Genomic insights into HSFs as candidate genes for high-temperature stress adaptation and gene editing with minimal off-target effects in flax

Abstract

Flax ( Linum usitatissimum ) is a cool season crop commercially cultivated for seed oil and stem fibre production. A comprehensive characterization of the heat shock factor ( HSF ) candidate genes in flax can accelerate genetic improvement and adaptive breeding for high temperature stress tolerance. We report the genome-wide identification of 34 putative HSF genes from the flax genome, which we mapped on 14 of the 15 chromosomes. Through comparative homology analysis, we classified these genes into three broad groups, and sub-groups. The arrangement of HSF-specific protein motifs, DNA-binding domain (DBD) and hydrophobic heptad repeat (HR-A/B), and exon-intron boundaries substantiated the phylogenetic separation of these genes. Orthologous relationships and evolutionary analysis revealed that the co-evolution of the LusHSF genes was due to recent genome duplication events. Digital and RT-qPCR analyses provided significant evidence of the differential expression of the LusHSF genes in various tissues, at various developmental stages, and in response to high-temperature stress. The co-localization of diverse cis-acting elements in the promoters of the LusHSF genes further emphasized their regulatory roles in the abiotic stress response. We further confirmed DNA-binding sites on the LusHSF proteins and designed guide RNA sequences for gene editing with minimal off-target effects. These results will hasten functional investigations of LusHSFs or assist in devising genome engineering strategies to develop high-temperature stress tolerant flax cultivars.

Genome-wide identification, classification and expression profile analysis of the HSF gene family in Hypericum perforatum

“…Previous studies have shown that transcription activation in vivo requires HSEs in HSF protein binding. The HSF recognizes a typical 5 bp sequence, 5-nGAAn-3, which forms at least three contiguous inverted repeats in the downstream HSP promoter ( Saha et al, 2019 ). The highly conserved DNA-binding domain (DBD) in the N-terminus includes one three-helical bundle (α1, α2, α3) and one antiparallel four-stranded β -sheet ( β 1, β 2, β 3, β 4) to form a helix-turn-helix structure.…”

“…Recently, HSF gene families were analyzed in different species, including maize, rice, pepper, tomato, soybean, and flax. Genome-wide analysis indicated that HSF proteins in various species may have different functions in reducing damage to high-temperature stress and also provide resources for evolutionary analysis ( Guo et al, 2015 ; Saha et al, 2019 ; Yang et al, 2016 ).…”

Adaptation Strategies to Improve the Resistance of Oilseed Crops to Heat Stress Under a Changing Climate: An Overview

“…The promoter regions of the soybean HSFs contained cis-elements that likely participate in drought, low temperature, and ABA stress responses. GmHsp90A2, GmRAR1, GmSGT1, GmSBH1P, and GmSBH1 are essential chaperones of the protective stress response in soybean (Chen et al, 2019 ; Huang et al, 2019b ), while LusHSF responds in linseed (Saha et al, 2019 ). These regulators play their role in making the interaction between the MBF1c ethylene activated pathway and HSP allied signaling.…”

Genome Sequencing of Fiber Flax Cultivar Atlant Using Oxford Nanopore and Illumina Platforms

“…Scaffold-level genome assemblies of linseed cultivar Longya-10, fiber cultivar Heiya-14, and pale flax were generated in 2020, based on Illumina sequencing, Hi-C technology, and genetic mapping (Zhang et al, 2020b ). These results are extremely important for further progress in molecular studies of flax, the development of genome editing, and marker-assisted and genomic selection (Saha et al, 2019 ; Morello et al, 2020 ; You and Cloutier, 2020 ). A high-quality genome can be used as a reference for genome and transcriptome assemblies of different flax cultivars/lines, and the identification of polymorphisms and differences in gene expression within L. usitatissimum genotypes (Dmitriev et al, 2019 , 2020b ; Guo et al, 2019 ; Wu et al, 2019 ).…”