EDS1, an essential component of R gene-mediated disease resistance in Arabidopsis has homology to eukaryotic lipases - PubMed (original) (raw)
EDS1, an essential component of R gene-mediated disease resistance in Arabidopsis has homology to eukaryotic lipases
A Falk et al. Proc Natl Acad Sci U S A. 1999.
Abstract
A major class of plant disease resistance (R) genes encodes leucine-rich-repeat proteins that possess a nucleotide binding site and amino-terminal similarity to the cytoplasmic domains of the Drosophila Toll and human IL-1 receptors. In Arabidopsis thaliana, EDS1 is indispensable for the function of these R genes. The EDS1 gene was cloned by targeted transposon tagging and found to encode a protein that has similarity in its amino-terminal portion to the catalytic site of eukaryotic lipases. Thus, hydrolase activity, possibly on a lipid-based substrate, is anticipated to be central to EDS1 function. The predicted EDS1 carboxyl terminus has no significant sequence homologies, although analysis of eight defective eds1 alleles reveals it to be essential for EDS1 function. Two plant defense pathways have been defined previously that depend on salicylic acid, a phenolic compound, or jasmonic acid, a lipid-derived molecule. We examined the expression of EDS1 mRNA and marker mRNAs (PR1 and PDF1.2, respectively) for these two pathways in wild-type and eds1 mutant plants after different challenges. The results suggest that EDS1 functions upstream of salicylic acid-dependent PR1 mRNA accumulation and is not required for jasmonic acid-induced PDF1.2 mRNA expression.
Figures
Figure 1
High-resolution mapping and transposon tagging of EDS1. (A) Genetic map. Recombinant analysis placed EDS1 0.2 cM centromeric to I18, an restriction fragment length polymorphism (RFLP) marker derived from a I/dSpm transposon insertion in Ler. (B) P1 contig. I18 was used to identify two P1 phage clones, 73I2 and 69D23. An RFLP was detected between Ler and Col-0 DNA with the 73I2 centromeric end-probe, allowing orientation of P1 clones relative to EDS1. I/dSpm insertions into EDS1 were located in Ler DNA corresponding to a 5.7-kb internal _Bgl_II fragment of P1 clones 105H5 and 5N12. (C) A blot of _Bgl_II-digested genomic DNA was probed with a 32P-labeled inverse-PCR product derived from an I/dSpm insertion shared by eds1 lines T1–T5. The blot shows the wild-type Ler 5.7-kb band and deletions of ≈1 or ≈0.5 kb, respectively, in the FN-derived Ler mutants eds1–2 and eds1–3. Lines T1 and T2 possess an additional 7.9-kb band caused by insertion of a 2.2-kb I/dSpm element. In contrast to a Noco2-susceptible F1 plant (S1) derived from a cross between T1 and eds1–3, three independent Noco2-resistant (revertant) F1 plants (R1, R2, and R3) have lost the I/dSpm insertion.
Figure 2
Nucleotide sequence of the EDS1 gene and derived amino acid sequence. The isolated EDS1 cDNA encodes a predicted ORF of 1,869 nt with untranslated 5′ and 3′ leader sequences of 37 and 180 nt, respectively. The L-family lipase consensus sequence around the predicted catalytic serine (S123) is underlined. The three predicted lipase catalytic residues, a serine (S123), an aspartate (D187), and a histidine (H317) are indicated by a double underline.
Figure 3
Homology of EDS1 to eukaryotic lipases. (A) Alignment of EDS1 amino acids 95–205 containing the serine (S) and aspartic acid (D) residues that form part of a putative lipase catalytic triad, to lipases from R. miehei (Rhimi_), R. niveus,_ (Rhiniv), P. camembertii (Penca) and Humicola lanuginosa (Humlan), an esterase from A. niger (Aspnig), and hypothetical lipases from A. thaliana (Isologs 1–4), Ipomoea nil (Ipomoea), and C. elegans (Cael1–2). Crystal structures for the Rhimi, Penca and Humlan lipases have been determined (–28), and their active site S and D residues are indicated by an arrow above the sequence. Identical amino acids are shown in black boxes while conserved amino acid changes are shaded in gray. A pairwise alignment of the EDS1 and R. miehei lipase amino acid sequences shown here reveals overall identity of 29% and a similarity score of 46%. Numbers to the right refer to amino acid positions of the full-length proteins (see Methods). (B) Alignment of EDS1 amino acids 307–321 around the putative catalytic histidine (H) to fungal lipase/esterases (see A for details). The arrow marks the catalytic histidine determined from crystal structures for Rhimi, Penca, and Humlan (–28). For the other putative plant lipases it was not possible to generate a consensus alignment around the known catalytic histidine of the above fungal lipases. (C) Conservation of secondary structure elements around the lipase catalytic serine. All sequences shown in A conform to this pattern. The arrow indicates the conserved conformational presentation of the catalytic serine in lipases. Amino acid positions are indicated on the right and left sides.
Figure 4
RNA gel blot of Arabidopsis Ler and eds1–2 plants after various treatments. Total RNA was extracted from wild-type Ler (A) and Ler eds1–2 (B) at indicated times: healthy leaves (untreated), leaves infiltrated with suspensions of avirulent P. syringae strain DC3000 expressing avrRps4 (avr+), or with virulent strain DC3000 containing no functional avr gene (avr−), wounded leaves (wound), and leaves sprayed with SA or JA. Blots were probed simultaneously with 32P-labeled EDS1, PR1, and PDF1.2 sequences and stripped before reprobing with an 18S ribosomal DNA fragment. A second, independent experiment gave similar results.
References
- Baker B, Zambryski P, Staskawicz B, Dinesh-Kumar S P. Science. 1997;276:726–733. - PubMed
- Delaney T P, Uknes S, Vernooij B, Friedrich L, Weymann K, Negrotto D, Gaffney T, Gut-Rella M, Kessmann H, Ward E, Ryals J. Science. 1994;266:1247–1250. - PubMed
- Zhou J, Loh Y-T, Bressan R A, Martin G B. Cell. 1995;83:925–935. - PubMed
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