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Cloning and Functional Analysis of Hexose Kinase and Inhibitor of Apoptosis Protein Genes from Magnaporthe Grisea

Author: ZhangLiSha
Tutor: ZhangZhengGuang
School: Nanjing Agricultural College
Course: Genetics
Keywords: M. grisea Glucokinase Hexokinase Glucose sensor Inhibitor of apoptosis protein BIR domain
CLC: S435.111.41
Type: Master's thesis
Year: 2008
Downloads: 91
Quote: 2
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Abstract


Rice blast fungus (Magnaporthe grisea) heterothallic ascomycetes infested rice due to rice blast caused a huge loss to the world rice production every year. Understanding of the molecular genetic basis of rice blast fungus growth and development of the pathogenic process can provide theoretical guidance for rice blast damage control. Glucose signal transduction pathway, hexokinase (hexokinase, HXK) as the glucose receptors involved in feelings of glucose. The cloning of the gene encoding M.grisea glucokinase and hexokinase to and were named MgGLK1 and MgHXK1. The Southem blot results showed that the in M.grisea genome MgGLK1 and MgHXK1 are single copy. Protein sequence analysis showed close genetic relationship with the other fungal homologues, and distant genetic relationship with the plants and animals. Yeast functional complementation the analysis showed MgGLK1 and MgHXK1 encodes a protein having functions of glucokinase and hexokinase. By knockout, and found that the mutant conidia production M.grisea △ glk1 compared with Guy11 and △ hxk1,, mutants have decreased. In addition, △ glk1 the mutant culture conditions using glucose as the carbon source will reduce the use of ammonium salt; improve glucose concentration or exogenous glucose -6 - phosphate (G-6-P) can restore the the △ glk1 mutant on the use of ammonium salts. MgGLK1 gene and its loss of catalytic activity allele able to complement the to △ glk1 mutant ability to use ammonium salt. The semi-quantitative RT-PCR analysis showed that the, MgGLK1 involved in transcriptional regulation of the ATPase gene. The above results indicate that the the MgGLK1 as a glucose sensor does not depend on its glucokinase activity to regulate the use of the ammonium salt for M.grisea may also exist; and in M.grisea in other receptors by the feel high concentrations of G-6-P content change Adjust the use of the ammonium salt. The inhibitor of apoptosis protein (inhibitor of apoptosis protein, IAP) plays an important negative role in the apoptotic process. The cloned M.grisea apoptosis inhibitory protein gene, named MgIAP. MgIAP cDNA length of 2646 bp, encoding a protein of 881 amino acids. The protein of MgIAP contains two conserved BIR domain. The phylogenetic tree analysis showed that Survivin, Bruce and Drosophila MgIAP mammals dBruce closest genetic relatives, baculovirus and mammalian apoptosis inhibitory protein kinship far. The BIR domain phylogenetic analysis found, MgIAP BIR domains belong to the type Ⅱ BIR. Southern blot analysis, MgIAP M.grisea genome as a single copy. Generation of ROS MgIAP heterologous expression in yeast, can inhibit the generation of ROS in the process of cell death induced by oxidative stress and cell death, yeast cells can also postpone the timing of aging and the aging process. The domain deletion mutant analysis shows that the carboxyl terminal of the MgIAP in the postponed yeast cell timing aging play an important role.

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CLC: > Agricultural Sciences > Plant Protection > Pest and Disease Control > Crop pests and diseases and their prevention > Cereal crop pests and diseases > Rice pests and diseases > Disease > Transgression ( pass ) an infectious agent harmful > Rice blast
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