1. Introduction
1.1 Schizosaccharomyces pombe as a Model System
S. pombe functions as a suitable model system since it is easy and inexpensive to rear, has a convenient size, a short life cycle, and is genetically manipulable. As a unicellular eukaryote, the fission yeast S. pombe can exist either in a haploid or diploid state and possesses two different mating types (h+ and h-). The wild type, however, is h90, which means it can switch mating type.
Figure 1.01: Left, picture of S. pombe cells
At top are two dividing cells in late mitotic phase, showing the fission yeast typical septum at the point of cytoplasmic division. The lower cell is in early M phase, having its chromosomes already segregated.
Figure 1.02: Right, fission yeast cell cycle
Diagrammatic representation of the S. pombe cell cycles with the interchange between the two occurring in G1 phase (Figure obtained and used with permission from Trevor Pemberton, University of Sussex).
[...]
S. pombe can undergo two different life cycles, either the vegetative (mitotic) cycle or the sporulation (meiotic) cycle, depending on the environment it is living in. These two cycles are shown in figure 2 with the change between the two occurring in cells at the G1 stage of the mitotic cycle. Under laboratory conditions, given all nutrients required, S. pombe prefers the haploid state. This makes it a favorable organism for genetic research since it ensures that introduced mutations are not masked by another wild type allele. [...]
Table of Contents
1. Introduction
1.1 Schizosaccharomyces pombe as a Model System
1.2 Why Investigating Signaling Cascades?
1.3 The SAPK Pathway
1.4 The cAMP Pathway
1.5 The Pheromone Pathway
1.6 Downstream the PKA and SAPK pathways
1.7 Transporters in Fission and Budding Yeast
1.8 Glycolysis, Gluconeogenesis and Glycerol Metabolism
1.9 Microarrays
1.10 Thesis Goal
2. Materials
2.1 Sources of Used Chemicals, Enzymes, and Kits
2.2 S. pombe Strains
2.3 Solutions and Yeast Media
2.4 Equipment
3. Methods
3.1 Experimental Design
3.2 Growth of S.pombe Strains
3.3 Harvesting Cells
3.4 RNA Extraction
3.5 Sample Preparation
3.6 Microarray
4. Results
4.1 Genes Up-Regulated upon Glucose Starvation
4.2 Highly Induced Genes of the Carbohydrate Metabolism
4.3 Genes Involved in Mating/Meiosis
4.4 Genes Involved in Global Transcriptional Regulation
4.5 Hexose Transporters
4.6 cAMP, SAPK, Pheromone Pathway Genes
4.7 Glucose Starvation vs. Oxidative Stress
4.8 Glucose starvation vs. Nitrogen starvation
4.9 Down-Regulated Processes
4.10 How does Gene Expression Change in a spc1 Deletion Mutant?
4.11 PombePerl
5. Discussion
5.1 Changes in Metabolic Pathways
5.2 Stress Activated Signaling Pathways
5.3 Changes of a Cell’s Global Processes
5.4 Effects of a spc1 Deletion on Signaling Pathways
5.5 Summary and Outlook
Research Objectives and Themes
The primary objective of this thesis is to comprehensively characterize the global transcriptional response of the fission yeast Schizosaccharomyces pombe to glucose starvation stress. The research addresses how the genome-wide expression profile shifts under these conditions, how metabolic and signaling pathways are reorganized, and how the loss of the stress-activated protein kinase Spc1p alters these transcriptional regulatory patterns.
- Genome-wide transcriptional response to glucose deprivation and re-feeding.
- Reorganization of metabolic pathways, including glycolysis and gluconeogenesis.
- Interplay between cAMP, SAPK, and pheromone signaling pathways.
- Impact of the spc1 mutation on stress-induced gene expression.
- Development of bioinformatics tools (PombePerl) for automated gene annotation.
Excerpt from the Book
1.1 Schizosaccharomyces pombe as a Model System
S. pombe functions as a suitable model system since it is easy and inexpensive to rear, has a convenient size, a short life cycle, and is genetically manipulable. As a unicellular eukaryote, the fission yeast S. pombe can exist either in a haploid or diploid state and possesses two different mating types (h+ and h-). The wild type, however, is h90, which means it can switch mating type.
S. pombe can undergo two different life cycles, either the vegetative (mitotic) cycle or the sporulation (meiotic) cycle, depending on the environment it is living in. These two cycles are shown in figure 2 with the change between the two occurring in cells at the G1 stage of the mitotic cycle. Under laboratory conditions, given all nutrients required, S. pombe prefers the haploid state. This makes it a favorable organism for genetic research since it ensures that introduced mutations are not masked by another wild type allele.
There are many similarities between relevant physiological processes in yeast and mammalian cells as supported by the cloning of the human homologs of yeast genes. Some physiological processes (e.g., mitosis, cell division) of S. pombe are more similar to those of human cells than those of the budding yeast Saccharomyces cerevisiae. Like human cells, S. pombe has a distinct G2 phase so a major checkpoint control is the decision to go from G2 to M (Russell and Nurse, 1986).
Summary of Chapters
1. Introduction: Provides an overview of S. pombe as a biological model and describes the signaling pathways (cAMP, SAPK, pheromone) and metabolic processes that are fundamental to yeast cell physiology.
2. Materials: Lists the specific chemicals, yeast strains, reagents, and laboratory equipment used throughout the experiments.
3. Methods: Details the experimental design, including growth conditions, RNA extraction, microarray processing, and the computational approaches used for data normalization and clustering.
4. Results: Presents the findings of the genome-wide transcriptional analysis, categorizing induced and repressed genes and examining the effects of the spc1 deletion on pathway regulation.
5. Discussion: Synthesizes the results to explain how S. pombe reprograms its metabolic and transcriptional state to survive glucose starvation and explores the broader implications for signaling network analysis.
Keywords
Schizosaccharomyces pombe, Glucose starvation, Microarrays, Transcriptional response, SAPK pathway, cAMP signaling, Pheromone pathway, Gene expression, Gluconeogenesis, Metabolic pathways, Spc1p, Signal transduction, Bioinformatics, Stress response, Mating and meiosis
Frequently Asked Questions
What is the core focus of this research?
The work focuses on investigating the global changes in gene expression in S. pombe when the cells are subjected to glucose starvation and subsequent re-feeding, using microarray technology to map these changes.
Which specific signaling pathways are examined?
The study primarily investigates the SAPK (Stress Activated Protein Kinase) pathway, the cAMP signaling pathway, and the pheromone-activated MAPK cascade, focusing on their roles in responding to nutritional stress.
What is the primary goal of the study?
The goal is to determine how the complete genome of S. pombe responds transcriptionally to glucose starvation, how specific pathways are affected, and how the absence of the MAP kinase Spc1p influences these responses.
Which scientific methodology is applied?
The methodology includes time-course microarray experiments, RNA extraction, cDNA synthesis, fluorescent labeling, and computational cluster analysis to identify co-regulated gene groups.
What topics are covered in the main section of the document?
The main sections cover the identification of up-regulated and down-regulated genes, analysis of carbohydrate metabolism, mating/meiosis regulation, and a comparative study between glucose and oxidative stress.
What characterize the key terms associated with this thesis?
The thesis is characterized by the integration of molecular genetics, transcriptomics, and computational biology to understand cellular adaptation strategies in eukaryotic model organisms.
How does the deletion of the spc1 gene affect the cells?
The deletion of the spc1 MAP kinase gene primarily disrupts the stress-activated signaling pathway, leading to altered transcriptional behavior of genes in the PKA and pheromone pathways compared to wild-type strains.
What role does the PombePerl tool play in this project?
PombePerl is a custom software tool developed by the author to automate the matching of gene names from microarray datasets with functional descriptions from online databases like GeneDB.
- Quote paper
- Michael Sassen (Author), 2005, Global Transcriptional Responses of Fission Yeast to Glucose Starvation Stress, Munich, GRIN Verlag, https://www.grin.com/document/41887
