Genetic dissection of shoot traits and proline content under control and drought conditions in barley

Drought is the most severe threat to world crop cultivation and production especially in water shortage areas. Wild barley diversity contains notable variation in phenotype that is essential for its adaptation to abiotic stress like drought. In the current study, we performed QTL mapping for shoot traits and proline content accumulation under control and drought conditions. A library of 73 (BC3S4:S10) S42ILs derived from German cultivar Scarlett and wild accession from Israel (ISR42-8) was used in this experimental study and genotyped for shoot traits with a 1,536-SNP Illumina BOPA1 set. Plants were analyzed and phenotypic data was collected for eight shoot traits and physiological trait i.e; proline content. All studied traits showed high significant differences between both treatments. Genetic mapping reveals total twenty QTLs for shoot traits and five QTLs for drought inducible proline accumulation all over the barley genome and had main effects on improving or reducing the traits under control and drought stress conditions. The most important QTL which have been obtained in the current study is for proline content on 1H chromosome. Further mapping and validation in a high resolution population revealed that Qpro.S42-1H underlie a previously unknown HvP5CS1 allele originated from wild barley. The functional mutations were found in the promoter motifs for DNA binding transcription factor i.e; ABRE-binding factors (ABF1, ABF2), where the number and arrangements of ABFs binding motifs in the wild P5CS1 allele in ISR42-8 appeared to imply transcriptional up regulation and excessive proline accumulation under extreme drought conditions. Higher proline accumulation in QTL allele bearing ILs S42IL-143 and S42IL-141 conferred improved physiological activity and photosynthetic yield, thus confirming functionality of an exotic P5CS1 allele in the cultivated barley. The present findings brought up a first insight on the molecular and evolutionary regulation of an essential drought physiological traits in crop plant. These resources offer opportunity to understand adaptive biology of crop plants and can serve as direct target for trait improvement in barley and related species.Die Kultivierung und der Ertrag von Feldfrüchten wird hauptsächlich durch Trockenheit insbesondere in Anbaugebieten mit zunehmender Wasserverknappung gefährdet. Die Wildformen unserer Kulturgerste bieten ein hohes Maß an Variation des Phänotyps und somit an Anpassungsmöglichkeiten an diverse abiotische Stressszenarien wie etwa Trockenheit. Für die vorliegende Arbeit wurde eine Kartierung quantitativer Merkmale für Sprossparameter, sowie des Gehalts von Prolin, einem Pflanzenhormon, unter Kontroll- und Stressbedingungen durchgeführt. Eine Population bestehend aus 73 (BC3S4:S10) S42-Introgressionslinien abstammend von der deutschen Kultursorte Scarlett und der exotischen Linie ISR42-8 aus Israel wurde im Hinblick auf Sprossmerkmale mithilfe eines Illumina BOPA1 Sets anhand von 1536 SNPs genotypisiert. Die Versuchspflanzen wurden im Hinblick auf acht phänotypische Sprossmerkmale, sowie physiologische Merkmale, wie z.B. Prolingehalt analysiert. Alle untersuchten Parameter wiesen hochsignifikante Unterschiede zwischen den beiden Behandlungen auf. Die genetische Kartierung ergab insgesamt 20 QTLs für Sprossmerkmale und fünf QTLs für trockeninduzierte Prolinanreicherung, verteilt auf das gesamte Gerstengenom und hatte wichtige Effekte in Bezug auf die Ausprägung der entsprechenden Merkmale unter Kontroll-bzw. Stressbedingungen. Für Prolingehalt konnte in der aktuellen Studie ein wichtiges QTL auf Chromosom 1H lokalisiert werden. Die weitere Kartierung und Validierung in einer höher auflösenden Population ergab, dass der Genort Qpro.S42-1H einem bislang unbekannten HvP5CS1 Allel aus Wildgerste entstammt. Funktionelle Mutationen wurden in der Promotorregion für DNA-bindende Transkriptionsfaktoren wie z.B. ABRE-Bindungsfaktoren (ABF1, ABF2) entdeckt, wobei deren Anzahl und Anordnung im exotischen P5CS1 Allel in ISR42-8 eine deutliche Prolinanreicherung unter extremen Trockenstressbedingungen reguliert. Die Introgressionslinien S42IL-143 und S42IL-14, welche das entsprechende QTL-Allel für eine Prolinanhäufung tragen, zeigten eine verbesserte physiologische Aktivität und Photosyntheserate und bestätigten damit die Funktionalität des P5CS1-Allels in der Kulturgerste. Die vorliegenden Ergebnisse geben einen ersten Einblick in die Regulierung eines entscheidenden physiologischen Merkmals auf molekularer Ebene in Pflanzen. Dadurch werden Möglichkeiten zum Verständnis der Anpassung von Pflanzen an Trockenstress und die Nutzung dieser wertvollen Ressourcen als Quelle zur Leistungsverbesserung von Gerste und verwandten Spezies eröffnet

Correction: Genetic Mapping Reveals Broader Role of Vrn-H3 Gene in Root and Shoot Development beyond Heading in Barley.

[This corrects the article DOI: 10.1371/journal.pone.0158718.]

Genetic Mapping Reveals Broader Role of Vrn-H3 Gene in Root and Shoot Development beyond Heading in Barley.

The aim of the present study was to dissect the genetic inheritance and interplay of root, shoot and heading attributes for a better understanding of these traits in crop production. For this, we utilized quantitative trait loci (QTL) and candidate gene analysis approach using a second filial (F2) population originated from a cross between spring cultivar Cheri and wild barley accession ICB181160. The F2 population comprising 182 plants was phenotyped for root dry weight (RDW), root volume (RV), root length (RL) and shoot dry weight (SDW), tiller number per plant (TIL) and days to heading (HEA). In parallel, this population was genotyped using polymerase chain reaction (PCR) based cleaved amplified polymorphic sequence (CAPS) markers distributed across the whole genome. Marker by trait analysis revealed 16 QTL for root and shoot traits localized on chromosomes 1H, 3H, 4H, 5H and 7H. The strongest and a common QTL effect for root, shoot and heading traits was identified on chromosome 7H at the putative region of Vrn-H3 gene. Later, we have established PCR based gene specific marker HvVrnH3 revealing polymorphism for early heading Vrn-H3 allele in Cheri and late heading allele vrn-H3 in ICB181160. Genotyping of these alleles revealed a clear co-segregation of early heading Vrn-H3 allele with lower root and shoot attributes, while late heading vrn-H3 allele with more TIL and higher root biomass suggesting a primary insight on the function of Vrn-H3 gene beyond flowering. Genetic interactions of vernalization genes Vrn-H3 with Vrn-H2 and Vrn-H1 also suggested the major role of Vrn-H3 alleles in determining root and shoot trait variations in barley. We believe, these data provide an opportunity for further research to test a precise significance of early heading on yield components and root associated sustainability in crops like barley and wheat

Evolution of Deeper Rooting 1-like homoeologs in wheat entails the C-terminus mutations as well as gain and loss of auxin response elements.

Root growth angle (RGA) in response to gravity controlled by auxin is a pertinent target trait for obtainment of higher yield in cereals. But molecular basis of this root architecture trait remain obscure in wheat and barley. We selected four cultivars two each for wheat and barley to unveil the molecular genetic mechanism of Deeper Rooting 1-like gene which controls RGA in rice leading to higher yield under drought imposition. Morphological analyses revealed a deeper and vertically oriented root growth in "NARC 2009" variety of wheat than "Galaxy" and two other barley cultivars "Scarlet" and "ISR42-8". Three new homoeologs designated as TaANDRO1-like, TaBNDRO1-like and TaDNDRO1-like corresponding to A, B and D genomes of wheat could be isolated from "NARC 2009". Due to frameshift and intronization/exonization events the gene structures of these paralogs exhibit variations in size. DRO1-like genes with five distinct domains prevail in diverse plant phyla from mosses to angiosperms but in lower plants their differentiation from LAZY, NGR and TAC1 (root and shoot angle genes) is enigmatic. Instead of IGT as denominator motif of this family, a new C-terminus motif WxxTD in the V-domain is proposed as family specific motif. The EAR-like motif IVLEM at the C-terminus of the TaADRO1-like and TaDDRO1-like that diverged to KLHTLIPNK in TaBDRO1-like and HvDRO1-like is the hallmark of these proteins. Split-YFP and yeast two hybrid assays complemented the interaction of TaDRO1-like with TOPLESS-a repressor of auxin regulated root promoting genes in plants-through IVLEM/KLHTLIPNK motif. Quantitative RT-PCR revealed abundance of DRO1-like RNA in root tips and spikelets while transcript signals were barely detectable in shoot and leaf tissues. Interestingly, wheat exhibited stronger expression of TaBDRO1-like than barley (HvDRO1-like), but TaBDRO1-like was the least expressing among three paralogs. The underlying cause of this expression divergence seems to be the presence of AuxRE motif TGTCTC and core TGTC with a coupling AuxRE-like motif ATTTTCTT proximal to the transcriptional start site in TaBDRO1-like and HvDRO1-like promoters. This is evident from binding of ARF1 to TGTCTC and TGTC motifs of TaBDRO1-like as revealed by yeast one-hybrid assay. Thus, evolution of DRO1-like wheat homoeologs might incorporate the C-terminus mutations as well as gain and loss of AuxREs and other cis-regulatory elements during expression divergence. Since root architecture is an important target trait for wheat crop improvement, therefore DRO1-like genes have potential applications in plant breeding for enhancement of plant productivity by the use of modern genome editing approaches

QTL map of root, shoot and heading traits detected in the F2 population of a cross between spring barley cultivar Cheri and wild accession ICB181160.

<p>QTL map of root, shoot and heading traits detected in the F2 population of a cross between spring barley cultivar Cheri and wild accession ICB181160.</p

Genetic interaction plot of <i>Vrn-H3</i> and <i>Vrn-H1</i> alleles from Cheri and ICB181160 among the F2 population for heading.

<p>All nine allele combinations of both genes are calculated using R-interaction plots.</p

Barley haplotype scoring from six SNPs and one indel in the intron 1 and exon 2 of <i>Vrn-H3</i> gene.

<p>Barley haplotype scoring from six SNPs and one indel in the intron 1 and exon 2 of <i>Vrn-H3</i> gene.</p

Mean comparisons of the root and shoot traits among Cheri, ICB181160 and the F2 population.

<p>Mean comparisons of the root and shoot traits among Cheri, ICB181160 and the F2 population.</p

Correlation coefficients among the root and shoot traits in the F2 population.

<p>Correlation coefficients among the root and shoot traits in the F2 population.</p

Genotypic and phenotypic trait variations in the F2 population for the gene-specific marker HvVrnH3.

<p>A) Genotypic scores of marker HvVrnH3 showing polymorphic <i>Vrn-H3</i> and <i>vrn-H3</i> alleles in barley. B-F) Phenotypic quantification of QTL showing the common effect of <i>Vrn-H3</i> gene on root, shoot and heading traits. Cultivar Cheri allele (AA), heterozygous (Aa) and ICB181160 allele (aa).</p