Cultivar specificity gene from the rice pathogen Magnaporthe...

Details Cultivar specificity gene from the rice pathogen Magnaporthe... Cultivar specificity gene from the rice pathogen Magnaporthe...

1999-02-25

2004-09-21

Nelson, Amy J. (Department: 1638)

Chemistry: molecular biology and microbiology

Micro-organism, per se ; compositions thereof; proces of...

Bacteria or actinomycetales; media therefor

C536S023600, C435S320100

Reexamination Certificate

active

06794176

ABSTRACT:

FIELD OF THE INVENTION
This invention relates to the field of disease resistance in plants. In particular, the invention provides a novel avirulence gene from the rice blast pathogen,
Magnaporthe grisea
, and methods of using the gene and its encoded products for improving resistance of rice to this pathogen.
BACKGROUND OF THE INVENTION
Rice is a major staple food for about two-thirds of the world's population. More than ninety percent of the world's rice is grown and consumed in developing countries. Rice blast disease, caused by the fungus
Magnaporthe grisea
, threatens rice crops worldwide. The disease can cause yield losses of ten to thirty percent in infested fields. Rice blast has been an ongoing problem in rice growing areas of the southern United States. It has now become a significant problem in rice growing areas of California, as well.
The “gene-for-gene” hypothesis has been advanced to explain the very specific disease resistance/susceptibility relationship that often exists between races of a plant pathogen and cultivars of its host species. The gene-for-gene hypothesis has been found applicable to many host-pathogen interactions, including that of the rice blast fungus,
Magnaporthe grisea
, and its host,
Oryza sativa
. To be able to understand and manipulate this host-pathogen relationship is of great practical interest as
M. grisea
is rapidly able to overcome new disease resistance in rice soon after their deployment. Moreover,
M. grisea
exists as a complex genus with many subspecific groups that are infertile, but differ in their host range. How these different subspecific groups interrelate evolutionarily is of great concern to plant breeders since some of these alternate hosts are frequently found growing in close proximity to, or in rotation with rice, and
M. grisea
isolates infecting these alternate hosts can sometimes also infect rice.
Gene-for-gene resistance (also known as hypersensitive resistance (HR) or race-specific resistance) depends for its activation on specific recognition of the invading pathogen by the plant. Many individual plant genes have been identified that control gene-for-gene resistance. These genes are referred to as resistance (R) genes. The function of a particular R gene depends on the genotype of the pathogen. A pathogen gene is referred to as an Avr gene if its expression causes the pathogen to produce a signal that triggers a strong defense response in a plant having a corresponding R gene. This response is not observed in the absence of either the Avr gene in the pathogen or the corresponding R gene in the plant. It should be noted that a single plant may have many R genes, and a single pathogen may have many Avr genes. However, strong resistance occurs only when an Avr gene and its specific R gene are matched in a host-pathogen interaction. In this instance, resistance generally occurs as activation of a HR response, in which the cells in the immediate vicinity of the infection undergo programmed necrosis in order to prevent the further advance of the pathogen into living plant tissue. Other features of the resistance response may also include synthesis of antimicrobial metabolites or pathogen-inhibiting enzymes, reinforcement of plant cell walls in the infected area, and induction of signal transduction pathways leading to systemic acquired resistance (SAR) in the plant.
The molecular basis of host-cultivar specificity and pathogenic variability in
M. grisea
is only beginning to be elucidated with the identification, mapping and, in some instances, cloning of specific Avr genes from pathogenic isolates of
M. grisea
. For instance, AVR2-YAMO (cultivar specificity) and PWL2 (host specificity) (Valent & Chumley, pp. 3.113-3.134 in
Rice Blast Disease
(R. Zeigler, S. A. Leong, P. Teng, Eds.), Wallingford: CAB International, 1994) both function as classic avirulence genes by preventing infection of a specific cultivar or host. AVR2-YAMO encodes a 223-amino acid protein with homology to proteases, while PWL2 encodes a 145-amino acid polypeptide which is glycine-rich. Based on the predicted amino acid sequences of the proteins, both may be secreted.
Homologs of both AVR2-YAMO and PWL2 appear to be widely distributed in rice and in other grass-infecting isolates of
M. grisea
, thereby confirming that
M. grisea
isolates which do not infect rice still may carry host or cultivar specificity genes for rice. In some cases, homologs of AVR2-YAMO and PWL2 have been shown to be functional and to exhibit the same host or cultivar specificity as AVR2-YAMO or PWL2.
As another example of a potentially useful Avr gene, the cultivar specificity gene AVR1-CO39, which determines avirulence on rice cultivar CO39, has been identified (Valent et al., Genetics 127: 87-101, 1991) and mapped to a position on
M. grisea
chromosome 1 (Smith & Leong, Theor. Appl. Genet. 88: 901-908, 1994). A segment of chromosome 1 that appears to contain the AVR1-CO39 gene has been isolated and cloned into a cosmid vector (Leong et al., pp. 846-852 in
Rice Genetics III, Proceedings of the Third Annual Rice Genetics Symposium
, G. S. Khush, Ed., Island Harbor Press, Manila, 1996); however, the gene itself heretofore has not been identified and characterized.
The availability of cloned cultivar and host specificity genes from
M. grisea
and, ultimately, the corresponding R genes from rice provides useful tools for manipulating and augmenting resistance to this pathogen in the field. Accordingly, it is an object of the present invention to provide a new cloned
M. grisea
cultivar specificity gene, AVR1-CO39, and its functional homologs for such use.
SUMMARY OF THE INVENTION
According to one aspect of the invention, there is provided an isolated nucleic acid, AVR1-CO39, from
Magnaporthe grisea
that confers rice cultivar CO39-specific avirulence to fungal plant pathogens that contain the nucleic acid. The nucleic acid preferably comprises part or all of Sequence I.D. No. 1, or hybridizes with part or all of Sequence I.D. No. 1 or its complement.
According to another aspect of the invention, there is provided a polypeptide encoded by part or all of the isolated nucleic acid of claim 1. Preferably, the polypeptide is selected from the group of polypeptides encoded by ORFS 1, 2, 3, 4, 5, 6 and 7, corresponding to Sequence ID No's. 2, 3, 4, 5, 6, 7 and 8, respectively, and most preferably is encoded by ORF 3.
According to another aspect of the invention, a transgenic epiphytic bacterium is provided, which expresses a portion of an AVR1-CO39 gene effective to confer rice cultivar CO39-specific avirulence to microorganisms expressing the gene. Preferably, the transgenic epiphytic bacterium expresses ORF3 of Sequence ID No. 1, or a functional equivalent.
According to another aspect of the invention, a method of enhancing the scope of resistance of rice cultivar CO39 plants to pathogenic microorganisms is provided. The method comprises treating the plants with an epiphytic bacterium that expresses a portion of an AVR1-CO39 gene effective to trigger expression of a CO39-specific R gene in the plants.
According to another aspect of the invention, a second method of enhancing the scope of resistance of rice cultivar CO39 plants to pathogenic microorganisms is provided. This method comprises treating the plants with a protein extract comprising polypeptides produced by expression of AVR1-CO39, in an amount effective to trigger expression of a CO39-specific R gene in the plants.
These and other features and advantages of the present invention will be described in greater detail in the description and examples set forth below.
DETAILED DESCRIPTION OF THE INVENTION
I. Definitions
Various terms relating to the biological molecules of the present invention are used hereinabove and also throughout the specifications and claims. The terms “substantially the same,” “percent similarity” and “percent identity” are defined in detail below.
With reference to nucleic acids of the invention, the term “isolated nucleic acid” is sometimes used. This term, when applied

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