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Evolution and selection of Rhg1, a copy-number variant nematode-resistance locus.

ABSTRACT: The soybean cyst nematode (SCN) resistance locus Rhg1 is a tandem repeat of a 31.2 kb unit of the soybean genome. Each 31.2-kb unit contains four genes. One allele of Rhg1, Rhg1-b, is responsible for protecting most US soybean production from SCN. Whole-genome sequencing was performed, and PCR assays were developed to investigate allelic variation in sequence and copy number of the Rhg1 locus across a population of soybean germplasm accessions. Four distinct sequences of the 31.2-kb repeat unit were identified, and some Rhg1 alleles carry up to three different types of repeat unit. The total number of copies of the repeat varies from 1 to 10 per haploid genome. Both copy number and sequence of the repeat correlate with the resistance phenotype, and the Rhg1 locus shows strong signatures of selection. Significant linkage disequilibrium in the genome outside the boundaries of the repeat allowed the Rhg1 genotype to be inferred using high-density single nucleotide polymorphism genotyping of 15 996 accessions. Over 860 germplasm accessions were found likely to possess Rhg1 alleles. The regions surrounding the repeat show indications of non-neutral evolution and high genetic variability in populations from different geographic locations, but without evidence of fixation of the resistant genotype. A compelling explanation of these results is that balancing selection is in operation at Rhg1.

SUBMITTER: Lee TG

PROVIDER: S-EPMC4413360 | biostudies-literature | 2015 Apr

REPOSITORIES: biostudies-literature

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Evolution and selection of Rhg1, a copy-number variant nematode-resistance locus.

Lee Tong Geon TG Kumar Indrajit I Diers Brian W BW Hudson Matthew E ME

Molecular ecology 20150330 8

The soybean cyst nematode (SCN) resistance locus Rhg1 is a tandem repeat of a 31.2 kb unit of the soybean genome. Each 31.2-kb unit contains four genes. One allele of Rhg1, Rhg1-b, is responsible for protecting most US soybean production from SCN. Whole-genome sequencing was performed, and PCR assays were developed to investigate allelic variation in sequence and copy number of the Rhg1 locus across a population of soybean germplasm accessions. Four distinct sequences of the 31.2-kb repeat unit ...[more]

PMID: 25735447

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Project description:Soybean growers widely use the Resistance to Heterodera glycines 1 (Rhg1) locus to reduce yield losses caused by soybean cyst nematode (SCN). Rhg1 is a tandemly repeated four gene block. Two classes of SCN resistance-conferring Rhg1 haplotypes are recognized: rhg1-a ("Peking-type," low-copy number, three or fewer Rhg1 repeats) and rhg1-b ("PI 88788-type," high-copy number, four or more Rhg1 repeats). The rhg1-a and rhg1-b haplotypes encode α-SNAP (alpha-Soluble NSF Attachment Protein) variants α-SNAP Rhg1 LC and α-SNAP Rhg1 HC, respectively, with differing atypical C-terminal domains, that contribute to SCN resistance. Here we report that rhg1-a soybean accessions harbor a copia retrotransposon within their Rhg1 Glyma.18G022500 (α-SNAP-encoding) gene. We termed this retrotransposon "RAC," for Rhg1 alpha-SNAP copia. Soybean carries multiple RAC-like retrotransposon sequences. The Rhg1 RAC insertion is in the Glyma.18G022500 genes of all true rhg1-a haplotypes we tested and was not detected in any examined rhg1-b or Rhg1WT (single-copy) soybeans. RAC is an intact element residing within intron 1, anti-sense to the rhg1-a α-SNAP open reading frame. RAC has intrinsic promoter activities, but overt impacts of RAC on transgenic α-SNAP Rhg1 LC mRNA and protein abundance were not detected. From the native rhg1-a RAC+ genomic context, elevated α-SNAP Rhg1 LC protein abundance was observed in syncytium cells, as was previously observed for α-SNAP Rhg1 HC (whose rhg1-b does not carry RAC). Using a SoySNP50K SNP corresponding with RAC presence, just ~42% of USDA accessions bearing previously identified rhg1-a SoySNP50K SNP signatures harbor the RAC insertion. Subsequent analysis of several of these putative rhg1-a accessions lacking RAC revealed that none encoded α-SNAPRhg1LC, and thus, they are not rhg1-a. rhg1-a haplotypes are of rising interest, with Rhg4, for combating SCN populations that exhibit increased virulence against the widely used rhg1-b resistance. The present study reveals another unexpected structural feature of many Rhg1 loci, and a selectable feature that is predictive of rhg1-a haplotypes.

Project description:BackgroundSoybean cyst nematode (Heterodera glycines, SCN) is the most economically damaging pathogen of soybean (Glycine max) in the U.S. The Rhg1 locus is repeatedly observed as the quantitative trait locus with the greatest impact on SCN resistance. The Glyma18g02680.1 gene at the Rhg1 locus that encodes an apparent leucine-rich repeat transmembrane receptor-kinase (LRR-kinase) has been proposed to be the SCN resistance gene, but its function has not been confirmed. Generation of fertile transgenic soybean lines is difficult but methods have been published that test SCN resistance in transgenic roots generated with Agrobacterium rhizogenes.ResultsWe report use of artificial microRNA (amiRNA) for gene silencing in soybean, refinements to transgenic root SCN resistance assays, and functional tests of the Rhg1 locus LRR-kinase gene. A nematode demographics assay monitored infecting nematode populations for their progress through developmental stages two weeks after inoculation, as a metric for SCN resistance. Significant differences were observed between resistant and susceptible control genotypes. Introduction of the Rhg1 locus LRR-kinase gene (genomic promoter/coding region/terminator; Peking/PI 437654-derived SCN-resistant source), into rhg1- SCN-susceptible plant lines carrying the resistant-source Rhg4+ locus, provided no significant increases in SCN resistance. Use of amiRNA to reduce expression of the LRR-kinase gene from the Rhg1 locus of Fayette (PI 88788 source of Rhg1) also did not detectably alter resistance to SCN. However, silencing of the LRR-kinase gene did have impacts on root development.ConclusionThe nematode demographics assay can expedite testing of transgenic roots for SCN resistance. amiRNAs and the pSM103 vector that drives interchangeable amiRNA constructs through a soybean polyubiqutin promoter (Gmubi), with an intron-GFP marker for detection of transgenic roots, may have widespread use in legume biology. Studies in which expression of the Rhg1 locus LRR-kinase gene from different resistance sources was either reduced or complemented did not reveal significant impacts on SCN resistance.

Project description:Two coding variants in the APOL1 gene (G1 and G2) explain most of the high rate of kidney disease in African Americans. APOL1-associated kidney disease risk inheritance follows an autosomal recessive pattern: The relative risk of kidney disease associated with inheritance of two high-risk variants is 7-30 fold, depending on the specific kidney phenotype. We wished to determine if the variability in phenotype might in part reflect structural differences in APOL1 gene. We analyzed sequence coverage from 1000 Genomes Project Phase 3 samples as well as exome sequencing data from African American kidney disease cases for copy number variation. 8 samples sequenced in the 1000 Genomes Project showed increased coverage over a ~100kb region that includes APOL2, APOL1 and part of MYH9, suggesting the presence of APOL1 copy number greater than 2. We reasoned that such duplications should be enriched in apparent G1 heterozygotes with kidney disease. Using a PCR-based assay, we observed the presence of this duplication in additional samples from apparent G0G1 or G0G2 individuals. The frequency of this APOL1 duplication was compared among cases (n = 123) and controls (n = 255) with apparent G0G1 heterozygosity. The presence of APOL1 duplication was observed in 4.06% of cases and 0.78% controls, preliminary evidence that this APOL1 duplication may alter susceptibility to kidney disease (p = 0.03). Taqman-based copy number assays confirmed the presence of 3 APOL1 copies in individuals positive for this specific duplication by PCR assay, but also identified a small number of individuals with additional APOL1 copies of presumably different structure. These observations motivate further studies to better assess the contribution of APOL1 copy number on kidney disease risk and on APOL1 function. Investigators and clinicians genotyping APOL1 should also consider whether the particular genotyping platform used is subject to technical errors when more than two copies of APOL1 are present.

Dataset Information

Evolution and selection of Rhg1, a copy-number variant nematode-resistance locus.

Publications

Evolution and selection of Rhg1, a copy-number variant nematode-resistance locus.

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