Biology subjects

Dilkes, B. P.

Publications and source records attributed to Dilkes, B. P..

4 recordsLinked to original sources

bioRxiv2018

Adult Plant Resistance in Maize to Northern Leaf Spot Is a Feature of Partial Loss-of-function Alleles of Hm1

Adult plant resistance (APR) is an enigmatic phenomenon in which resistance genes are ineffective in protecting seedlings from disease but confer robust resistance at maturity. Maize has multiple cases in which genes confer APR to northern leaf spot, a lethal disease caused by Cochliobolus carbonum race 1 (CCR1). The first identified case of APR in maize is encoded by a hypomorphic allele, Hm1A, at the hm1 locus. In contrast, wild type alleles of hm1 provide complete protection at all developmental stages and in every part of the maize plant. Hm1 encodes an NADPH-dependent reductase, which inactivates HC-toxin, a key virulence effector of CCR1. Cloning and characterization of Hm1A ruled out differential transcription or translation for its APR phenotype and identified an amino acid substitution that reduced HC-toxin reductase (HCTR) activity. The possibility of a causal relationship between the weak nature of Hm1A and its APR phenotype was confirmed by the generation of two new APR alleles of Hm1 by mutagenesis. The HCTRs encoded by these new APR alleles had undergone relatively conservative missense changes that partially reduced their enzymatic activity similar to HM1A. No difference in accumulation of HCTR was observed between adult and juvenile plants, suggesting that the susceptibility of seedlings derives from a greater need for HCTR activity, not reduced accumulation of the gene product. Conditions and treatments that altered the photosynthetic output of the host had a dramatic effect on resistance imparted by the APR alleles, demonstrating a link between the energetic or metabolic status of the host and disease resistance affected by HC-toxin catabolism by the APR alleles of HCTR.\n\nAUTHOR SUMMARYAdult plant resistance (APR) is a phenomenon in which disease resistance genes are able to confer resistance at the adult stages of the plant but somehow fail to do so at the seedling stages. Despite the widespread occurrence of APR in various plant diseases, the mechanism underlying this trait remains obscure. It is not due to the differential transcription of these genes, and here we show that it is also not due to the differential translation or activity of the APR alleles of the maize hm1 gene at different stages of development. Using a combination of molecular genetics, biochemistry and physiology, we present multiple lines of evidence that demonstrate that APR is a feature or symptom of weak forms of resistance. While the mature parts of the plant are metabolically robust enough to manifest resistance, seedling tissues are not, leaving them vulnerable to disease. Growth conditions that compromise the photosynthetic output of the plant further deteriorate the ability of the seedlings to protect themselves from pathogens.\n\nOne sentence summaryCharacterization of adult plant resistance in the maize-CCR1 pathosystem reveals a causal link between weak resistance and APR.

pathology↗

bioRxiv2017

A Very Oil Yellow1 modifier of the Oil Yellow1-N1989 allele uncovers a cryptic phenotypic impact of cis-regulatory variation in maize

Forward genetics determines the function of genes underlying trait variation by identifying the change in DNA responsible for changes in phenotype. Detecting phenotypically-relevant variation outside protein coding sequences and distinguishing this from neutral variants is not trivial; partly because the mechanisms by which DNA polymorphisms in the intergenic regions affect gene regulation are poorly understood. Here we utilized a dominant genetic marker with a convenient phenotype to investigate the effect of cis and trans-acting regulatory variation. We performed a forward genetic screen for natural variation that suppress or enhance the semi-dominant mutant allele Oy1-N1989, encoding the magnesium chelatase subunit I of maize. This mutant permits rapid phenotyping of leaf color as a reporter for chlorophyll accumulation, and mapping of natural variation in maize affecting chlorophyll metabolism. We identified a single modifier locus segregating between B73 and Mo17 that was linked to the reporter gene itself, which we call very oil yellow1. Based on the variation in OY1 transcript abundance and genome-wide association data, vey1 is predicted to consist of multiple cis-acting regulatory sequence polymorphisms encoded at the wild-type oy1 alleles. The vey1 allele appears to be a common polymorphism in the maize germplasm that alters the expression level of a key gene in chlorophyll biosynthesis. These vey1 alleles have no discernable impact on leaf chlorophyll in the absence of the Oy1-N1989 reporter. Thus, use of a mutant as a simple and efficient reporter for magnesium chelatase activity resulted in the detection of expression-level polymorphisms not readily visible in the laboratory.

genetics↗

bioRxiv2017

Phytohormone Inhibitor Treatments Phenocopy Brassinosteroid And Gibberellin Dwarf Mutant Interactions In Maize

Phytohormone biosynthesis produces metabolites with profound effects on plant growth and development. Modulation of hormone levels during developmental events, in response to the environment, by genetic polymorphism, or by chemical application can reveal the plant processes most responsive to a phytohormone. In many cases, chemical inhibitors are applied and the levels of specific phytohormones are measured to determine if, and which, phytohormone is affected by a molecule. In many cases, the sensitivity of biochemical testing has determined multiple pathways affected by a single inhibitor. Genetic studies are not subject to this problem, and a wealth of data about the morphological impacts of hormone biosynthetic inhibition has accumulated through the study of enzyme mutants. We previously identified a complex interplay between brassinosteroid (BR) and gibberellin (GA) in maize, where the interdependence of the two differs dependent on the developmental context. We found that: GA is required for loss of BR to induce retained pistils in the tassel florets (POPIT); BR is required for the loss of GA to induce tiller outgrowth; BR and GA are additive for plant height; BR has no effect on the induction of anther retention in ear florets of GA mutants. In this work, we sought to assess the specificity of three triazole inhibitors of cytochrome P450s by determining their abilities to recapitulate the phenotype of double mutants. The GA biosynthetic inhibitors uniconazole (UCZ) and paclobutrazol (PAC) were applied to the BR biosynthetic mutant na2 and all double mutant phenotypes were recovered in the UCZ treatment. PAC was unable to suppress the retention of pistils in the tassels of nana plant2 (na2) mutant plants. The BR biosynthetic inhibitor propiconazole (PCZ) suppressed tiller outgrowth in the GA biosynthetic mutant dwarf5 (d5). All treatments were additive with genetic mutants for effects on plant height. Due to additional measurements done here but not in previous studies of the double mutants, we detected new interactions between GA and BR biosynthesis affecting plastochron index and tassel branching. These experiments, a refinement of our previous model, and a discussion of the extension of this type of work are presented.

plant biology↗

bioRxiv2016

Whole genome sequence accuracy is improved by replication in a population of mutagenized sorghum.

The accurate detection of induced mutations is critical for both forward and reverse genetics studies. Experimental chemical mutagenesis induces relatively few single base changes per individual. In a complex eukaryotic genome, false positive detection of mutations can occur at or above this mutagenesis rate. We demonstrate here, using a population of ethyl methanesulfonate (EMS) treated Sorghum bicolor BTx623 individuals, that using replication to detect false positive induced variants in next-generation sequencing data permits higher throughput variant detection with greater accuracy. We used a lower sequence coverage depth (average of 7X) from 586 independently mutagenized individuals and detected 5,399,493 homozygous SNPs. Of these, 76% originated from only 57,872 genomic positions prone to false positive variant calling. These positions are characterized by high copy number paralogs where the error-prone SNP positions are at copies containing a variant at the SNP position. The ability of short stretches of homology to generate these error prone positions suggests that incompletely assembled or poorly mapped repeated sequences are one driver of these error prone positions. Removal of these false positives left 1,275,872 homozygous and 477,531 heterozygous EMS-induced SNPs which, congruent with the mutagenic mechanism of EMS, were greater than 98% G:C to A:T transitions. Through this analysis we generated a database of sequence indexed mutants of Sorghum. This collection contains 4,035 high impact homozygous mutations in 3,637 genes and 56,514 homozygous missense mutations in 23,227 genes. Each line contains, on average, 2,177 annotated homozygous SNPs per genome, including seven likely gene knockouts and 96 missense mutations. The number of mutations in a transcript was linearly correlated with the transcript length and also the G+C count, but not with the GC/AT ratio. Analysis of the detected mutagenized positions identified CG-rich patches, and flanking sequences strongly influenced EMS-induced mutation rates. Our method for detecting false-positive induced mutations is generally applicable to any organism, is independent of the choice of in silico variant-calling algorithm, and is most valuable when the true mutation rate is likely to be low, such as in laboratory induced mutations or somatic mutation detection in medicine.

genetics↗