Two more families supporting the existence of monogenic spinocerebellar ataxia 48.

The reduced penetrance of TBP intermediate alleles and the recently proposed possible digenic TBP/STUB1 inheritance raised questions on the possible mechanism involved opening a debate on the existence of SCA48 as a monogenic disorder. We here report clinical and genetic results of two apparently unrelated patients carrying the same STUB1 variant(c.244G > T;p.Asp82Tyr) with normal TBP alleles and a clinical picture fully resembling SCA48, including cerebellar ataxia, dysarthria and mild cognitive impairment. This report provides supportive evidence that this specific ataxia can also occur as a monogenic disease, considering classical TBP allelic ranges.

Recessive MECR pathogenic variants cause an LHON-like optic neuropathy.

BACKGROUND: Leber's hereditary optic neuropathy (LHON) is a mitochondrial disorder characterised by complex I defect leading to sudden degeneration of retinal ganglion cells. Although typically associated with pathogenic variants in mitochondrial DNA, LHON was recently described in patients carrying biallelic variants in nuclear genes DNAJC30, NDUFS2 and MCAT. MCAT is part of mitochondrial fatty acid synthesis (mtFAS), as also MECR, the mitochondrial trans-2-enoyl-CoA reductase. MECR mutations lead to a recessive childhood-onset syndromic disorder with dystonia, optic atrophy and basal ganglia abnormalities. METHODS: We studied through whole exome sequencing two sisters affected by sudden and painless visual loss at young age, with partial recovery and persistent central scotoma. We modelled the candidate variant in yeast and studied mitochondrial dysfunction in yeast and fibroblasts. We tested protein lipoylation and cell response to oxidative stress in yeast. RESULTS: Both sisters carried a homozygous pathogenic variant in MECR (p.Arg258Trp). In yeast, the MECR-R258W mutant showed an impaired oxidative growth, 30% reduction in oxygen consumption rate and 80% decrease in protein levels, pointing to structure destabilisation. Fibroblasts confirmed the reduced amount of MECR protein, but failed to reproduce the OXPHOS defect. Respiratory complexes assembly was normal. Finally, the yeast mutant lacked lipoylation of key metabolic enzymes and was more sensitive to H2O2 treatment. Lipoic Acid supplementation partially rescued the growth defect. CONCLUSION: We report the first family with homozygous MECR variant causing an LHON-like optic neuropathy, which pairs the recent MCAT findings, reinforcing the impairment of mtFAS as novel pathogenic mechanism in LHON.

Genomic tools for durum wheat breeding: de novo assembly of Svevo transcriptome and SNP discovery in elite germplasm.

BACKGROUND: The tetraploid durum wheat (Triticum turgidum L. ssp. durum Desf. Husnot) is an important crop which provides the raw material for pasta production and a valuable source of genetic diversity for breeding hexaploid wheat (Triticum aestivum L.). Future breeding efforts to enhance yield potential and climate resilience will increasingly rely on genomics-based approaches to identify and select beneficial alleles. A deeper characterisation of the molecular and functional diversity of the durum wheat transcriptome will be instrumental to more effectively harness its genetic diversity. RESULTS: We report on the de novo transcriptome assembly of durum wheat cultivar 'Svevo'. The transcriptome of four tissues/organs (shoots and roots at the seedling stage, reproductive organs and developing grains) was assembled de novo, yielding 180,108 contigs, with a N50 length of 1121 bp and mean contig length of 883 bp. Alignment against the transcriptome of nine plant species identified 43% of transcripts with homology to at least one reference transcriptome. The functional annotation was completed by means of a combination of complementary software. The presence of differential expression between the A- and B-homoeolog copies of the durum wheat tetraploid genome was ascertained by phase reconstruction of polymorphic sites based on the T. urartu transcripts and inferring homoeolog-specific sequences. We observed greater expression divergence between A and B homoeologs in grains rather than in leaves and roots. The transcriptomes of 13 durum wheat cultivars spanning the breeding period from 1969 to 2005 were analysed for SNP diversity, leading to 95,358 non-rare, hemi-SNPs shared among two or more cultivars and 33,747 locus-specific (diploid inheritance) SNPs. CONCLUSIONS: Our study updates and expands the de novo transcriptome reference assembly available for durum wheat. Out of 180,108 assembled transcripts, 13,636 were specific to the Svevo cultivar as compared to the only other reference transcriptome available for durum, thus contributing to the identification of the tetraploid wheat pan-transcriptome. Additionally, the analysis of 13 historically relevant hallmark varieties produced a SNP dataset that could successfully validate the genotyping in tetraploid wheat and provide a valuable resource for genomics-assisted breeding of both tetraploid and hexaploid wheats.

Genome-wide association study on stem rust resistance in Kazakh spring barley lines.

BACKGROUND: Stem rust (SR) is one of the most economically devastating barley diseases in Kazakhstan, and in some years it causes up to 50 % of yield losses. Routine conventional breeding for resistance to stem rust is almost always in progress in all Kazakhstan breeding stations. However, molecular marker based approach towards new SR resistance genes identification and relevant marker-assisted selection had never been employed in Kazakhstan yet. In this study, as a preliminary step the GWAS (genome-wide association study) mapping was applied in attempt to identify reliable SNP markers and quantitative trait loci (QTL) associated with resistance to SR. RESULTS: Barley collection of 92 commercial cultivars and promising lines was genotyped using a high-throughput single nucleotide polymorphism (9,000 SNP) Illumina iSelect array. 6,970 SNPs out of 9,000 total were polymorphic and scorable. 5,050 SNPs out of 6,970 passed filtering threshold and were used for association mapping (AM). All 92 accessions were phenotyped for resistance to SR by observing adult plants in artificially infected plots at the Research Institute for Biological Safety Problems in Dzhambul region of Kazakhstan. GLM analysis allowed the identification of 15 SNPs associated with the resistance at the heading time (HA) phase, and 2 SNPs at the seed's milky-waxy maturity (SM) phase. However, after application of 5 % Bonferroni multiple test correction, only 2 SNPs at the HA stage on the same position of chromosome 6H can be claimed as reliable markers for SR resistance. The MLM analysis after the Bonferroni correction did not reveal any associations in this study, although distribution lines in the quantile-quantile (QQ) plot indicates that overcorrection in the test due to both Q and K matrices usage. CONCLUSIONS: Obtained data suggest that genome wide genotyping of 92 spring barley accessions from Kazakhstan with 9 K Illumina SNP array was highly efficient. Linkage disequilibrium based mapping approach allowed the identification of highly significant QTL for the SR resistance at the HA phase of growth on chromosome 6H. On the other hand, no significant QTL was detected at the SM phase, assuming that for a successful GWASmapping experiment a larger size population with more diverse genetic background should be tested. Obtained results provide additional information towards better understanding of SR resistance in barley.

AFG3L2 and ACO2-Linked Dominant Optic Atrophy: Genotype-Phenotype Characterization Compared to OPA1 Patients.

PURPOSE: Heterozygous mutations in the AFG3L2 gene (encoding a mitochondrial protease indirectly reflecting on OPA1 cleavage) and ACO2 gene (encoding the mitochondrial enzyme aconitase) are associated with isolated forms of Dominant Optic Atrophy (DOA). We aimed at describing their neuro-ophthalmological phenotype as compared with classic OPA1-related DOA. DESIGN: Cross-sectional study. METHODS: The following neuro-ophthalmological parameters were collected: logMAR visual acuity (VA), color vision, mean deviation and foveal threshold at visual fields, average and sectorial retinal nerve fiber layer (RNFL), and ganglion cell layer (GCL) thickness on optical coherence tomography. ACO2 and AFG3L2 patients were compared with an age- and sex-matched group of OPA1 patients with a 1:2 ratio. All eyes were analyzed using a clustered Wilcoxon rank sum test with the Rosner-Glynn-Lee method. RESULTS: A total of 44 eyes from 23 ACO2 patients and 26 eyes from 13 AFG3L2 patients were compared with 143 eyes from 72 OPA1 patients. All cases presented with bilateral temporal-predominant optic atrophy with various degree of visual impairment. Comparison between AFG3L2 and OPA1 failed to reveal any significant difference. ACO2 patients compared to both AFG3L2 and OPA1 presented overall higher values of nasal RNFL thickness (P = .029, P = .023), average thickness (P = .012, P = .0007), and sectorial GCL thickness. These results were confirmed also comparing separately affected and subclinical patients. CONCLUSIONS: Clinically, DOA remains a fairly homogeneous entity despite the growing genetic heterogeneity. ACO2 seems to be associated with an overall better preservation of retinal ganglion cells, probably depending on the different pathogenic mechanism involving mtDNA maintenance, as opposed to AFG3L2, which is involved in OPA1 processing and is virtually indistinguishable from classic OPA1-DOA.

The genetic puzzle of a SOD1-patient with ocular ptosis and a motor neuron disease: a case report.

Amyotrophic lateral sclerosis (ALS) is a neurodegenerative disease with a complex genetic architecture, showing monogenic, oligogenic, and polygenic inheritance. In this study, we describe the case of a 71 years-old man diagnosed with ALS with atypical clinical features consisting in progressive ocular ptosis and sensorineural deafness. Genetic analyses revealed two heterozygous variants, in the SOD1 (OMIM*147450) and the TBK1 (OMIM*604834) genes respectively, and furthermore mitochondrial DNA (mtDNA) sequencing identified the homoplasmic m.14484T>C variant usually associated with Leber's Hereditary Optic Neuropathy (LHON). We discuss how all these variants may synergically impinge on mitochondrial function, possibly contributing to the pathogenic mechanisms which might ultimately lead to the neurodegenerative process, shaping the clinical ALS phenotype enriched by adjunctive clinical features.

Dissecting the multifaceted contribution of the mitochondrial genome to autism spectrum disorder.

Autism spectrum disorder (ASD) is a clinically heterogeneous class of neurodevelopmental conditions with a strong, albeit complex, genetic basis. The genetic architecture of ASD includes different genetic models, from monogenic transmission at one end, to polygenic risk given by thousands of common variants with small effects at the other end. The mitochondrial DNA (mtDNA) was also proposed as a genetic modifier for ASD, mostly focusing on maternal mtDNA, since the paternal mitogenome is not transmitted to offspring. We extensively studied the potential contribution of mtDNA in ASD pathogenesis and risk through deep next generation sequencing and quantitative PCR in a cohort of 98 families. While the maternally-inherited mtDNA did not seem to predispose to ASD, neither for haplogroups nor for the presence of pathogenic mutations, an unexpected influence of paternal mtDNA, apparently centered on haplogroup U, came from the Italian families extrapolated from the test cohort (n = 74) when compared to the control population. However, this result was not replicated in an independent Italian cohort of 127 families and it is likely due to the elevated paternal age at time of conception. In addition, ASD probands showed a reduced mtDNA content when compared to their unaffected siblings. Multivariable regression analyses indicated that variants with 15%-5% heteroplasmy in probands are associated to a greater severity of ASD based on ADOS-2 criteria, whereas paternal super-haplogroups H and JT were associated with milder phenotypes. In conclusion, our results suggest that the mtDNA impacts on ASD, significantly modifying the phenotypic expression in the Italian population. The unexpected finding of protection induced by paternal mitogenome in term of severity may derive from a role of mtDNA in influencing the accumulation of nuclear de novo mutations or epigenetic alterations in fathers' germinal cells, affecting the neurodevelopment in the offspring. This result remains preliminary and needs further confirmation in independent cohorts of larger size. If confirmed, it potentially opens a different perspective on how paternal non-inherited mtDNA may predispose or modulate other complex diseases.

The Global Durum Wheat Panel (GDP): An International Platform to Identify and Exchange Beneficial Alleles.

Representative, broad and diverse collections are a primary resource to dissect genetic diversity and meet pre-breeding and breeding goals through the identification of beneficial alleles for target traits. From 2,500 tetraploid wheat accessions obtained through an international collaborative effort, a Global Durum wheat Panel (GDP) of 1,011 genotypes was assembled that captured 94-97% of the original diversity. The GDP consists of a wide representation of Triticum turgidum ssp. durum modern germplasm and landraces, along with a selection of emmer and primitive tetraploid wheats to maximize diversity. GDP accessions were genotyped using the wheat iSelect 90K SNP array. Among modern durum accessions, breeding programs from Italy, France and Central Asia provided the highest level of genetic diversity, with only a moderate decrease in genetic diversity observed across nearly 50 years of breeding (1970-2018). Further, the breeding programs from Europe had the largest sets of unique alleles. LD was lower in the landraces (0.4 Mbp) than in modern germplasm (1.8 Mbp) at r 2 = 0.5. ADMIXTURE analysis of modern germplasm defined a minimum of 13 distinct genetic clusters (k), which could be traced to the breeding program of origin. Chromosome regions putatively subjected to strong selection pressure were identified from fixation index (F st ) and diversity reduction index (DRI) metrics in pairwise comparisons among decades of release and breeding programs. Clusters of putative selection sweeps (PSW) were identified as co-localized with major loci controlling phenology (Ppd and Vrn), plant height (Rht) and quality (gliadins and glutenins), underlining the role of the corresponding genes as driving elements in modern breeding. Public seed availability and deep genetic characterization of the GDP make this collection a unique and ideal resource to identify and map useful genetic diversity at loci of interest to any breeding program.

Durum wheat genome highlights past domestication signatures and future improvement targets.

The domestication of wild emmer wheat led to the selection of modern durum wheat, grown mainly for pasta production. We describe the 10.45 gigabase (Gb) assembly of the genome of durum wheat cultivar Svevo. The assembly enabled genome-wide genetic diversity analyses revealing the changes imposed by thousands of years of empirical selection and breeding. Regions exhibiting strong signatures of genetic divergence associated with domestication and breeding were widespread in the genome with several major diversity losses in the pericentromeric regions. A locus on chromosome 5B carries a gene encoding a metal transporter (TdHMA3-B1) with a non-functional variant causing high accumulation of cadmium in grain. The high-cadmium allele, widespread among durum cultivars but undetected in wild emmer accessions, increased in frequency from domesticated emmer to modern durum wheat. The rapid cloning of TdHMA3-B1 rescues a wild beneficial allele and demonstrates the practical use of the Svevo genome for wheat improvement.

Fine-scale spatial genetic structure in predominantly selfing plants with limited seed dispersal: A rule or exception?

Gene flow at a fine scale is still poorly understood despite its recognized importance for plant population demographic and genetic processes. We tested the hypothesis that intensity of gene flow will be lower and strength of spatial genetic structure (SGS) will be higher in more peripheral populations because of lower population density. The study was performed on the predominantly selfing Avena sterilis and included: (1) direct measurement of dispersal in a controlled environment; and (2) analyses of SGS in three natural populations, sampled in linear transects at fixed increasing inter-plant distances. We found that in A. sterilis major seed dispersal is by gravity in close (less than 2 m) vicinity of the mother plant, with a minor additional effect of wind. Analysis of SGS with six nuclear SSRs revealed a significant autocorrelation for the distance class of 1 m only in the most peripheral desert population, while in the two core populations with Mediterranean conditions, no genetic structure was found. Our results support the hypothesis that intensity of SGS increases from the species core to periphery as a result of decreased within-population gene flow related to low plant density. Our findings also show that predominant self-pollination and highly localized seed dispersal lead to SGS at a very fine scale, but only if plant density is not too high.

Role of phenotypic plasticity and population differentiation in adaptation to novel environmental conditions.

Species can adapt to new environmental conditions either through individual phenotypic plasticity, intraspecific genetic differentiation in adaptive traits, or both. Wild emmer wheat, Triticum dicoccoides, an annual grass with major distribution in Eastern Mediterranean region, is predicted to experience in the near future, as a result of global climate change, conditions more arid than in any part of the current species distribution. To understand the role of the above two means of adaptation, and the effect of population range position, we analyzed reaction norms, extent of plasticity, and phenotypic selection across two experimental environments of high and low water availability in two core and two peripheral populations of this species. We studied 12 quantitative traits, but focused primarily on the onset of reproduction and maternal investment, which are traits that are closely related to fitness and presumably involved in local adaptation in the studied species. We hypothesized that the population showing superior performance under novel environmental conditions will either be genetically differentiated in quantitative traits or exhibit higher phenotypic plasticity than the less successful populations. We found the core population K to be the most plastic in all three trait categories (phenology, reproductive traits, and fitness) and most successful among populations studied, in both experimental environments; at the same time, the core K population was clearly genetically differentiated from the two edge populations. Our results suggest that (1) two means of successful adaptation to new environmental conditions, phenotypic plasticity and adaptive genetic differentiation, are not mutually exclusive ways of achieving high adaptive ability; and (2) colonists from some core populations can be more successful in establishing beyond the current species range than colonists from the range extreme periphery with conditions seemingly closest to those in the new environment.

Multi-approaches analysis reveals local adaptation in the emmer wheat (Triticum dicoccoides) at macro- but not micro-geographical scale.

Detecting local adaptation and its spatial scale is one of the most important questions of evolutionary biology. However, recognition of the effect of local selection can be challenging when there is considerable environmental variation across the distance at the whole species range. We analyzed patterns of local adaptation in emmer wheat, Triticum dicoccoides, at two spatial scales, small (inter-population distance less than one km) and large (inter-population distance more than 50 km) using several approaches. Plants originating from four distinct habitats at two geographic scales (cold edge, arid edge and two topographically dissimilar core locations) were reciprocally transplanted and their success over time was measured as 1) lifetime fitness in a year of planting, and 2) population growth four years after planting. In addition, we analyzed molecular (SSR) and quantitative trait variation and calculated the QST/FST ratio. No home advantage was detected at the small spatial scale. At the large spatial scale, home advantage was detected for the core population and the cold edge population in the year of introduction via measuring life-time plant performance. However, superior performance of the arid edge population in its own environment was evident only after several generations via measuring experimental population growth rate through genotyping with SSRs allowing counting the number of plants and seeds per introduced genotype per site. These results highlight the importance of multi-generation surveys of population growth rate in local adaptation testing. Despite predominant self-fertilization of T. dicoccoides and the associated high degree of structuring of genetic variation, the results of the QST - FST comparison were in general agreement with the pattern of local adaptation at the two spatial scales detected by reciprocal transplanting.