Cell-Autonomous Expression of Barley Mla1 Confers Race-Specific Resistance to the Powdery Mildew Fungus via a Rar1-Independent Signaling Pathway

Abstract

The barley Mla locus encodes 28 characterized resistance specificities to the biotrophic fungal pathogen barley powdery mildew. We describe a single-cell transient expression assay using entire cosmid DNAs to pinpoint Mla1 within the complex 240-kb Mla locus. The MLA1 cDNA encodes a 108-kD protein containing an N-terminal coiled-coil structure, a central nucleotide binding domain, and a C-terminal leucine-rich repeat region; it also contains a second short open reading frame at the 5' end that has a possible regulatory function. Although most Mla-encoded resistance specificities require Rar1 for their function, we used the single-cell expression system to demonstrate that Mla1 triggers full resistance in the presence of the severely defective rar1-2 mutant allele. Wheat contains an ortholog of barley Mla, designated TaMla, that is tightly linked to (0.7 centimorgan) but distinct from a tested resistance specificity at the complex Pm3 locus to wheat powdery mildew. Thus, the most polymorphic powdery mildew resistance loci in barley and wheat may have evolved in parallel at two closely linked homeoloci. Barley Mla1 expressed in wheat using the single-cell transformation system failed to trigger a response to any of the wheat powdery mildew Avr genes tested, indicating that AvrMla1 is not genetically fixed in wheat mildew strains.

Recognition Specificity and RAR1/SGT1 Dependence in Barley Mla Disease Resistance Genes to the Powdery Mildew Fungus

“…Although Mla12 might be an allele of Mla1 and Mla6 , it differs from them by belonging to a subgroup of Mla variants that trigger delayed resistance responses (Freialdenhoven et al, 1994;Boyd et al, 1995). Using a single-cell transient gene expression assay (Shirasu et al, 1999b;Zhou et al, 2001), we demonstrate that Mla12 overexpression shifts the slow Mla12 -triggered response to a rapid Mla1 / Mla6 -like resistance. We gained insights into structurefunction relationships of MLA proteins by analyzing a series of reciprocal domain swaps between MLA1 and MLA6.…”

“…Two clones, designated Sp14-1 and Sp14-4, contain identical RGHs showing ‫ف‬ 90% sequence identity to Mla1 and Mla6 in deduced exon and intron sequences. A closer comparison of the NB-LRR gene in Sp14-4 with Mla1 and Mla6 revealed an identical 5 Ј untranslated small open reading frame of nine amino acids and the same intron-exon structure (Halterman et al, 2001;Zhou et al, 2001). These genes share a simple sequence repeat (AT) n in intron 3, although the exact numbers of the repeats differ (see Figure 6 below).…”

“…The RGHs belong to three dissimilar families sharing Ͻ 43% amino acid sequence similarity between families (Wei et al, 1999(Wei et al, , 2002. Because Morex lacks a known Mla resistance specificity, the first two identified Mla powdery mildew R genes, Mla1 and Mla6 , were isolated from other barley accessions (Halterman et al, 2001;Zhou et al, 2001). The deduced proteins share 91% identical residues and show highest overall similarity to the deduced Morex RGH1bcd family member (83 and 79% identity to MLA1 and MLA6, respectively) (Halterman et al, 2001;Wei et al, 2002).…”

Genetically engineered stem rust resistance in barley using the Rpg1 gene

“…The Mla1 gene conferring resistance to AvrMla1 of B. graminis f. sp. hordei is an R gene encoding an N-terminal coiled-coil (CC) protein structure as well as NBS and LRR domains (25). It was pinpointed by isolating a ␥-ray induced susceptible mutant with an Mla deletion, identifying a clone corresponding to the deletion in a cosmid library, and carrying out a functional test with the cosmid subclones by using a three-component epidermal single-cell expression system.…”

Map-based isolation of the leaf rust disease resistance gene Lr10 from the hexaploid wheat ( Triticum aestivum L.) genome

“…Mlo, the first barley disease resistance gene isolated by map-based cloning (3), was identified through the use of a yeast artificial chromosome (YAC) library and the subsequent construction of a bacterial artificial chromosome (BAC) library from the YAC clone spanning the resistance locus. More recently, the construction of a BAC library from the barley cultivar Morex (4) has greatly facilitated the map-based isolation of the powdery mildew resistance genes Mla1 and Mla6 (5,6) as well as the stem rust resistance gene Rpg1 (7). In wheat, BAC libraries have been constructed from the diploid species Triticum monococcum (8) and Ae.…”