Histone Deacetylase (HDAC) Gene Family in Allotetraploid Cotton and Its Diploid Progenitors: In Silico Identification, Molecular Characterization, and Gene Expression Analysis under Multiple Abiotic Stresses, DNA Damage and Phytohormone Treatments.

Histone deacetylases (HDACs) play a significant role in a plant's development and response to various environmental stimuli by regulating the gene transcription. However, HDACs remain unidentified in cotton. In this study, a total of 29 HDACs were identified in allotetraploid Gossypium hirsutum, while 15 and 13 HDACs were identified in Gossypium arboretum and Gossypium raimondii, respectively. Gossypium HDACs were classified into three groups (reduced potassium dependency 3 (RPD3)/HDA1, HD2-like, and Sir2-like (SRT) based on their sequences, and Gossypium HDACs within each subgroup shared a similar gene structure, conserved catalytic domains and motifs. Further analysis revealed that Gossypium HDACs were under a strong purifying selection and were unevenly distributed on their chromosomes. Gene expression data revealed that G. hirsutum HDACs were differentially expressed in various vegetative and reproductive tissues, as well as at different developmental stages of cotton fiber. Furthermore, some G. hirsutum HDACs were co-localized with quantitative trait loci (QTLs) and single-nucleotide polymorphism (SNPs) of fiber-related traits, indicating their function in fiber-related traits. We also showed that G. hirsutum HDACs were differentially regulated in response to plant hormones (abscisic acid (ABA) and auxin), DNA damage agent (methyl methanesulfonate (MMS)), and abiotic stresses (cold, salt, heavy metals and drought), indicating the functional diversity and specification of HDACs in response to developmental and environmental cues. In brief, our results provide fundamental information regarding G. hirsutum HDACs and highlight their potential functions in cotton growth, fiber development and stress adaptations, which will be helpful for devising innovative strategies for the improvement of cotton fiber and stress tolerance.

Comparative Genome-wide Analysis and Expression Profiling of Histone Acetyltransferase (HAT) Gene Family in Response to Hormonal Applications, Metal and Abiotic Stresses in Cotton.

Post-translational modifications are involved in regulating diverse developmental processes. Histone acetyltransferases (HATs) play vital roles in the regulation of chromation structure and activate the gene transcription implicated in various cellular processes. However, HATs in cotton, as well as their regulation in response to developmental and environmental cues, remain unidentified. In this study, 9 HATs were identified from Gossypium raimondi and Gossypium arboretum, while 18 HATs were identified from Gossypium hirsutum. Based on their amino acid sequences, Gossypium HATs were divided into three groups: CPB, GNAT, and TAFII250. Almost all the HATs within each subgroup share similar gene structure and conserved motifs. Gossypium HATs are unevenly distributed on the chromosomes, and duplication analysis suggests that Gossypium HATs are under strong purifying selection. Gene expression analysis showed that Gossypium HATs were differentially expressed in various vegetative tissues and at different stages of fiber development. Furthermore, all the HATs were differentially regulated in response to various stresses (salt, drought, cold, heavy metal and DNA damage) and hormones (abscisic acid (ABA) and auxin (NAA)). Finally, co-localization of HAT genes with reported quantitative trait loci (QTL) of fiber development were reported. Altogether, these results highlight the functional diversification of HATs in cotton growth and fiber development, as well as in response to different environmental cues. This study enhances our understanding of function of histone acetylation in cotton growth, fiber development, and stress adaptation, which will eventually lead to the long-term improvement of stress tolerance and fiber quality in cotton.

Roegneria alashanica Keng: a species with the StStStYStY genome constitution.

The genome constitution of tetraploid Roegneria alashanica Keng has been in question for a long time. Most scientific studies have suggested that R. alashanica had two versions of the St genome, St1St2, similar to that of Pseudoroegneria elytrigioides (C. Yen & J.L. Yang) B.R. Lu. A study, however, concluded that R. alashanica had the StY genome formula typical for tetraploid species of Roegneria. For the present study, R. alashanica, Elymus longearistatus (Bioss.) Tzvelev (StY genomes), Pseudoroegneria strigosa (M. Bieb.) Á. Löve (St), Pseudoroegneria libanoctica (Hackel) D.R. Dewey (St), and Pseudoroegneria spicata (Pursh) Á. Löve (St) were screened for the Y-genome specific marker B14(F+R)269. All E. longearistatus plants expressed intense bands specific to the Y genome. Only 6 of 10 R. alashanica plants exhibited relatively faint bands for the STS marker. Previously, the genome in species of Pseudoroegneria exhibiting such faint Y-genome specific marker was designated as StY. Based on these results, R. alashanica lacks the Y genome in E. longearistatus but likely possess two remotely related St genomes, St and StY. According to its genome constitution, R. alashanica should be classified in the genus Pseudoroenera and given the new name Pseudoroegneria alashanica (Keng) R.R.-C. Wang and K.B. Jensen.

Differential transferability of EST-SSR primers developed from the diploid species Pseudoroegneria spicata, Thinopyrum bessarabicum, and Thinopyrum elongatum.

Simple sequence repeat technology based on expressed sequence tag (EST-SSR) is a useful genomic tool for genome mapping, characterizing plant species relationships, elucidating genome evolution, and tracing genes on alien chromosome segments. EST-SSR primers developed from three perennial diploid species of Triticeae, Pseudoroegneria spicata (Pursh) Á. Löve (having St genome), Thinopyrum bessarabicum (Savul. & Rayss) Á. Löve (Jb = Eb = J), and Thinopyrum elongatum (Host) D.R. Dewey (Je = Ee = E), were used to produce amplicons in these three species to (i) assess relative transferability, (ii) identify polymorphic species-specific markers, and (iii) determine genome relationships among the three species. Because of the close relationship between Jb and Je genomes, EST-SSR primers derived from Th. bessarabicum and Th. elongatum had greater transferability to each other than those derived from the St-genome P. spicata. A large number of polymorphic species- and genome-specific EST-SSR amplicons were identified that will be used for construction of genetic maps of these diploid species, and tracing economically useful genes in breeding or gene transfer programs in various species of Triticeae.

Genome evolution of intermediate wheatgrass as revealed by EST-SSR markers developed from its three progenitor diploid species.

Intermediate wheatgrass (Thinopyrum intermedium (Host) Barkworth & D.R. Dewey), a segmental autoallohexaploid (2n = 6x = 42), is not only an important forage crop but also a valuable gene reservoir for wheat (Triticum aestivum L.) improvement. Throughout the scientific literature, there continues to be disagreement as to the origin of the different genomes in intermediate wheatgrass. Genotypic data obtained from newly developed EST-SSR primers derived from the putative progenitor diploid species Pseudoroegneria spicata (Pursh) Á. Löve (St genome), Thinopyrum bessarabicum (Savul. & Rayss) Á. Löve (J = J(b) = E(b)), and Thinopyrum elongatum (Host) D. Dewey (E = J(e) = E(e)) indicate that the V genome of Dasypyrum (Coss. & Durieu) T. Durand is not one of the three genomes in intermediate wheatgrass. Based on all available information in the literature and findings in this study, the genomic designation of intermediate wheatgrass should be changed to J(vs)J(r)St, where J(vs) and J(r) represent ancestral genomes of present-day J(b) of Th. bessarabicum and J(e) of Th. elongatum, with J(vs) being more ancient. Furthermore, the information suggests that the St genome in intermediate wheatgrass is most similar to the present-day St found in diploid species of Pseudoroegneria from Eurasia.

Genetic control of rhizomes and genomic localization of a major-effect growth habit QTL in perennial wildrye.

Rhizomes are prostrate subterranean stems that provide primitive mechanisms of vegetative dispersal, survival, and regrowth of perennial grasses and other monocots. The extent of rhizome proliferation varies greatly among grasses, being absent in cereals and other annuals, strictly confined in caespitose perennials, or highly invasive in some perennial weeds. However, genetic studies of rhizome proliferation are limited and genes controlling rhizomatous growth habit have not been elucidated. Quantitative trait loci (QTLs) controlling rhizome spreading were compared in reciprocal backcross populations derived from hybrids of rhizomatous creeping wildrye (Leymus triticoides) and caespitose basin wildrye (L. cinereus), which are perennial relatives of wheat. Two recessive QTLs were unique to the creeping wildrye backcross, one dominant QTL was unique to the basin wildrye backcross, and one additive QTL was detectable in reciprocal backcrosses with high log odds (LOD = 31.6) in the basin wildrye background. The dominant QTL located on linkage group (LG)-2a was aligned to a dominant rhizome orthogene (Rhz3) of perennial rice (Oryza longistamina) and perennial sorghum (Sorghum propinquum). Nonparametric 99 % confidence bounds of the 31.6-LOD QTL were localized to a distal 3.8-centiMorgan region of LG-6a, which corresponds to a 0.7-Mb region of Brachypodium Chromosome 3 containing 106 genes. An Aux/IAA auxin signal factor gene was located at the 31.6-LOD peak, which could explain the gravitropic and aphototropic behavior of rhizomes. Findings elucidate genetic mechanisms controlling rhizome development and architectural growth habit differences among plant species. Results have possible applications to improve perennial forage and turf grasses, extend the vegetative life cycle of annual cereals, such as wheat, or control the invasiveness of highly rhizomatous weeds such as quackgrass (Elymus repens).

Genes and QTLs controlling inflorescence and stem branch architecture in Leymus (Poaceae: Triticeae) Wildrye.

Grass inflorescence and stem branches show recognizable architectural differences among species. The inflorescence branches of Triticeae cereals and grasses, including wheat, barley, and 400-500 wild species, are usually contracted into a spike formation, with the number of flowering branches (spikelets) per node conserved within species and genera. Perennial Triticeae grasses of genus Leymus are unusual in that the number of spikelets per node varies, inflorescences may have panicle branches, and vegetative stems may form subterranean rhizomes. Leymus cinereus and L. triticoides show discrete differences in inflorescence length, branching architecture, node number, and density; number of spikelets per node and florets per spikelet; culm length and width; and perimeter of rhizomatous spreading. Quantitative trait loci controlling these traits were detected in 2 pseudo-backcross populations derived from the interspecific hybrids using a linkage map with 360 expressed gene sequence markers from Leymus tiller and rhizome branch meristems. Alignments of genes, mutations, and quantitative trait loci controlling similar traits in other grass species were identified using the Brachypodium genome reference sequence. Evidence suggests that loci controlling inflorescence and stem branch architecture in Leymus are conserved among the grasses, are governed by natural selection, and can serve as possible gene targets for improving seed, forage, and grain production.

Leymus EST linkage maps identify 4NsL-5NsL reciprocal translocation, wheat-Leymus chromosome introgressions, and functionally important gene loci.

Allotetraploid (2n = 4x = 28) Leymus triticoides and Leymus cinereus are divergent perennial grasses, which form fertile hybrids. Genetic maps with n = 14 linkage groups (LG) comprised with 1,583 AFLP and 67 heterologous anchor markers were previously used for mapping quantitative trait loci (QTLs) in these hybrids, and chromosomes of other Leymus wildryes have been transferred to wheat. However, identifications of the x = 7 homoeologous groups were tenuous and genetic research has been encumbered by a lack of functional, conserved gene marker sequences. Herein, we mapped 350 simple sequence repeats and 26 putative lignin biosynthesis genes from a new Leymus EST library and constructed one integrated consensus map with 799 markers, including 375 AFLPs and 48 heterologous markers, spanning 2,381 centiMorgans. LG1b and LG6b were reassigned as LG6b* and LG1b*, respectively, and LG4Ns and LG4Xm were inverted so that all 14 linkage groups are aligned to the x = 7 Triticeae chromosomes based on EST alignments to barley and other reference genomes. Amplification of 146 mapped Leymus ESTs representing six of the seven homoeologous groups was shown for 17 wheat-Leymus chromosome introgression lines. Reciprocal translocations between 4L and 5L in both Leymus and Triticum monococcum were aligned to the same regions of Brachypodium chromosome 1. A caffeic acid O-methyltransferase locus aligned to fiber QTL peaks on Leymus LG7a and brown midrib mutations of maize and sorghum. Glaucousness genes on Leymus and wheat chromosome 2 were aligned to the same region of Brachypodium chromosome 5. Markers linked to the S self-incompatibility gene on Leymus LG1a cosegregated with markers on LG2b, possibly cross-linked by gametophytic selection. Homoeologous chromosomes 1 and 2 harbor the S and Z gametophytic self-incompatibility genes of Phalaris, Secale, and Lolium, but the Leymus chromosome-2 self-incompatibility gene aligns to a different region on Brachypodium chromosome 5. Nevertheless, cosegregation of self-incompatibility genes on Leymus presents a powerful system for mapping these loci.

A molecular genetic linkage map identifying the St and H subgenomes of Elymus (Poaceae: Triticeae) wheatgrass.

Elymus L. is the largest and most complex genus in the Triticeae tribe of grasses with approximately 150 polyploid perennial species occurring worldwide. We report here the first genetic linkage map for Elymus. Backcross mapping populations were created by crossing caespitose Elymus wawawaiensis (EW) (Snake River wheatgrass) and rhizomatous Elymus lanceolatus (EL) (thickspike wheatgrass) to produce F(1) interspecific hybrids that were then backcrossed to the same EL male to generate progeny with segregating phenotypes. EW and EL are both allotetraploid species (n = 14) containing the St (Pseudoroegneria) and H (Hordeum) genomes. A total of 387 backcross progeny from four populations were genotyped using 399 AFLP and 116 EST-based SSR and STS markers. The resulting consensus map was 2574 cM in length apportioned among the expected number of 14 linkage groups. EST-based SSR and STS markers with homology to rice genome sequences were used to identify Elymus linkage groups homoeologous to chromosomes 1-7 of wheat. The frequency of St-derived genome markers on each linkage group was used to assign genome designations to all linkage groups, resulting in the identification of the seven St and seven H linkage groups of Elymus. This map also confirms the alloploidy and disomic chromosome pairing and segregation of Elymus and will be useful in identifying QTLs controlling perennial grass traits in this genus.

Orchardgrass (Dactylis glomerata L.) EST and SSR marker development, annotation, and transferability.

Orchardgrass, or cocksfoot [Dactylis glomerata (L.)], has been naturalized on nearly every continent and is a commonly used species for forage and hay production. All major cultivated varieties of orchardgrass are autotetraploid, and few tools or information are available for functional and comparative genetic analyses and improvement of the species. To improve the genetic resources for orchardgrass, we have developed an EST library and SSR markers from salt, drought, and cold stressed tissues. The ESTs were bi-directionally sequenced from clones and combined into 17,373 unigenes. Unigenes were annotated based on putative orthology to genes from rice, Triticeae grasses, other Poaceae, Arabidopsis, and the non-redundant database of the NCBI. Of 1,162 SSR markers developed, approximately 80% showed amplification products across a set of orchardgrass germplasm, and 40% across related Festuca and Lolium species. When orchardgrass subspecies were genotyped using 33 SSR markers their within-accession similarity values ranged from 0.44 to 0.71, with Mediterranean accessions having a higher similarity. The total number of genotyped bands was greater for tetraploid accessions compared to diploid accessions. Clustering analysis indicated grouping of Mediterranean subspecies and central Asian subspecies, while the D. glomerata ssp. aschersoniana was closest related to three cultivated varieties.

Analyses of Thinopyrum bessarabicum, T. elongatum, and T. junceum chromosomes using EST-SSR markers.

Wild Thinopyrum grasses are important gene pools for forage and cereal crops. Knowledge of their chromosome organizations is pivotal for efficient utilization of this important gene pool in germplasm enhancement programs. Expressed sequence tags derived simple sequence repeat (EST-SSR) markers for Thinopyrum bessarabicum, T. elongatum, and T. junceum chromosomes were identified among amplicons produced from three series of wheat-Thinopyrum addition lines using 193 primer pairs designed from the Leymus EST unigenes. The homology of T. junceum chromosomes in 13 wheat addition lines was tentatively established to reveal that homologous groups 3, 4, 5, 6, and 7 were represented by HD3515, HD3505, AJDAj11, AJDAj1, and HD3508, whereas groups 1 and 2 were represented by AJADj7-AJDAj9 and AJDAj2-AJDAj4, respectively. AJDAj5 and AJDAj6 had complexly reconstituted T. junceum chromosomes that might have resulted from fusion or translocations of large chromosomal segments from two or more chromosomes, that is (1+5) and (2+5+1), respectively. The identified EST-SSR markers will be useful in comparative gene mapping, chromosome tracing, taxonomic studies, gene introgression, and cultivar identification.

Population structure in Pseudoroegneria spicata (Poaceae: Triticeae) modeled by Bayesian clustering of AFLP genotypes.

Pseudoroegneria spicata (Poaceae: Triticeae) is an abundant, allogamous species widely adapted to the temperate, semiarid steppe and open woodland regions of western North America. Amplified fragment length polymorphism (AFLP), model-based Bayesian clustering, and other methods of hypothesis testing were used to investigate genetic diversity and population structure among 565 P. spicata plants from 82 localities representing much of the species distribution. Comparisons with four Asiatic Pseudoroegneria species and two North American Elymus wawawaiensis accessions demonstrate cohesiveness in P. spicata. However, P. spicata genotypes group by locality and geographic region based on genetic distance analysis. Average DNA polymorphism among P. spicata localities was significantly correlated (r = 0.58) with geographical distance. The optimum Bayesian cluster model included 21 P. spicata groups, indicating that dispersal among sampling locations was not sufficient to group genotypes into one unstructured population. Approximately 18.3% of the DNA polymorphism was partitioned among the 21 regional groups, 14.9% among localities within groups, and 66.8% within accessions. Average DNA polymorphism among Bayesian groups was correlated (r = 0.53) with the average geographic distance among Bayesian groups, which partly reflects isolation by distance. However, conspicuous regional boundaries were discernable among several divergent genetic groups.

Molecular genetic linkage maps for allotetraploid Leymus wildryes (Gramineae: Triticeae).

Molecular genetic maps were constructed for two full-sib populations, TTC1 and TTC2, derived from two Leymus triticoides x Leymus cinereus hybrids and one common Leymus triticoides tester. Informative DNA markers were detected using 21 EcoRI-MseI and 17 PstI-MseI AFLP primer combinations, 36 anchored SSR or STS primer pairs, and 9 anchored RFLP probes. The 164-sib TTC1 map includes 1069 AFLP markers and 38 anchor loci in 14 linkage groups spanning 2001 cM. The 170-sib TTC2 map contains 1002 AFLP markers and 36 anchor loci in 14 linkage groups spanning 2066 cM. Some 488 homologous AFLP loci and 24 anchor markers detected in both populations showed similar map order. Thus, 1583 AFLP markers and 50 anchor loci were mapped into 14 linkage groups, which evidently correspond to the 14 chromosomes of allotetraploid Leymus (2n = 4x = 28). Synteny of two or more anchor markers from each of the seven homoeologous wheat and barley chromosomes was detected for 12 of the 14 Leymus linkage groups. Moreover, two distinct sets of genome-specific STS markers were identified in these allotetraploid Leymus species. These Leymus genetic maps and populations will provide a useful system to evaluate the inheritance of functionally important traits of two divergent perennial grass species.