The Chalcone Isomerase Family in Cotton: Whole-Genome Bioinformatic and Expression Analyses of the Gossypium barbadense L. Response to Fusarium Wilt Infection.

Chalcone isomerase (CHI) is a key component of phenylalanine metabolism that can produce a variety of flavonoids. However, little information and no systematic analysis of CHI genes is available for cotton. Here, we identified 33 CHI genes in the complete genome sequences of four cotton species (Gossypium arboretum L., Gossypium raimondii L., Gossypium hirsutum L., and Gossypium barbadense L.). Cotton CHI proteins were classified into two main groups, and whole-genome/segmental and dispersed duplication events were important in CHI gene family expansion. qRT-PCR and semiquantitative RT-PCR results suggest that CHI genes exhibit temporal and spatial variation and respond to infection with Fusarium wilt race 7. A preliminary model of CHI gene involvement in cotton evolution was established. Pairwise comparison revealed that seven CHI genes showed higher expression in cultivar 06-146 than in cultivar Xinhai 14. Overall, this whole-genome identification unlocks a new approach to the comprehensive functional analysis of the CHI gene family, which may be involved in adaptation to plant pathogen stress.

[Electroacupuncture Intervention Improves Cartilage Degeneration and Subchondral Bone Osteoporosis of Knee-joint Possibly by Adjusting OPG/RANK/RANKL Signaling in Ovariectomized Rats].

OBJECTIVE: To observe the effect of electroacupuncture (EA) on histopathological changes of cartilage and subchondral bone and osteoprotegerin (OPG)，receptor activator of nuclear factor-κB (RANK), and RANK ligand (RANKL) (OPG/RANK/RANKL) signaling and the expression of matrix metalloproteinase-13 (MMP-13) in ovariectomized(OVX)rats, so as to explore its mechanism underlying improvement of osteoporosis. METHODS: Three-month female SD rats were randomly divided into sham operation, model and EA groups (n＝8 in each group). The ovoariectomy model was established by resection of bilateral ovaries. Rats of the sham group were treated by simple removal of a piece of adipose tissue around the bilateral ovaries. EA (3 Hz/15 Hz, 1 mA) was applied to bilateral "Sanyinjiao" (SP 6), "Yanglingquan" (GB 34), "Yinlingquan" (SP 9) and "Zusanli" (ST 36) for 30 min, once daily (except the weekends) for 12 weeks. The histopathological changes of the subchondral bone of the right knee-joint were observed after Saffron O dyeing and evaluated by Mankin's score, and its anatomical structure including the bone volume fraction (bone volume/tissue volume, BV/TV), trabecular bone number (Tb. N) and trabecular thickness (Tb.Th), trabecular separation (Tb.Sp) was observed by using Micro CT imaging. The urine C-terminal cross-linking telopeptide of type Ⅰ collagen (CTX-Ⅰ), CTX-Ⅱ (two bone resorption markers) and serum estrogen (E 2) contents were assayed by ELISA, and the expression levels of OPG, RANKL and MMP-13 mRNAs in the cartilage tissue of the left knee-joint were detected by quantitative RT-PCR. RESULTS: Following modeling, the BV/TV, Tb. N and Tb.Th levels were significantly decreased (P<0.01) and Tb.Sp and Mankin's score obviously increased in the model group compared with the sham group (P<0.01), suggesting a formation of osteoporosis and degeneration of the cartilage tissue. The serum E 2 content and OPG mRNA level in the cartilagetissue were significantly down-regulated (P<0.01), and urine CTX-Ⅰ and CTX-Ⅱ contents and RANKL mRNA and MMP-13 mRNA expression levels cartilagetissue were considerably up-regulated in the model group relevant to the sham group (P<0.01). After EA intervention, modeling-induced decrease of BV/TV, Tb.N, Tb.Th, E 2 and OPG mRNA levels and OVX-induced increase of Tb.Sp, Mankin's score, CTX-Ⅰ， CTX-Ⅱ， RANKL mRNA and MMP-13 mRNA levels were all completely reversed in the EA group relevant to the model group (P<0.01，P<0.05)．. CONCLUSION: EA intervention can inhibit subchondral bone osteoporosis and articular cartilage degeneration of knee-joint in OVX rats, which is closely associated with its effects in inhibiting the down-regulation of serum E 2 and OPG mRNA expression and up-regulation of CTX-Ⅰ， CTX-Ⅱ， RANKL mRNA and MMP-13 mRNA levels, including adjusting OPG/RANK/RANKL signaling.

W55a encodes a novel protein kinase that is involved in multiple stress responses.

Protein kinases play crucial roles in response to external environment stress signals. A putative protein kinase, W55a, belonging to SNF1-related protein kinase 2 (SnRK2) subfamily, was isolated from a cDNA library of drought-treated wheat seedlings. The entire length of W55a was obtained using rapid amplification of 5' cDNA ends (5'-RACE) and reverse transcription-polymerase chain reaction(RT-PCR). It contains a 1,029 -bp open reading frame (ORF) encoding 342 amino acids. The deduced amino acid sequence of W55a had eleven conserved catalytic subdomains and one Ser/Thr protein kinase active-site that characterize Ser/Thr protein kinases. Phylogenetic analysis showed that W55a was 90.38% homologous with rice SAPK1, a member of the SnRK2 family. Using nullisomic-tetrasomic and ditelocentric lines of Chinese Spring, W55a was located on chromosome 2BS. Expression pattern analysis revealed that W55a was upregulated by drought and salt, exogenous abscisic acid, salicylic acid, ethylene and methyl jasmonate, but was not responsive to cold stress. In addition, W55a transcripts were abundant in leaves, but not in roots or stems, under environmental stresses. Transgenic Arabidopsis plants overexpressing W55a exhibited higher tolerance to drought. Based on these findings, W55a encodes a novel dehydration-responsive protein kinase that is involved in multiple stress signal transductions.

Isolation and functional characterization of HvDREB1-a gene encoding a dehydration-responsive element binding protein in Hordeum vulgare.

A gene encoding Hordeum vulgare dehydration-responsive element binding protein 1 (HvDREB1), a member of the A-2 subgroup of the DREB subfamily, was isolated from barley seedlings. A subcellular localization assay revealed accumulation of HvDREB1 protein in the nucleus. As a trans-acting factor, HvDREB1 was able to bind to DRE/CRT elements and transactivate reporter gene expression in yeast cells. A study of various deletion mutants of HvDREB1 proteins indicated that the transactivation activity was localized to the N-terminal region. Expression of the HvDREB1 gene in barley leaves was significantly induced by salt, drought, and low-temperature. In contrast to most A-2 subgroup members in Arabidopsis thaliana, HvDREB1 also responded to exogenous ABA. Overexpression of HvDREB1 activated a downstream gene, RD29A, under normal growth conditions and led to increased tolerance to salt stress in Arabidopsis plants. These results suggest that HvDREB1 produces a DRE-/CRT-binding transcription factor that may have an important role in improving salt tolerance in plants.

Characterization of the TaAIDFa gene encoding a CRT/DRE-binding factor responsive to drought, high-salt, and cold stress in wheat.

Dehydration responsive element-binding factors (DBFs) belong to the AP2/ERF superfamily and play vital regulatory roles in abiotic stress responses in plants. In this study, we isolated three novel homologs of the DBF gene family in wheat (Triticum aestivum L.) by screening a drought-induced cDNA library and designated them as TaAIDFs (T. aestivum abiotic stress-induced DBFs). Compared to TaAIDFb and TaAIDFc, TaAIDFa lacks a short Ser/Thr-rich region, a putative phosphorylation site, following the AP2/ERF domain. The TaAIDFa gene, located on chromosome 3BS, is interrupted by a single intron at the 17th Arg (R) in the N-terminal domain. The N-terminal region of the TaAIDFa protein modulates nuclear localization. The TaAIDFa protein is capable of binding to CRT/DRE elements in vitro and in vivo, and of trans-activating reporter gene expression in yeast cells. The TaAIDFa promoter, with various stress-related cis-acting elements, drives expression of the GUS reporter gene in wheat calli under stress conditions. This was further confirmed by responses of TaAIDFa transcripts to drought, salinity, low-temperature, and exogenous ABA. Furthermore, overexpression of TaAIDFa activated CRT/DRE-containing genes under normal growth conditions, and improved drought and osmotic stress tolerances in transgenic Arabidopsis plants. These results suggested that TaAIDFa encodes a CRT/DRE element-binding factor that might be involved in multiple abiotic stress signal transduction pathways.

Isolation and molecular characterization of the Triticum aestivum L. ethylene-responsive factor 1 (TaERF1) that increases multiple stress tolerance.

ERF transcription factors play important roles in regulating gene expression under abiotic and biotic stresses. The first member of the ERF gene family in wheat (Triticum aestivum L.) was isolated by screening a drought-induced cDNA library and designated as T. aestivum ethylene-responsive factor 1 (TaERF1), which encoded a putative protein of 355 amino acids with a conserved DNA-binding domain and a conserved N-terminal motif (MCGGAIL). The TaERF1 gene was located on chromosome 7A. Protein interaction assays indicated that TaERF1, with a putative phosphorylation site (TPDITS) in the C-terminal region, was a potential phosphorylation substrate for TaMAPK1 protein kinase. Deletion of the N-terminal motif enhanced the interaction of TaERF1 with TaMAPK1. The predicted TaERF1 protein contained three putative nuclear localization signals (NLSs), and three NLSs modulated synergistically the activity of subcellular localization. As a trans-acting factor, TaERF1 was capable of binding to the GCC-box and CRT/DRE elements in vitro, and of trans-activating reporter gene expression in tobacco (Nicotiana tabacum L.) leaves. Transcription of the TaERF1 gene was induced not only by drought, salinity and low-temperature stresses and exogenous ABA, ethylene and salicylic acid, but also by infection with Blumeria graminis f. sp. tritici. Furthermore, overexpression of TaERF1 activated stress-related genes, including PR and COR/RD genes, under normal growth conditions, and improved pathogen and abiotic stress tolerance in transgenic plants. These results suggested that the TaERF1 gene encodes a GCC-box and CRT/DRE element binding factor that might be involved in multiple stress signal transduction pathways.