Os04g0494100

GeneOs04g0494100 ,namelyOsChia4a,is a class IV chitinase that has similarity (about 70z identity) to both maize ChiA and Arabidopsis ChIV of class IV chitinases.

Annotated Information

Function

File:Mutation analysis.png

Fig. 1 Mutation analysis of candidate cis regulatory elements in the promoter. (A) Nucleotide sequence of the region from −515 to −265 upstream of the ATG translation initiation site of OsChia4a. The three E-boxes are denoted by E1–E3. (B) Mutated E-boxes (indicated by X), in which the CANNTG sequence was altered to AANNTG, were used for the dual LUC assays as performed in Fig. 3. In each construct (m1, m2 and m3), mutation was introduced into each E-box (E1, E2 and E3), respectively. Black and open bars indicate the relative LUC activities (FLUC/RLUC) of the deletion derivatives after 12 h of incubation of the rice cells with and without 100 μM JA, respectively. The data are expressed as mean values plus/minus the standard deviation of three replicates. A statistical analysis was performed using the t-test, and the significant differences (p < 0.01) between the relative LUC activities in the treated and in the untreated cells were denoted by asterisks.

Fig. 2 Family 19 Chitinases of Nipponbare and Other Cultivars .

File:Antifungal activity.png

Fig. 3Antifungal activity of purified OsChia4a-His protein. (A) Purified OsChia4a-His protein (lane P) was analysed by 12% SDS-PAGE along with molecular standards (lane M); it is shown by Coomassie Brilliant Blue staining. Arrowhead shows the purified OsChia4a-His protein (31.4 kDa). (B) A typical image of hyphae elongation of washed conidia of M. oryzae, which were mixed with purified OsChia4a-His protein or bovine serum albumin (BSA). After incubation for one day at 28 °C, the growth of M. oryzae was examined under the microscope. (C) Hyphal length of incubated conidia as shown in (B) was measured. The results are the average of more than 100 conidia examined in each condition; bars indicate the standard error from the mean value. .

• OsChia4a, which was identified to be one of the highest JA-inductive genes. The recombinant protein of His-tagged OsChia4a exhibited an inhibitory effect against the spore germination and hyphal growth of Magnaporthe oryzae .The promoter analysis of OsChia4a revealed that the region from −515 bp to −265 bp upstream of the ATG translation initiation site was required for the responsiveness to JA. A subsequent mutation analysis indicated that an E-box (CANNTG) in this region act as a JA-responsive cis element. These results imply that a basic helix-loop-helix transcription factor is likely to be involved in the regulation of the OsChia4a expression in a JA-dependent manner(Fig. 1).
  
  
• The high sequence conservation (95z identity) between the catalytic domains of OsChia4a (class IV) and OsChia2b (class II) provides an exam- ple of the direct and recent diversion of their genes via either deletion or insertion mechanisms within rice.The class IV catalytic domains have three deletions that correspond to the loops between a- helices C and D, F and G, and G and H.These loops do not appear to signicantly contribute to stability and activity towards chitin polymers (Fig. 2).
  
  
• OsChia4a is one of the highest induced chitinase genes (in terms of fold induction compared to the reference, untreated sample) both in the leaves and in the suspension cell. Unlike class IV, class I chitinases, OsChia1a and OsChia1c, which have been reported to be induced by the chitin elicitor in suspension cells, were only induced in the leaves and not in the suspension cells after the JA treatment .
  
  
• The antifungal activity of OsChia4a was demonstrated using the recombinant His-tagged OsChia4a protein that was produced in N. benthamiana using the Cowpea mosaic virus vector system .The protein was purified by using MagneHis Protein Purification System (Promega) ( Fig. 1A) and subjected to the hyphal extension-inhibition assay against M. oryzae as described in section “Materials and methods”(Fig. 3).
  
  

Expression

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Evolution

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Structured Information

Gene Name	Os04g0494100
Description	Similar to Chitinase
Version	NM_001059721.1 GI:115459171 GeneID:4336265
Length	1169 bp
Definition	Oryza sativa Japonica Group Os04g0494100, complete gene.
Source	Oryza sativa Japonica Group ORGANISM Oryza sativa Japonica Group Eukaryota; Viridiplantae; Streptophyta; Embryophyta; Tracheophyta; Spermatophyta; Magnoliophyta; Liliopsida; Poales; Poaceae; BEP clade; Ehrhartoideae; Oryzeae; Oryza.
Chromosome	Chromosome 4
Location	Chromosome 4:25107288..25108456
Sequence Coding Region	25107468..25107874,25107963..25108422
Expression	GEO Profiles:Os04g0494100
Genome Context	<gbrowseImage1> name=NC_008397:25107288..25108456 source=RiceChromosome04 preset=GeneLocation </gbrowseImage1>
Gene Structure	<gbrowseImage2> name=NC_008397:25107288..25108456 source=RiceChromosome04 preset=GeneLocation </gbrowseImage2>
Coding Sequence	<cdnaseq>atggcgaactcaccgacgccgacaatgctggcgttcctggctcttgggctagcgctcctcctctccgccaccggccaggcgagcgcgcagaactgcggctgccagtcgaacatgtgctgcagcaaatgggggtactgcggcacgggcaaggactactgcggagatgggtgccgctctggcccgtgctacggcggcggcggcggtggaggaggaggaggcggaggtggtggaggcggaggcggaggcagcggcgtgtctgtagagagcgtggtcaccgaggcgttcttcaatgggatcaagaaccaggccccgaacggttgcgccggcaagaacttttacacacgacagtcgtttcttaacgctgcccactcctactcgggcttcgccagggaccgcaccaacgatgactccaagcgtgagatcgctgccttctttgcccacgtcactcatgagaccggacatatgtgctacatcaacgagataaacggggcgagcatggactactgcgacaagaacaacaagcagtggccgtgccagccggggaagaagtactacgggcgcgggccgctgcagatctcgtggaactacaactacgggcctgcggggcagaacatcgggttcgacgggctgagggacccggacagggtggcgcaggacccgacgatctccttcaagacggcgctctggttctggatgaacaacgtgcaccaggtgatgttgcaggggttcggcgccaccatccgggccatcaacggtgcgctcgagtgcaacggcaagaaccccggcgccgtcaacgcaagggtaaactactacaaagactactgccgccaattcggcgttgacccgggtggcaacctttactgttga</cdnaseq>
Protein Sequence	<aaseq>MANSPTPTMLAFLALGLALLLSATGQASAQNCGCQSNMCCSKWG YCGTGKDYCGDGCRSGPCYGGGGGGGGGGGGGGGGGGGSGVSVESVVTEAFFNGIKNQ APNGCAGKNFYTRQSFLNAAHSYSGFARDRTNDDSKREIAAFFAHVTHETGHMCYINE INGASMDYCDKNNKQWPCQPGKKYYGRGPLQISWNYNYGPAGQNIGFDGLRDPDRVAQ DPTISFKTALWFWMNNVHQVMLQGFGATIRAINGALECNGKNPGAVNARVNYYKDYCR QFGVDPGGNLYC</aaseq>
Gene Sequence	<dnaseqindica>583..989#35..494#atcaaccatcacctctttcacaccatatcctataatggcgaactcaccgacgccgacaatgctggcgttcctggctcttgggctagcgctcctcctctccgccaccggccaggcgagcgcgcagaactgcggctgccagtcgaacatgtgctgcagcaaatgggggtactgcggcacgggcaaggactactgcggagatgggtgccgctctggcccgtgctacggcggcggcggcggtggaggaggaggaggcggaggtggtggaggcggaggcggaggcagcggcgtgtctgtagagagcgtggtcaccgaggcgttcttcaatgggatcaagaaccaggccccgaacggttgcgccggcaagaacttttacacacgacagtcgtttcttaacgctgcccactcctactcgggcttcgccagggaccgcaccaacgatgactccaagcgtgagatcgctgccttctttgcccacgtcactcatgagaccggacgtaaggaaaatatttaattacatactcctggtatttgtacatatatgcatgtaactaatgaggatgataaatgataatgcactcgcagatatgtgctacatcaacgagataaacggggcgagcatggactactgcgacaagaacaacaagcagtggccgtgccagccggggaagaagtactacgggcgcgggccgctgcagatctcgtggaactacaactacgggcctgcggggcagaacatcgggttcgacgggctgagggacccggacagggtggcgcaggacccgacgatctccttcaagacggcgctctggttctggatgaacaacgtgcaccaggtgatgttgcaggggttcggcgccaccatccgggccatcaacggtgcgctcgagtgcaacggcaagaaccccggcgccgtcaacgcaagggtaaactactacaaagactactgccgccaattcggcgttgacccgggtggcaacctttactgttgagttacatgcacacatgctgcgcggctcatcgatcaggactgaataataaacaaagtaataacgaataaaacgtttggtgtgtacgtacattgcatgcacgtacggtacatgttgcaagtgacgtcaagagtttggtacggataaatatgaaaaataaaatgttgggatcaggattaaatg</dnaseqindica>
External Link(s)	NCBI Gene:Os04g0494100, RefSeq:Os04g0494100