Temporal Gene Expression in Apical Culms Shows Early Changes in Cell Wall Biosynthesis Genes in Sugarcane

Multiple genes in sugarcane control sucrose accumulation and the biosynthesis of cell wall components; however, it is unclear how these genes are expressed in its apical culms. To better understand this process, we sequenced mRNA from +1 stem internodes collected from four genotypes with different concentrations of soluble solids. Culms were collected at four different time points, ranging from six to 12-month-old plants. Here we show differentially expressed genes related to sucrose metabolism and cell wall biosynthesis, including genes encoding invertases, sucrose synthase and cellulose synthase. Our results showed increased expression of invertases in IN84-58, the genotype with lower sugar and higher fiber content, as well as delayed expression of secondary cell wall-related cellulose synthase for the other genotypes. Interestingly, genes involved with hormone metabolism were differentially expressed across time points in the three genotypes with higher soluble solids content. A similar result was observed for genes controlling maturation and transition to reproductive stages, possibly a result of selection against flowering in sugarcane breeding programs. These results indicate that carbon partitioning in apical culms of contrasting genotypes is mainly associated with differential cell wall biosynthesis, and may include early modifications for subsequent sucrose accumulation. Co-expression network analysis identified transcription factors related to growth and development, showing a probable time shift for carbon partitioning occurred in 10-month-old plants.

Microtubule-associated IQD9 guides cellulose synthase velocity to shape seed mucilage

Arabidopsis seeds release large capsules of mucilaginous polysaccharides, which are shaped by an intricate network of cellulosic microfibrils. Cellulose synthase complexes is guided by the microtubule cytoskeleton, but it is unclear which proteins mediate this process in the seed coat epidermis (SCE). Using reverse genetics, we identified IQ67 DOMAIN 9 (IQD9) and KINESIN LIGHT CHAIN-RELATED 1 (KLCR1) as two highly expressed genes during seed development and comprehensively characterized their roles for cell wall polysaccharide biosynthesis and cortical microtubule (MT) organization. Mutations in IQD9 as well as in KLCR1 lead to compact mucilage capsules with aberrant cellulose distribution, which can be rescued by transgene complementation. Double mutant analyses revealed that their closest paralogs (IQD10 and KLCR2, respectively) are not required for mucilage biosynthesis. IQD9 physically interacts with KLCR1 and localizes to cortical MTs to maintain their organization in SCE cells. Similar to the previously identified TONNEAU1 (TON1) RECRUITING MOTIF 4 (TRM4) protein, IQD9 is required to maintain the velocity of cellulose synthases. Our results demonstrate that IQD9, KLCR1 and TRM4 are MT-associated proteins that are required for seed mucilage architecture. This study provides the first direct evidence that members of the IQD, KLCR and TRM families have overlapping roles in guiding the distribution of cell wall polysaccharides. Therefore, SCE cells provide an attractive system to further decipher the complex genetic regulation of polarized cellulose deposition.

Rice Brittle Culm19 Encoding Cellulose Synthase Subunit CESA4 Causes Dominant Brittle Phenotype But has No Distinct Influence on Growth and Grain Yield

Background Mechanical strength is a crucial agronomic trait in rice (Oryza sativa), and brittle mutants are thought suitable materials to investigate the mechanism of cell wall formation. So far, almost all brittle mutants are recessive, and most of them are defected in multiple morphologies and/or grain yield, limiting their application in hybrid breeding and in rice straw recycling. Results We identified a semi-dominant brittle mutant Brittle culm19 (Bc19) isolated from the japonica variety Nipponbare through chemical mutagenesis. The mutant showed the same apparent morphologies and grain yield to the wild type plant except for its weak mechanical strength. Its development of secondary cell wall in sclerenchyma cells was affected, along with reduced contents of cellulose, hemicellulose, lignin and sugars in culms and leaves. Positional cloning suggested that the Bc19 gene was allelic to OsCESA4, encoding one of the cellulose synthase A (CESA) catalytic subunits. In this mutant, a C-to-T substitution occurred in the coding sequence of BC19, causing the P507S missense mutation in its encoded product, which was located in the second cytoplasmic region of the OsCESA4 protein. Furthermore, introducing mutant gene Bc19 into the wild-type plant resulted in brittle plants, confirming that the P507S point mutation in OsCESA4 protein was responsible for the semi-dominant brittle phenotype of Bc19 mutant. Reverse correlation was revealed between cellulose contents and expression levels of mutant gene Bc19 among the homozygous mutant, the hybrid F1 plant, and the Bc19 overexpression transgenic plants, implying that gene Bc19 might affect cellulose synthesis in a dosage-dependent manner. Conclusions Bc19, a semi-dominant brittle mutant allele of gene OsCESA4, was identified using map-based cloning approach. The mutated protein of Bc19 possessing the P507S missense mutation behaved in a dosage-dependent semi-dominant manner. Unique brittle effect on phenotype and semi-dominant genetic quality of gene Bc19 indicated its potential application in grain-straw dual-purpose hybrid rice breeding.

Deep transcriptomic study reveals the role of cell wall biosynthesis and organization networks in the developing shell of peanut pod

Background Peanut (Arachis hypogaea L.) belongs to an exceptional group of legume plants, wherein the flowers are produced aerially, but the pods develop under the ground. In such a unique environment, the pod’s outer shell plays a vital role as a barrier against mechanical damage and soilborne pathogens. Recent studies have reported the uniqueness and importance of gene expression patterns that accompany peanut pods’ biogenesis. These studies focused on biogenesis and pod development during the early stages, but the late developmental stages and disease resistance aspects still have gaps. To extend this information, we analyzed the transcriptome generated from four pod developmental stages of two genotypes, Hanoch (Virginia-type) and IGC53 (Peruvian-type), which differs significantly in their pod shell characteristics and pathogen resistance. Results The transcriptome study revealed a significant reprogramming of the number and nature of differentially expressed (DE) genes during shell development. Generally, the numbers of DE genes were higher in IGC53 than in Hanoch, and the R5-R6 transition was the most dynamic in terms of transcriptomic changes. Genes related to cell wall biosynthesis, modification and transcription factors (TFs) dominated these changes therefore, we focused on their differential, temporal and spatial expression patterns. Analysis of the cellulose synthase superfamily identified specific Cellulose synthase (CesAs) and Cellulose synthase-like (Csl) genes and their coordinated interplay with other cell wall-related genes during the peanut shell development was demonstrated. TFs were also identified as being involved in the shell development process, and their pattern of expression differed in the two peanut genotypes. The shell component analysis showed that overall crude fiber, cellulose, lignin, hemicelluloses and dry matter increased with shell development, whereas K, N, protein, and ash content decreased. Genotype IGC53 contained a higher level of crude fiber, cellulose, NDF, ADF, K, ash, and dry matter percentage, while Hanoch had higher protein and nitrogen content. Conclusions The comparative transcriptome analysis identified differentially expressed genes, enriched processes, and molecular processes like cell wall biosynthesis/modifications, carbohydrate metabolic process, signaling, transcription factors, transport, stress, and lignin biosynthesis during the peanut shell development between two contrasting genotypes. TFs and other genes like chitinases were also enriched in peanut shells known for pathogen resistance against soilborne major pathogens causing pod wart disease and pod damages. This study will shed new light on the biological processes involved with underground pod development in an important legume crop.

Expression Patterns and Gene Analysis of the Cellulose Synthase Gene Superfamily in Eucalyptus grandis

Cellulose is the world’s most abundant renewable energy resource, and a variety of cellulose synthase genes are involved in the biosynthesis of cellulose. In the process of cellulose synthesis, all cellulose synthases are interrelated and act synergistically. In this study, we analyzed the contents of cellulose, hemicellulose, and lignin in the different parts and tissues of E. grandis. The results showed that the cellulose content had greater differences among three different heights. On this basis, we carried out the transcriptome-wide profiling of gene expression patterns using RNA sequencing. A total of 2066 differentially expressed genes were identified for three pairwise comparisons between three different heights, most of which were related to the programmed photosynthetic membrane and photosystem. A total of 100 transcripts of CSs (58 CesA and 42 Csl) were obtained from transcriptome libraries. The expression pattern of these genes indicated that different CS genes had a wide range of expression profiles. A phylogenetic analysis of 135 reference CS genes showed that the CSs of E. grandis were clustered into six major groups (CesA1-9, CslA, CslB/H, CslD, CslE, and CslG). Based on the weighted gene co-expression network analysis, a dual-directional regulation mechanism between Csl and CesA proteins in the cellulose biosynthesis was identified. The gene expression profile analysis, using qRT-PCR in different tissues of E. grandis, demonstrated that the CSs were highly expressed in xylem, and CesAs had a higher relative expression than Csls. The analysis of sequence similarity combined with the expression pattern indicated that the CesA1, 3, and 6 transcripts were associated with the biosynthesis of the secondary cell wall, and CesA4, 5, and 7 transcripts were more likely to associate with the biosynthesis of the primary cell wall. Finally, the qRT-PCR analysis confirmed the expression of 11 selected CSs in three different parts of E. grandis.