Novel induced mlo mutant alleles in combination with site-directed mutagenesis reveal functionally important domains in the heptahelical barley Mlo protein

Abstract

BackgroundRecessively inherited natural and induced mutations in the barley Mlo gene confer durable broad-spectrum resistance against the powdery mildew pathogen, Blumeria graminis f.sp. hordei. Mlo codes for a member of a plant-specific family of polytopic integral membrane proteins with unknown biochemical activity. Resistant barley mlo mutant alleles identify amino acid residues that are critical for Mlo function in the context of powdery mildew susceptibility.ResultsWe molecularly analyzed a novel set of induced barley mlo mutants and used site-directed mutagenesis in combination with transient gene expression to unravel novel amino acid residues of functional significance. We integrate these results with previous findings to map functionally important regions of the heptahelical Mlo protein. Our data reveal the second and third cytoplasmic loop as being particularly sensitive to functional impediment by mutational perturbation, suggesting that these regions are critical for the susceptibility-conferring activity of the Mlo protein. In contrast, only mutations in the second but not the third cytoplasmic loop appear to trigger the Endoplasmic Reticulum-localized quality control machinery that ensures the biogenesis of properly folded membrane proteins.ConclusionOur findings identify functionally important regions of the polytopic barley Mlo protein and reveal the differential sensitivity of individual protein domains to cellular quality control.

Genome-wide Identification, Classification and Expression Analysis of the Mildew Resistance Locus O ( MLO ) Gene Family in Sweet Orange ( Citrus sinensis )

“…Interestingly, when displayed in the evolutionary trees of MLO protein family, all MLO isoforms from dicots experimentally shown to be necessary for PM immunity group in the clade V in the scientific documents [ [8][9][10][20][21][22] ]. Furthermore, clade IV harbors all featured monocot PM perceptivity proteins [ 23 ]. These show that phylogenetic studies on MLO proteins may predict factor candidates for PM perceptivity.…”

Expression Profiling of MLO Family Genes under Podosphaera xanthii Infection and Exogenous Application of Phytohormones in Cucumis melo L.

“…1 a), which might facilitate PM resistance conferred by CmMLO genes. As the second and third cytoplasmic loops of MLO proteins play a critical role in powdery mildew susceptibility [40], any mutation of these positions might be involved in PM resistance.…”

“…The invariable cysteine and proline residues in extracellular loops or in TM domains are necessary for MLO protein function and/or durability [11]. In addition, second and third cytoplasmic loops in MLO proteins are important for PM susceptibility [40]. Moreover, histological reports…”

Identification of novel Mlo family members in wheat and their genetic characterization

“…The accumulation of the mlo transcript was also detected in rice infected with blast fungi and in wheat injected with powdery mildew-derived carbohydrate (Piffanelli et al, 2002). Recently, using Mlo mutant alleles in barley, Reinstädler et al (2010) revealed the regions of the MLO protein that are functionally important for resistance to powdery mildew. However, microarray analyses found that not only the Mlo genes but also more than 300 other genes showed expressional changes after powdery mildew infection in barley, and some of them were…”