Characterization and fine mapping of a yellow leaf gene regulating chlorophyll biosynthesis and chloroplast development in cotton (Gossypium arboreum).

Chlorophyll biosynthesis and chloroplast development are essential for photosynthesis and plant growth. Gossypium arboreum, a valuable source of genetic variation for cotton improvement, remains poorly studied for the mechanisms regulating chlorophyll biosynthesis and chloroplast development. Here we created a G. arboreum etiolated leaf and stuntedness (els) mutant that displayed a distinct yellow color of leaves, bracts and stems throughout the whole growth, where chlorophyll accumulation in leaves was reduced and chloroplast development was delayed. The GaCHLH gene, which encodes the H subunit of magnesium chelatase (Mg-chelatase), was screened by MutMap and KASP analysis. Compared to GaCHLH, the gene Gachlh of the mutant had a single nucleotide transition (G to A) at 1549 bp, which causes the substitution of a glycine (G) by a serine (S) at the 517th amino acid, resulting in an abnormal secondary structure of the Gachlh protein. GaCHLH-silenced SXY1 and ZM24 plants exhibited a lower GaCHLH expression level, a lower chlorophyll content, and the yellow-leaf phenotype. Gachlh expression affected the expression of key genes in the tetrapyrrole pathway. GaCHLH and Gachlh were located in the chloroplasts and that alteration of the mutation site did not affect the final target position. The BiFC assay result indicated that Gachlh could not bind to GaCHLD properly, which prevented the assembly of Mg-chelatase and thus led to the failure of chlorophyll synthesis. In this study, the Gachlh gene of G. arboreum els was finely localized and identified for the first time, providing new insights into the chlorophyll biosynthesis pathway in cotton.

GhBES1 mediates brassinosteroid regulation of leaf size by activating expression of GhEXO2 in cotton (Gossypium hirsutum).

KEY MESSAGE: We proposed a working model of BR to promote leaf size through cell expansion. In the BR signaling pathway, GhBES1 affects cotton leaf size by binding to and activating the expression of the E-box element in the GhEXO2 promoter region. Brassinosteroid (BR) is an essential phytohormone that controls plant growth. However, the mechanisms of BR regulation of leaf size remain to be determined. Here, we found that the BR deficient cotton mutant pagoda1 (pag1) had a smaller leaf size than wild-type CRI24. The expression of EXORDIUM (GhEXO2) gene, was significantly downregulated in pag1. Silencing of BRI1-EMS-SUPPRESSOR 1 (GhBES1), inhibited leaf cell expansion and reduced leaf size. Overexpression of GhBES1.4 promoted leaf cell expansion and enlarged leaf size. Expression analysis showed GhEXO2 expression positively correlated with GhBES1 expression. In plants, altered expression of GhEXO2 promoted leaf cell expansion affecting leaf size. Furthermore, GhBES1.4 specifically binds to the E-box elements in the GhEXO2 promoter, inducing its expression. RNA-seq data revealed many down-regulated genes related to cell expansion in GhEXO2 silenced plants. In summary, we discovered a novel mechanism of BR regulation of leaf size through GhBES1 directly activating the expression of GhEXO2.

Influence of Ca2+ on Early Degradation of Cast-In-Situ Mortar Induced by Sulfate-Magnesium Multiple Combined Attack.

Early degradation of cast-in-situ concrete induced by multiple internal-external sulfate combined attacks significantly affects the development of concrete strength. An experimental study regarding the effects of Ca2+ on the early degradation of cast-in-situ mortars subjected to internal-external sulfate and magnesium combined attacks is investigated in this paper. In particular, a specific method for accurately simulating the degradation of cast-in-situ structures was proposed in this experiment. Physical properties (including weight, size changes, and porosity), mechanical properties (including flexural strength and compressive strength), sulfate concentration, and microstructural properties were monitored during 28 days of immersion. The results show that an internal sulfate and magnesium combined attack (ISA-IMA) obviously retards the development of early strength and accelerates the degradation induced by external sulfate attack (ESA). The diffusion path of sulfate ions from outside is blocked by flake-shaped magnesium hydrates, delaying the penetration of external sulfate attacks. However, it is far from neutralizing the strength loss induced by an internal magnesium attack (IMA) at an early age. Premixed excessive Ca2+ would improve the strength development and pore structure of concrete or mortar, enhancing durability against corrosive conditions.

Genome-wide analysis of ZAT gene family revealed GhZAT6 regulates salt stress tolerance in G. hirsutum.

High salt environments can induce stress in different plants. The genes containing the ZAT domain constitute a family that belongs to a branch of the C2H2 family, which plays a vital role in responding to abiotic stresses. In this study, we identified 169 ZAT genes from seven plant species, including 44 ZAT genes from G. hirsutum. Phylogenetic tree analysis divided ZAT genes in six groups with conserved gene structure, protein motifs. Two C2H2 domains and an EAR domain and even chromosomal distribution on At and Dt sub-genome chromosomes of G. hirsutum was observed. GhZAT6 was primarily expressed in the root tissue and responded to NaCl and ABA treatments. Subcellular localization found that GhZAT6 was located in the nucleus and demonstrated transactivation activity during a transactivation activity assay. Arabidopsis transgenic lines overexpressing the GhZAT6 gene showed salt tolerance and grew more vigorously than WT on MS medium supplemented with 100 mmol NaCl. Additionally, the silencing of the GhZAT6 gene in cotton plants showed more obvious leaf wilting than the control plants, which were subjected to 400 mmol NaCl treatment. Next, the expressions of GhAPX1, GhFSD1, GhFSD2, and GhSOS3 were significantly lower in the GhZAT6-silenced plants treated with NaCl than the control. Based on these findings, GhZAT6 may be involved in the ABA pathway and mediate salt stress tolerance by regulating ROS-related gene expression.

Identification of BR biosynthesis genes in cotton reveals that GhCPD-3 restores BR biosynthesis and mediates plant growth and development.

MAIN CONCLUSION: Brassinosteroid (BR) synthesis genes in different cotton species was comprehensively identified, and the participation of GhCPD-3 in the BR synthesis signaling pathway for regulating plant development was verified. Brassinosteroid is a natural steroidal phytohormone that plays fundamental roles in plant growth and development. In cotton, detailed characterization and functional validation of BR biosynthesis genes remain rare. Here, 16, 8 and 9 BR biosynthesis genes were identified in Gossypium hirsutum, Gossypium raimondii and Gossypium arboreum, respectively, and their phylogenetic relationships, gene structures, conserved motifs of the encoded proteins, chromosomal locations were determined and a synteny analysis was performed. Gossypium hirsutum and Arabidopsis BR biosynthesis genes closely clustered in the phylogenetic tree and fragment duplication was likely the primary cause promoting gene family expansion in G. hirsutum. Gene Ontology (GO) and KEGG (Kyoto Encyclopedia of Genes and Genomes) enrichment analysis showed their relevance as BR biosynthesis genes. GhCPD-3 was highly expressed in roots and stems and the loci of single nucleotide polymorphisms (SNPs) were significantly associated with these traits.Ectopic overexpression of GhCPD-3 in the cpd91 Arabidopsis mutant rescued the mutant phenotype by increasing plant height and leaf size in comparison to those of cpd91 and WT plants. Moreover, overexpressed GhCPD-3 in cpd91 mutants showed greater hypocotyl and root lengths than those of cpd91 and WT plants under light and dark conditions, respectively, indicating that BR actively promotes hypocotyl and root growth. Similar to CPD (CONSTITUTIVE PHOTOMORPHOGENIC DWARF), GhCPD-3 restores BR biosynthesis thereby mediating plant growth and development.

Response of phytohormone mediated plant homeodomain (PHD) family to abiotic stress in upland cotton (Gossypium hirsutum spp.).

BACKGROUND: The sequencing and annotations of cotton genomes provide powerful theoretical support to unravel more physiological and functional information. Plant homeodomain (PHD) protein family has been reported to be involved in regulating various biological processes in plants. However, their functional studies have not yet been carried out in cotton. RESULTS: In this study, 108, 55, and 52 PHD genes were identified in G. hirsutum, G. raimondii, and G. arboreum, respectively. A total of 297 PHD genes from three cotton species, Arabidopsis, and rice were divided into five groups. We performed chromosomal location, phylogenetic relationship, gene structure, and conserved domain analysis for GhPHD genes. GhPHD genes were unevenly distributed on each chromosome. However, more GhPHD genes were distributed on At_05, Dt_05, and At_07 chromosomes. GhPHD proteins depicted conserved domains, and GhPHD genes exhibiting similar gene structure were clustered together. Further, whole genome duplication (WGD) analysis indicated that purification selection greatly contributed to the functional maintenance of GhPHD gene family. Expression pattern analysis based on RNA-seq data showed that most GhPHD genes showed clear tissue-specific spatiotemporal expression patterns elucidating the multiple functions of GhPHDs in plant growth and development. Moreover, analysis of cis-acting elements revealed that GhPHDs may respond to a variety of abiotic and phytohormonal stresses. In this regard, some GhPHD genes showed good response against abiotic and phytohormonal stresses. Additionally, co-expression network analysis indicated that GhPHDs are essential for plant growth and development, while GhPHD genes response against abiotic and phytohormonal stresses may help to improve plant tolerance in adverse environmental conditions. CONCLUSION: This study will provide useful information to facilitate further research related to the vital roles of GhPHD gene family in plant growth and development.

Degradation Mechanism of Concrete Subjected to External Sulfate Attack: Comparison of Different Curing Conditions.

Sulfate induced degradation of concrete brings great damage to concrete structures in saline or offshore areas. The degradation mechanism of cast-in-situ concrete still remains unclear. This paper investigates the degradation process and corresponding mechanism of cast-in-situ concrete when immersed in sulfate-rich corrosive environments. Concrete samples with different curing conditions were prepared and immersed in sulfate solutions for 12 months to simulate the corrosion of precast and cast-in-situ concrete structures, respectively. Tests regarding the changes of physical, chemical, and mechanical properties of concrete samples were conducted and recorded continuously during the immersion. Micro-structural and mineral methods were performed to analyze the changes of concrete samples after immersion. Results indicate that the corrosion process of cast-in-situ concrete is much faster than the degradation of precast concrete. Chemical attack is the main cause of degradation for both precast and cast-in-situ concrete. Concrete in the environment with higher sulfate concentration suffers more severe degradation. The water/cement ratio has a significant influence on the durability of concrete. A lower water/cement ratio results in obviously better resistance against sulfate attack for both precast and cast-in-situ concrete.

A novel glycosylated solution from Dioscorea zingiberensis C.H. Wright significantly improves the solvent productivity of Clostridium beijerinckii.

The economics of bio-solvent production are largely dependent on the cost of the fermentation substrate. Dioscorea zingiberensis C.H. Wright (DZW), the main raw material used to produce saponin, contains 13-18% starch which can be directly saccharified to a saccharification liquid of DZW starch (SLDS) that contains abundant nutrients. In this study, the water-soluble micromolecule compounds in SLDS were quantified through 1H NMR. Using SLDS as the substrate to conduct ABE fermentation by Clostridium beijerinckii, the fermentation cycle was shortened 24h, the maximum biomass and consumption rate of the glucose significantly increased, and the productivity of total solvents were increased by 0.244±0.010g/L/h compared to standard P2 medium. Expression analysis of genes encoding key enzymes involved in acetone and butanol production and glucose consumption showed that they were induced by SLDS. Taken together, SLDS is a useful and renewable glycosylated solution for ABE fermentation.