Abstract
The Vf locus, derived from the crabapple species Malus floribunda 821, confers resistance to five races of the fungal pathogen Venturia inaequalis, the causal agent of apple scab disease. In our previous research, the Vf locus was restricted to a BAC contig of approximately 290 kb covered by five overlapping BAC clones. Here, we report on cloning of the resistance gene(s) present in the Vf BAC contig using a highly reliable and straightforward approach. This approach relies on hybridization of labeled cDNAs to amplified inserts of subclones derived from BAC inserts, followed by recovery of full-size transcripts by rapid amplification of cDNA ends (RACE). A cluster of four resistance paralogs (Vfa1, Vfa2, Vfa3, and Vfa4) was identified in the Vf locus. Vfa1, Vfa2 and Vfa4 had no introns and are predicted to encode proteins characterized with extracellular leucine-rich repeats (LRRs) and transmembrane (TM) domains. However, Vfa3 contains an insertion of 780 bp at the end of the LRR motif, resulting in multiple truncated transcripts. Comparison of Vfa1, Vfa2, and Vfa4 paralogs revealed a high degree of overall homology in their deduced amino acid sequences, while divergences were mainly restricted within LRR domains, including variable LRR units, numerous amino acid substitutions, and several residue deletions/duplications. Differential expression profiles among the four paralogs were observed during leaf development. Vfa1, Vfa2, and Vfa3 were active in immature leaves, but slightly expressed in mature leaves, while Vfa4 was active in immature leaves and was highly expressed in mature leaves.
Full Text
The Full Text of this article is available as a PDF (203.3 KB).
Selected References
These references are in PubMed. This may not be the complete list of references from this article.
- Bergelson J., Kreitman M., Stahl E. A., Tian D. Evolutionary dynamics of plant R-genes. Science. 2001 Jun 22;292(5525):2281–2285. doi: 10.1126/science.1061337. [DOI] [PubMed] [Google Scholar]
- Buckler A. J., Chang D. D., Graw S. L., Brook J. D., Haber D. A., Sharp P. A., Housman D. E. Exon amplification: a strategy to isolate mammalian genes based on RNA splicing. Proc Natl Acad Sci U S A. 1991 May 1;88(9):4005–4009. doi: 10.1073/pnas.88.9.4005. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Cross S. H., Clark V. H., Bird A. P. Isolation of CpG islands from large genomic clones. Nucleic Acids Res. 1999 May 15;27(10):2099–2107. doi: 10.1093/nar/27.10.2099. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Dixon M. S., Hatzixanthis K., Jones D. A., Harrison K., Jones J. D. The tomato Cf-5 disease resistance gene and six homologs show pronounced allelic variation in leucine-rich repeat copy number. Plant Cell. 1998 Nov;10(11):1915–1925. doi: 10.1105/tpc.10.11.1915. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Dixon M. S., Jones D. A., Keddie J. S., Thomas C. M., Harrison K., Jones J. D. The tomato Cf-2 disease resistance locus comprises two functional genes encoding leucine-rich repeat proteins. Cell. 1996 Feb 9;84(3):451–459. doi: 10.1016/s0092-8674(00)81290-8. [DOI] [PubMed] [Google Scholar]
- Ellis J., Lawrence G., Ayliffe M., Anderson P., Collins N., Finnegan J., Frost D., Luck J., Pryor T. Advances in the molecular genetic analysis of the flax-flax rust interaction. Annu Rev Phytopathol. 1997;35:271–291. doi: 10.1146/annurev.phyto.35.1.271. [DOI] [PubMed] [Google Scholar]
- Elvin P., Slynn G., Black D., Graham A., Butler R., Riley J., Anand R., Markham A. F. Isolation of cDNA clones using yeast artificial chromosome probes. Nucleic Acids Res. 1990 Jul 11;18(13):3913–3917. doi: 10.1093/nar/18.13.3913. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Hammond-Kosack Kim E., Jones Jonathan D. G. PLANT DISEASE RESISTANCE GENES. Annu Rev Plant Physiol Plant Mol Biol. 1997 Jun;48(NaN):575–607. doi: 10.1146/annurev.arplant.48.1.575. [DOI] [PubMed] [Google Scholar]
- Hulbert S. H., Webb C. A., Smith S. M., Sun Q. Resistance gene complexes: evolution and utilization. Annu Rev Phytopathol. 2001;39:285–312. doi: 10.1146/annurev.phyto.39.1.285. [DOI] [PubMed] [Google Scholar]
- Jia Y., McAdams S. A., Bryan G. T., Hershey H. P., Valent B. Direct interaction of resistance gene and avirulence gene products confers rice blast resistance. EMBO J. 2000 Aug 1;19(15):4004–4014. doi: 10.1093/emboj/19.15.4004. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Kobe B., Deisenhofer J. A structural basis of the interactions between leucine-rich repeats and protein ligands. Nature. 1995 Mar 9;374(6518):183–186. doi: 10.1038/374183a0. [DOI] [PubMed] [Google Scholar]
- Leister D., Ballvora A., Salamini F., Gebhardt C. A PCR-based approach for isolating pathogen resistance genes from potato with potential for wide application in plants. Nat Genet. 1996 Dec;14(4):421–429. doi: 10.1038/ng1296-421. [DOI] [PubMed] [Google Scholar]
- Lovett M., Kere J., Hinton L. M. Direct selection: a method for the isolation of cDNAs encoded by large genomic regions. Proc Natl Acad Sci U S A. 1991 Nov 1;88(21):9628–9632. doi: 10.1073/pnas.88.21.9628. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Martin G. B., Brommonschenkel S. H., Chunwongse J., Frary A., Ganal M. W., Spivey R., Wu T., Earle E. D., Tanksley S. D. Map-based cloning of a protein kinase gene conferring disease resistance in tomato. Science. 1993 Nov 26;262(5138):1432–1436. doi: 10.1126/science.7902614. [DOI] [PubMed] [Google Scholar]
- Meyers B. C., Shen K. A., Rohani P., Gaut B. S., Michelmore R. W. Receptor-like genes in the major resistance locus of lettuce are subject to divergent selection. Plant Cell. 1998 Nov;10(11):1833–1846. doi: 10.1105/tpc.10.11.1833. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Michelmore R. W., Meyers B. C. Clusters of resistance genes in plants evolve by divergent selection and a birth-and-death process. Genome Res. 1998 Nov;8(11):1113–1130. doi: 10.1101/gr.8.11.1113. [DOI] [PubMed] [Google Scholar]
- Parimoo S., Patanjali S. R., Shukla H., Chaplin D. D., Weissman S. M. cDNA selection: efficient PCR approach for the selection of cDNAs encoded in large chromosomal DNA fragments. Proc Natl Acad Sci U S A. 1991 Nov 1;88(21):9623–9627. doi: 10.1073/pnas.88.21.9623. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Parniske M., Hammond-Kosack K. E., Golstein C., Thomas C. M., Jones D. A., Harrison K., Wulff B. B., Jones J. D. Novel disease resistance specificities result from sequence exchange between tandemly repeated genes at the Cf-4/9 locus of tomato. Cell. 1997 Dec 12;91(6):821–832. doi: 10.1016/s0092-8674(00)80470-5. [DOI] [PubMed] [Google Scholar]
- Richter T. E., Ronald P. C. The evolution of disease resistance genes. Plant Mol Biol. 2000 Jan;42(1):195–204. [PubMed] [Google Scholar]
- Salmeron J. M., Oldroyd G. E., Rommens C. M., Scofield S. R., Kim H. S., Lavelle D. T., Dahlbeck D., Staskawicz B. J. Tomato Prf is a member of the leucine-rich repeat class of plant disease resistance genes and lies embedded within the Pto kinase gene cluster. Cell. 1996 Jul 12;86(1):123–133. doi: 10.1016/s0092-8674(00)80083-5. [DOI] [PubMed] [Google Scholar]
- Scofield SR, Tobias CM, Rathjen JP, Chang JH, Lavelle DT, Michelmore RW, Staskawicz BJ. Molecular Basis of Gene-for-Gene Specificity in Bacterial Speck Disease of Tomato. Science. 1996 Dec 20;274(5295):2063–2065. doi: 10.1126/science.274.5295.2063. [DOI] [PubMed] [Google Scholar]
- Song W. Y., Wang G. L., Chen L. L., Kim H. S., Pi L. Y., Holsten T., Gardner J., Wang B., Zhai W. X., Zhu L. H. A receptor kinase-like protein encoded by the rice disease resistance gene, Xa21. Science. 1995 Dec 15;270(5243):1804–1806. doi: 10.1126/science.270.5243.1804. [DOI] [PubMed] [Google Scholar]
- Sood R., Bonner T. I., Makalowska I., Stephan D. A., Robbins C. M., Connors T. D., Morgenbesser S. D., Su K., Faruque M. U., Pinkett H. Cloning and characterization of 13 novel transcripts and the human RGS8 gene from the 1q25 region encompassing the hereditary prostate cancer (HPC1) locus. Genomics. 2001 Apr 15;73(2):211–222. doi: 10.1006/geno.2001.6500. [DOI] [PubMed] [Google Scholar]
- Xu Mingliang, Korban Schuyler S. AFLP-derived SCARs facilitate construction of a 1.1 Mb sequence-ready map of a region that spans the Vf locus in the apple genome. Plant Mol Biol. 2002 Nov;50(4-5):803–818. doi: 10.1023/a:1019912419709. [DOI] [PubMed] [Google Scholar]
- Young N. D. The genetic architecture of resistance. Curr Opin Plant Biol. 2000 Aug;3(4):285–290. doi: 10.1016/s1369-5266(00)00081-9. [DOI] [PubMed] [Google Scholar]
- Zhang M. Q. Identification of protein coding regions in the human genome by quadratic discriminant analysis. Proc Natl Acad Sci U S A. 1997 Jan 21;94(2):565–568. doi: 10.1073/pnas.94.2.565. [DOI] [PMC free article] [PubMed] [Google Scholar]