Expression analysis of the fungus Ustilago maydis WT strain FB2 (a2b2) at different pH values
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ABSTRACT: Elicidate the changes of the transcriptional program response of U. maydis WT strain (a2b2) when transfered from neutrality to acidic or alkaline medium The mutants strains CL211 and GP25 used as controls as well as the WT strain are described in Martínez-Espinoza et al., 1997; 2004;González-Prieto et al., 2014 and Banuett & Herskowitz, 1989 respectively.
Project description:Elicidate the changes of the transcriptional program response of U. maydis WT strain (a2b2) when transfered from neutrality to acidic or alkaline medium The mutants strains CL211 and GP25 used as controls as well as the WT strain are described in MartÃnez-Espinoza et al., 1997; 2004;González-Prieto et al., 2014 and Banuett & Herskowitz, 1989 respectively. High density-one color arrays from Nimblegen were used according to a design from Scott Gold from University of Georgia. The design has five different oligonucleotides of 60 bp in lenght in duplicate, representing each one of the 6883 gene of the U. maydis genome.
Project description:Study of gene regulation basidiocarps development in Ustilago maydis using transcriptomic analysis. In 2012, Cabrera-Ponce et al. established conditions allowing a completely different developmental program in U. maydis when grown on solid medium containing Dicamba (synthetic auxin) in dual cultures with maize embryogenic calli.
Project description:Fungal pathogens depend on sophisticated gene expression programs for successful infection. A crucial component is RNA regulation mediated by RNA-binding proteins. In the biotrophic fungal plant pathogen Ustilago maydis, loss of the multi KH domain containing RBP, Khd4, causes aberrant cell morphology, disturbed hyphal growth and impaired virulence. To investigate the role of Khd4 in the polar growth of infectious hyphae, we established the RNA-editing based hyperTRIBE method to detect in vivo mRNA targets.This technique is especially suited to investigate proteins of high molecular weight or low expression that are difficult to purify (McMahon et al., 2016, Xu et al., 2018, Rahman et al., 2018). HyperTRIBE exploits the RNA editing activity of the catalytic domain of ADAR (Adenosine Deaminase Acting on RNA) from D. melanogaster that additionally carries the mutation E488Q to increase editing (Kuttan and Bass, 2012; designated Ada). We fused the codon-optimized catalytic Ada domain with the green fluorescent protein (see Materials and methods; enhanced version, designated Gfp, Clontech) to the C terminus of Khd4 (Khd4-Ada-Gfp). As a control for background editing, we used the strain expressing Ada-Gfp protein with N-terminal Kat fusion in the wildtype background (control-Ada).
Project description:Fungal pathogens depend on sophisticated gene expression programs for successful infection. A crucial component is RNA regulation mediated by RNA-binding proteins. In the biotrophic fungal plant pathogen Ustilago maydis, loss of the multi KH domain containing RBP, Khd4, causes aberrant cell morphology, disturbed hyphal growth and impaired virulence. To investigate the role of Khd4 in the polar growth of infectious hyphae, we established the RNA-editing based hyperTRIBE method to detect in vivo mRNA targets.This technique is especially suited to investigate proteins of high molecular weight or low expression that are difficult to purify (McMahon et al., 2016, Xu et al., 2018, Rahman et al., 2018). HyperTRIBE exploits the RNA editing activity of the catalytic domain of ADAR (Adenosine Deaminase Acting on RNA) from D. melanogaster that additionally carries the mutation E488Q to increase editing (Kuttan and Bass, 2012; designated Ada). We fused the codon-optimized catalytic Ada domain with the green fluorescent protein (see Materials and methods; enhanced version, designated Gfp, Clontech) to the C terminus of Khd4 (Khd4-Ada-Gfp). As a control for background editing, we used the strain expressing Ada-Gfp protein with N-terminal Kat fusion in the wildtype background (control-Ada). We then performed differential gene expression analysis to determine the pathological consequence of dysregulated Khd4-mediated mRNA regulation.
Project description:In the phytopathogenic Basidiomcete Ustilago maydis, sexual and pathogenic development are tightly connected and controlled via the b-mating type locus. The b-mating type locus encodes two homeodomain transcription factors, bE and bW, which form an active heterodimeric complex when they are derived from different b-alleles. Rbf1 encodes a zinc-finger transcription factor that is expressed upon formation of an active bE/bW heterodimer. To analyse the dependency of b and rbf1, changes in gene expression were monitored in strain AB31 (Brachmann et al., 2001, Mol Microbiol 42, 1047-1063) and in the derivative AB31Δrbf1 in which rbf1 was deleted. AB31 harbours the active combination bE1 and bW2 under the control of the arabinose-inducible crg1 promoter. Samples were taken 3h, 5h and 12h after induction of bE/bW gene expression. Strain AB32, which harbors the incompatible bE2 and bW2 combination, was used as control.
Project description:The basidiomycete Ustilago maydis causes smut disease in maize. Colonization of the host plant is initiated by direct penetration of cuticle and cell wall of maize epidermis cells. The invading hyphae are surrounded by the plant plasma membrane and proliferate within the plant tissue. We identified a novel secreted protein, termed Pep1. Disruption mutants of pep1 are not affected in saprophytic growth and develop normal infection structures. However, Îpep1 mutants fail to penetrate the epidermal cell wall and elicit a strong plant defense response. Using Affymetrix maize arrays we identified about 110 plant genes which are differentially regulated in Îpep1 and wild type infections during the penetration stage. Experiment Overall Design: In three independent experiments plants were infected with the strain SG200Dpep1 which is derived from the solopathogenic U. maydis strain SG200. Samples from infected leaves were taken at 24 hours post infection. Samples were treated under the same conditions as described previosly (Doehlemann et al. (2008) Plant J, in press).
Project description:In the phytopathogenic Basidiomcete Ustilago maydis, sexual and pathogenic development are tightly connected and controlled via the b-mating type locus. The b-mating type locus encodes two homeodomain transcription factors, bE and bW, which form an active heterodimeric complex when they are derived from different b-alleles. Rbf1 encodes a zinc-finger transcription factor that is expressed upon formation of an active bE/bW heterodimer. To identify gene regulated by rbf1 independently from b, the b mating type locus was replaced by a copy of rbf1 under control of the arabinose-inducible crg1 promoter (strain CP27). Samples were taken 5h after induction of rbf1 gene expression. Strain JB2, carrying a deletion of the b-locus, was used as control.
Project description:The Brown Norway (BN) strain of rat is an inbred normotensive strain. BN rats of both sexes present some interesting pathophysiological phenotypes involving arteries and the kidneys. These include internal elastic lamina (IEL) ruptures in the abdominal aorta and iliac arteries, a deficit in aortic elastin content, a persistent ductus arteriosus, hydronephrosis and hematuria. Spontaneous rupture of the internal elastic lamina occurs in various arteries during growth and aging, in different rat strains, both normotensive and hypertensive. Most strains present such ruptures in their caudal and renal arteries, although to different extents (Osborne-Pellegrin 1985; Coutard and Osborne-Pellegrin 1991). However, the BN strain is the only rat strain to spontaneously develop numerous IEL ruptures in the abdominal aorta and iliac arteries (Osborne-Pellegrin et al., 1989; Behmoaras J et al., 2005). In this respect, it is exceptional. In this study samples of abdominal aortae of BN- and LOU-rats (here as control) were compared Keywords: strain effect
Project description:Nociceptive neurons develop a complex dendritic arbor to sense noxious stimuli, which enables animals to react to environmental insults and perform self-protective behaviours. The genetic programs controlling neuronal dendritic morphogenesis are poorly understood. In C. elegans, the PVD sensory neuron generates a complex dendritic arbor that envelops the body of the animal. This nociceptive neuron enables study of dendrite formation in vivo. MEC-3 is a transcription factor expressed in the PVD neuron (Chatzigeorgiou et al., 2010; Li et al., 2011; Way and Chalfie, 1989). mec- 3 mutant PVD neurons fail to elaborate lateral branches and show defective function (Smith et al., 2010; Tsalik et al., 2003) (Husson et al., 2012).