SUMO protease FUG1, histone reader AL3 and chromodomain protein LHP1 are integral to repeat expansion-induced gene silencing in Arabidopsis thaliana.

Epigenetic gene silencing induced by expanded repeats can cause diverse phenotypes ranging from severe growth defects in plants to genetic diseases such as Friedreich's ataxia in humans. The molecular mechanisms underlying repeat expansion-induced epigenetic silencing remain largely unknown. Using a plant model with a temperature-sensitive phenotype, we have previously shown that expanded repeats can induce small RNAs, which in turn can lead to epigenetic silencing through the RNA-dependent DNA methylation pathway. Here, using a genetic suppressor screen and yeast two-hybrid assays, we identified novel components required for epigenetic silencing caused by expanded repeats. We show that FOURTH ULP GENE CLASS 1 (FUG1)-an uncharacterized SUMO protease with no known role in gene silencing-is required for epigenetic silencing caused by expanded repeats. In addition, we demonstrate that FUG1 physically interacts with ALFIN-LIKE 3 (AL3)-a histone reader that is known to bind to active histone mark H3K4me2/3. Loss of function of AL3 abolishes epigenetic silencing caused by expanded repeats. AL3 physically interacts with the chromodomain protein LIKE HETEROCHROMATIN 1 (LHP1)-known to be associated with the spread of the repressive histone mark H3K27me3 to cause repeat expansion-induced epigenetic silencing. Loss of any of these components suppresses repeat expansion-associated phenotypes coupled with an increase in IIL1 expression with the reversal of gene silencing and associated change in epigenetic marks. Our findings suggest that the FUG1-AL3-LHP1 module is essential to confer repeat expansion-associated epigenetic silencing and highlight the importance of post-translational modifiers and histone readers in epigenetic silencing.

Quantitative Trait Loci for Genotype and Genotype by Environment Interaction Effects for Seed Yield Plasticity to Terminal Water-Deficit Conditions in Canola (Brassica napus L.).

Canola plants suffer severe crop yield and oil content reductions when exposed to water-deficit conditions, especially during the reproductive stages of plant development. There is a pressing need to develop canola cultivars that can perform better under increased water-deficit conditions with changing weather patterns. In this study, we analysed genetic determinants for the main effects of quantitative trait loci (QTL), (Q), and the interaction effects of QTL and Environment (QE) underlying seed yield and related traits utilising 223 doubled haploid (DH) lines of canola in well-watered and water-deficit conditions under a rainout shelter. Moderate water-deficit at the pre-flowering stage reduced the seed yield to 40.8%. Multi-environmental QTL analysis revealed 23 genomic regions associated with days to flower (DTF), plant height (PH) and seed yield (SY) under well-watered and water-deficit conditions. Three seed yield QTL for main effects were identified on chromosomes A09, C03, and C09, while two were related to QE interactions on A02 and C09. Two QTL regions were co-localised to similar genomic regions for flowering time and seed yield (A09) and the second for plant height and chlorophyll content. The A09 QTL was co-located with a previously mapped QTL for carbon isotope discrimination (Δ13C) that showed a positive relationship with seed yield in the same population. Opposite allelic effects for plasticity in seed yield were identified due to QE interactions in response to water stress on chromosomes A02 and C09. Our results showed that QTL's allelic effects for DTF, PH, and SY and their correlation with Δ13C are stable across environments (field conditions, previous study) and contrasting water regimes (this study). The QTL and DH lines that showed high yield under well-watered and water-deficit conditions could be used to manipulate water-use efficiency for breeding improved canola cultivars.

Quantifying splice-site usage: a simple yet powerful approach to analyze splicing.

RNA splicing, and variations in this process referred to as alternative splicing, are critical aspects of gene regulation in eukaryotes. From environmental responses in plants to being a primary link between genetic variation and disease in humans, splicing differences confer extensive phenotypic changes across diverse organisms (1-3). Regulation of splicing occurs through differential selection of splice sites in a splicing reaction, which results in variation in the abundance of isoforms and/or splicing events. However, genomic determinants that influence splice-site selection remain largely unknown. While traditional approaches for analyzing splicing rely on quantifying variant transcripts (i.e. isoforms) or splicing events (i.e. intron retention, exon skipping etc.) (4), recent approaches focus on analyzing complex/mutually exclusive splicing patterns (5-8). However, none of these approaches explicitly measure individual splice-site usage, which can provide valuable information about splice-site choice and its regulation. Here, we present a simple approach to quantify the empirical usage of individual splice sites reflecting their strength, which determines their selection in a splicing reaction. Splice-site strength/usage, as a quantitative phenotype, allows us to directly link genetic variation with usage of individual splice-sites. We demonstrate the power of this approach in defining the genomic determinants of splice-site choice through GWAS. Our pilot analysis with more than a thousand splice sites hints that sequence divergence in cis rather than trans is associated with variations in splicing among accessions of Arabidopsis thaliana. This approach allows deciphering principles of splicing and has broad implications from agriculture to medicine.

Genetics of Na+ exclusion and salinity tolerance in Afghani durum wheat landraces.

BACKGROUND: Selecting for low concentration of Na+ in the shoot provides one approach for tackling salinity stress that adversely affects crop production. Novel alleles for Na+ exclusion can be identified and then introduced into elite crop cultivars. RESULTS: We have identified loci associated with lower Na+ concentration in leaves of durum wheat landraces originating from Afghanistan. Seedlings of two F2 populations derived from crossings between Australian durum wheat (Jandaroi) and two Afghani landraces (AUS-14740 and AUS-14752) were grown hydroponically and evaluated for Na+ and K+ concentration in the third leaf. High heritability was found for both third leaf Na+ concentration and the K+/Na+ ratio in both populations. Further work focussed on line AUS-14740. Bulk segregant analysis using 9 K SNP markers identified two loci significantly associated with third leaf Na+ concentration. Marker regression analysis showed a strong association between all traits studied and a favourable allele originating from AUS-14740 located on the long arm of chromosome 4B. CONCLUSIONS: The candidate gene in the relevant region of chromosome 4B is likely to be the high affinity K+ transporter B1 (HKT1;5-B1). A second locus associated with third leaf Na+ concentration was located on chromosome 3BL, with the favourable allele originating from Jandaroi; however, no candidate gene can be identified.