. cinerea --- p.122 / Chapter 4.3.2 --- The endocytic pathway of C. cinerea --- p.127 / Chapter 4.3.2.1 --- Time-course of FM4-64 internalization --- p.127 / Chapter 4.3.2.2 --- Validation of active transport of FM4-64 --- p.129 / Chapter 4.3.3 --- Cloning of Cc.Rab5 and Cc.Rab7 --- p.131 / Chapter 4.3.4 --- Temporal expression of Cc.Rab5 and Cc.Rab7 --- p.133 / Chapter 4.3.5 --- Spatial expression of Cc.Rab5 and Cc.Rab7 --- p.136 / Chapter 4.3.6 --- Effects of Cc.Rab5 and Cc.Rab7 knock-down by siRNA --- p.140 / Chapter 4.3.6.1 --- Observation of effect of siRNA on fruiting body development --- p.140 / Chapter 4.3.6.2 --- Validation of the efficacy of external application of siRNA --- p.143 / Chapter 4.4 --- Discussion --- p.146 / Chapter Chapter 5 --- Concluding remarks --- p.152 / References --- p.156 / Appendix --- p.180

Thesis (M.S.)--University of Tennessee, Knoxville, 2002. / Title from title page screen (viewed on Sept. 25, 2002). Thesis advisor: Karen W. Hughes. Document formatted into pages (vii, 58 p. : ill. (some col.)). Vita. Includes bibliographical references (p. 51-55).

No description available.

The A mating type factor of the hymenomycete fungus Coprinus cinereus is a multi-allelic gene complex that controls mating compatibility and sexual development. It contains up to four pairs of specificity genes, the a, b, c, and d gene-pairs. Each gene-pair codes for two homeodomain transcription factors with distinct classes of homeodomain motifs. Mating compatibility between the A42 and A6 factors depends solely on the different alleles of the b gene-pair, b1-1 and b2-1 in A42 and b1-3 and b2-3 in A6. The b1-3 and b2-3 genes of A6 were isolated and the complete DNA sequences of genomic and cDNA clones were determined. Construction of chimeric genes using the A42 and A6 b genes identified the N-terminal regions of the A proteins as being responsible for allele specificity. Analysis of protein-protein interactions showed that b1 and b2 proteins from different alleles of the same gene-pair can dimerise, whereas proteins from the same allele pair can not. It was shown that a region of 90 amino acids at the N-terminus of the b2-3 protein is sufficient for dimerisation with b1-1. This region is predicted to contain an amphipathic helix. A comparison with the equivalent region in the b2-1 protein identifies a similar helix. This suggests that a compatible A mating type reaction and thus allele specificity is recognised by the ability to dimerise through this domain. Polyclonal antibodies were raised against the b1-1 protein and a heterologous yeast expression system was established for testing potential DNA target sites of the b1-1 and b2-3 proteins, both techniques offering potentially useful tools for further molecular analysis of the A mating type proteins.

Thesis (M.S.)--Oregon State University, 1988. / Typescript (photocopy). Includes bibliographical references (leaves 63-65). Also available on the World Wide Web.

Thesis (Ph. D.)--Oregon Graduate Institute of Science and Technology, 1990.

Thesis (Ph. D.)--Illinois State University, 1972. / Title from title page screen, viewed Sept. 28, 2004. Dissertation Committee: Anthony E. Liberta (chair), H.E. Brockman, H.W. Huizinga, D.F. Weber, E.I. Rhymer. Includes bibliographical references (leaves 157-160) and abstract. Also available in print.

No description available.

Submitted by Isaac Francisco de Souza Dias (isaac.souzadias@ufpe.br) on 2016-06-30T18:49:42Z No. of bitstreams: 2 license_rdf: 1232 bytes, checksum: 66e71c371cc565284e70f40736c94386 (MD5) Dissertação Carla Lira.pdf: 1678207 bytes, checksum: bd79649b073f38b1abb20903bb40fe70 (MD5) / Made available in DSpace on 2016-06-30T18:49:42Z (GMT). No. of bitstreams: 2 license_rdf: 1232 bytes, checksum: 66e71c371cc565284e70f40736c94386 (MD5) Dissertação Carla Lira.pdf: 1678207 bytes, checksum: bd79649b073f38b1abb20903bb40fe70 (MD5) Previous issue date: 2012-02-15 / CAPES / Os representantes da classe Agaricomycetes (filo Basidiomycota) são caracterizados por desenvolverem basidiomas onde produzem basídios e basidiosporos. Grande parte dos representantes deste grupo degrada componentes da madeira, sendo assim chamados de lignolíticos ou lignocelulolíticos, mas também agem como parasitas obrigatórios ou facultativos e como simbiontes. Para ampliar o conhecimento sobre a diversidade taxonômica e ecológica desse grupo de fungos do sertão de Pernambuco, foram realizadas seis coletas em duas áreas em estágio severo e moderado de desertificação em Cabrobó [Barro Branco (CBB) e Fazenda Mosquito (CFM), respectivamente] e um brejo de altitude em Triunfo [Sítio Carro Quebrado (SCQ)], totalizando 18 expedições. Para avaliar a diversidade e as relações desse grupo de fungos com os hospedeiros nas áreas de estudo, foram utilizados testes de χ2 e ANOSIM. No presente tudo, foram coletados 256 espécimes correspondendo a 47 espécies de Agaricomycetes lignocelulolíticos (12 em CBB, 19 em CFM e 41 em SCQ). Dentre estas espécies, há 31 novos registros para o estado de Pernambuco, região Nordeste, Caatinga de Pernambuco, bioma Caatinga, Brasil e América do Sul. Após 18 visitas a campo, as curvas cumulativas de espécies não se estabilizaram, indicando que mais coletas são necessárias. De acordo com a ANOSIM, há diferença significativa na composição de espécies entre as duas áreas de caatinga e brejo de altitude, assim como para abundância de acordo com o teste de χ2, sendo coletado um maior número de espécimes em SCQ. Entretanto, de acordo com este teste, não há diferença na riqueza entre as áreas. Não há diferença também na composição e na riqueza de espécies entre os períodos de coleta (sazonalidade), de acordo com a ANOSIM e com o teste de χ2, respectivamente. Já a abundância de espécies nessas áreas foi diferente quando utilizado o teste de χ2, sendo coletados mais espécimes na estação seca. A tendência em ocorrer em substratos mortos foi observada para Agaricomycetes lignocelulolíticos em todas as áreas de estudo. Entretanto, em áreas de caatinga sob processos de desertificação, os táxons de Hymenochaetales tendem a acompanhar a disponibilidade de substrato, ocorrendo preferencialmente em árvores vivas. Phellinus rimosus foi recorrente em Caesalpinia pyramidalis em CBB e específico no mesmo hospedeiro em CFM. / The members of Agaricomycetes (phylum Basidiomycota) are characterized by developing basidiomata in which basidia and basidiospores are produced. Most representatives of this group degrade wood components, the so called lignolytic or lignocellulolytic, but also act as facultative and obligate parasites or as symbionts. To increase the knowledge about the taxonomic and ecological diversity of this group of fungi in the semi-arid of Pernambuco state, six collections were undertaken in areas in moderate and severe stage of desertification in Cabrobó [Barro Branco (CBB) and Fazenda Mosquito (CFM), respectively] and an upland forest in Triunfo [Sítio Carro Quebrado (SCQ)], in a total of 18 expeditions. The χ2 test and the ANOSIM were used to assess the diversity and relationships of this group of fungi with the hosts in the study areas. The 256 specimens collected corresponded to 47 species of lignocellulolytic Agaricomycetes (12 in CBB, 19 in CFM and 41 in SCQ). Among these species, there are 31 new records for the Pernambuco state, for the Northeast, Caatinga of Pernambuco, Caatinga biome, Brazil and South America. After 18 field trips, the cumulative curves of species were not stabilized, indicating that the number of collections needs to be increase. According to the ANOSIM, there are significant difference in the species composition among the two areas of caatinga and the montane forest. The same was observed for abundance according to the χ2 test, more specimens being collected in SCQ. However the is no difference in richness among the areas according to this test. Also, there are no differences in species composition and richness among sampling periods (seasonality), according to the ANOSIM and the χ2 test, respectively. The abundance of the species was different according to the χ2 test, more specimens being collected during the dry season. The tendency to occur on dead substrates was found in lignocellulolytic Agaricomycetes. However, in areas of caatinga in different processes of desertification, taxa of Hymenochaetales tended to follow the availability of substrate, preferably occurring on living trees. Phellinus rimosus was recurrent on Caesalpinia pyramidalis in CBB and specific on the same host in CFM.

In the first of two parts of the investigation, the stimulation of Armiil aria mellea (Fr.) Quél., a common root-rotting fungus, by Aureobastdium pullulans (DeBary) Arnaud was studied. A. pullulans is an ubiquitous imperfect fungus, inhabiting soil and other substrates. It was shown to produce in culture a volatile, heat stable, neutral substance which stimulated the mycelial development, rhizomorph initiation and rhizomorph elongation of A. mellea. A. pullulans is the only microorganism reported so far to stimulate the growth of A. mellea. The effect of ethanol on the growth of A. mellea was similar to the effect of a cell-free filtrate from a liquid culture of A. pullulans. The cell-free filtrate was shown by gas chromatography to contain ethanol. Ethanol supplied daily as 50 ppm in a glucose-asparagine medium resulted in a ten-fold increase in the number of rhizomorphs produced by A. mellea. The effects of 3-indolylacetic acid, ɣ-(indole-3)-n-butyric acid, the sodium salt of 2,4-dichlorophenoxyacetic acid and tryptophane on the growth of A. mellea were tested, but the stimulatory effect of A. pullulans was not reproduced by them. One rhizomorph tip of A. mellea could develop an extensive rhizomorph system in autoclaved soil if the stimulatory substance produced by A. pullulans was available. In the second part of the investigation the effect of soil moisture and the related effect of some soil-inhabiting Fungi imperfecti on the spread through soil of the root-rotting Basidiomycete Coniopnora puteana (Schum. ex Fr.) Karst.were studied. A soil moisture level of 20 - 25% saturation was satisfactory for the growth of C. puteana in non-sterile soil. At this moisture level C. puteana was able to grow out from a small alder disc inoculum in the centre of a petri dish and invade alder discs at the periphery of the dish. In wetter conditions (50% saturation and higher) it was unable to grow out into the soil. In autoclaved soil the cptimum moisture level for the growth of C. puteana was between 75% and 100% saturation. Small amounts of non-sterile soil were added to autoclaved soil at different moisture levels, with an effect on the growth of C. puteanas similar to that of completely non-sterile soil. Trichoderma viride Pers. ex Fr., a known antibiotic producer, was inoculated in autoclaved soil and produced a greater inhibition of C. puteana in the wetter treatments than in drier ones. The inhibitory effect of Acti-dione (cycloheximide), an antifungal antibiotic active in the pH range 3-5, was examined in non-sterile soil end in autoclaved soil. The same concentration of antibiotic resulted in a greater inhibition of the growth of C. puteana at the higher moisture levels than at the lower ones. / Science, Faculty of / Botany, Department of / Graduate