Fangwei Zhou

Seasonal changes in leaf coloration are a key ecological and ornamental characteristic of Liquidambar formosana. To clarify the molecular basis of this process, we performed an integrated transcriptomic and metabolomic investigation comparing wild-type L. formosana (FX) with the autumn-red cultivar ‘Jinyu’ (JY). Leaves were sampled before and after the color transition. Analyses revealed distinct metabolic pathways driving coloration in each genotype. In JY, the red phenotype was primarily attributed to the activation of anthocyanin biosynthesis, characterized by coordinated upregulation of key structural genes (LfDFR, LfANS, LfBZ1) and significant accumulation of anthocyanins, especially pelargonidin derivatives. Conversely, FX exhibited enhanced flavonol biosynthesis and carotenoid/terpenoid metabolism, leading to greater yellow pigment accumulation and an orange-yellow hue. Weighted gene co-expression network analysis (WGCNA) identified a core module strongly correlated with anthocyanin content in JY, which was significantly enriched with transcription factors from the MYB, bHLH, and WRKY families. These results demonstrate that different L. formosana genotypes employ divergent metabolic strategies for autumn coloration, governed by specific transcriptional regulatory networks. This study provides crucial insights into pigment regulation in woody plants and offers valuable candidate genes for the molecular breeding of ornamental L. formosana cultivars.

Rhododendron lapponicum (L.) Wahlenb., prized for its vibrant and diverse floral displays, holds significant ornamental and ecological value. However, advances in its molecular breeding have been constrained by the absence of reliable tools for accurate gene expression analysis. A fundamental requirement for such studies is the identification of stable reference genes for qRT-PCR. To date, no systematically validated reference genes exist for normalizing gene expression across R. lapponicum cultivars with diverse flower colors, representing a major technical obstacle to elucidating the molecular mechanisms of color formation. This study aimed to fill this gap by systematically identifying and validating optimal reference genes for petal tissues in six distinct R. lapponicum cultivars. We assessed the expression stability of 11 candidate genes using four independent algorithms and integrated the results via RefFinder. Our comprehensive analysis across multiple algorithms consistently identified RlaEF1-α and RlaACT as the most stably expressed reference genes. Their reliability was robustly validated by normalizing the expression of RlaMYB113, a key anthocyanin regulator; the normalized expression levels showed an extremely significant difference between rose-red and white cultivars (p < 0.001) and produced a coherent, phenotype-correlated profile, in contrast to the distorted patterns obtained with unstable references. This study establishes RlaEF1-α and RlaACT as a precise dual-gene internal control for qRT-PCR. By providing a validated normalization framework, our work enables accurate quantification of color-related genes and directly supports molecular breeding efforts aimed at the targeted development and selection of novel R. lapponicum cultivars with desirable and stable flower colors.

Leaf color mutants serve as excellent models for investigating the metabolic pathways involved in chlorophyll biosynthesis, chloroplast development, and photosynthesis in plants. This study aimed to elucidate the mechanisms underlying color formation in the yellow–green leaf mutant (YL) of Magnolia sinostellata by employing physiological, cytological and transcriptomic analyses to compare the mutant with control plants (wild type Magnolia sinostellata, WT). Physiological assessments revealed a reduction in chlorophyll content, particularly chlorophyll b, alongside an increase in the flavonoid level in YL relative to WT. Cytological examinations indicated the presence of defective chloroplasts within the mesophyll cells of the mutants. Transcriptomic analysis identified 8205 differentially expressed genes, with 4159 upregulated and 4046 downregulated. Genes associated with chlorophyll metabolism, flavonoid metabolism, photosynthesis, and signaling pathways were found to play crucial roles in leaf yellowing. In conclusion, this study delineated the phenotypic, physiological, cytological, and transcriptomic differences between YL and WT leaves, offering novel insights into the mechanisms driving leaf yellowing in Magnolia sinostellata.

Paper Mulberry (Broussonetia papyrifera) possesses medicinal, economic, and ecological significance and is extensively used for feed production, papermaking, and ecological restoration due to its ease of propagation, rapid growth rate, and strong stress resistance. The recent completion of the sequencing of the Paper Mulberry genome has prompted further research into the genetic breeding and molecular biology of this important species. A highly stable reference gene is essential to enhance the quantitative analysis of functional genes in Paper Mulberry; however, none has been identified. Accordingly, in this study, the leaves, stems, roots, petioles, young fruits, and mature fruits of Paper Mulberry plants were selected as experimental materials, and nine candidate reference genes, namely, α-TUB1, α-TUB2, β-TUB, H2A, ACT, DnaJ, UBQ, CDC2, and TIP41, were identified by RT-qPCR. Their stability was assessed using the geNorm, Normfinder, Delta Ct, BestKeeper, and RefFinder algorithms, identifying ACT and UBQ as showing the greatest stability. The expression of BpMYB090, which regulates the production of trichomes, was examined in the leaves of plants of the wild type (which have more trichomes) and mutant (which have fewer trichomes) at various developmental stages to validate the results of this study. As a result, their identification addresses a critical gap in the field of Paper Mulberry research, providing a solid foundation for future research that will concentrate on the characterization of pertinent functional genes in this economically valuable species.

Liquidambar formosana Hance is renowned for its rich leaf color and possesses notable advantages, such as robust adaptability, strong resistance to diseases and pests, and rapid growth, making it a preferred choice for urban greening and carbon sequestration forest initiatives. The completion of whole-genome sequencing of L. formosana has spurred an increased interest in exploring the molecular mechanisms underlying seasonal changes in leaf color, marking a significant focus in L. formosana breeding research. However, there is currently a lack of stable reference genes suitable for analyzing the expression patterns of functional genes in L. formosana exhibiting varying leaf colors. This study selected five L. formosana varieties with significant differences in leaf colors. Through the RT-qPCR analysis, and evaluation using BestKeeper, geNorm, NormFinder, Delta Ct, and RefFinder, the expression stability of 14 candidate reference genes was examined. Consequently, two reference genes (LifEF1-α and LifACT) with stable expression, suitable for RT-qPCR of L. formosana with diverse leaf colors, were identified. The stability of these selected reference genes was further validated by examining the LifbHLH137 gene, which promoted the biosynthesis of anthocyanins. This advancement facilitated molecular biology and genetic breeding investigations of L. formosana, providing essential data for the precise quantification of functional genes associated with leaf color variation.

The biodiversity and activity of microorganisms are crucial for litter decomposition, but how litter traits at different stages of decomposition drive changes in microbial communities has yet to be thoroughly explored. In the typical alpine hilly area of the Qinghai–Tibet Plateau, three types of litter at different decomposition stages were selected under a natural Picea crassifolia (Picea crassifolia Kom.) forest: undecomposed (A-1), partially decomposed (A-2), and fully decomposed (A-3). By measuring physicochemical indicators, microbial diversity, and the composition of the litter at different decomposition stages, this study investigates the community changes and responses of bacteria to litter characteristic changes at different decomposition levels. The results show that with the increase in decomposition level, bacterial diversity increases, community structure changes, and network complexity gradually increases, while the changes in fungal communities are insignificant. Structural equation modeling indicates that the first principal component (PC1) of litter properties is significantly negatively correlated with bacterial diversity and positively correlated with bacterial community composition. There is no significant correlation between fungal diversity and community composition, indicating a closer relationship between bacteria and litter characteristics than fungi. In summary, with an increase in litter decomposition level, the diversity and network complexity of bacterial and fungal communities will significantly increase, which is related to the changes in various litter characteristics. This study provides a scientific basis for the regulatory mechanism of litter decomposition and turnover in the alpine hilly area of the Qinghai–Tibet Plateau, specifically in Picea crassifolia forests.

Expansins are a group of cell wall enzyme proteins that help to loosen cell walls by breaking hydrogen bonds between cellulose microfibrils and hemicellulose. Expansins are essential plant proteins that are involved in several key processes, including seed germination, the growth of pollen tubes and root hairs, fruit ripening and abscission processes. Currently, there is a lack of knowledge concerning the role of expansins in woody plants. In this study, we analyzed expansin genes using Populus genome as the study target. Thirty-six members of the expansin gene family were identified in Populus that were divided into four subfamilies (EXPA, EXPB, EXLA and EXLB). We analyzed the molecular structure, chromosome localization, evolutionary relationships and tissue specificity of these genes and investigated expression changes in responses to phytohormone and abiotic stresses of the expansin genes of Populus tremula L. (PtEXs). Molecular structure analysis revealed that each PtEX protein had several conserved motifs and all of the PtEXs genes had multiple exons. Chromosome structure analysis showed that the expansin gene family is distributed on 14 chromosomes. The PtEXs gene family expansion patterns showed segmental duplication. Transcriptome data of Populus revealed that 36 PtEXs genes were differently expressed in different tissues. Cis-element analysis showed that the PtEXs were closely associated with plant development and responses to phytohormone and abiotic stress. Quantitative real-time PCR showed that abscisic acid (ABA) and low-temperature treatment affected the expression of some PtEXs genes, suggesting that these genes are involved in responses to phytohormone and abiotic stress. This study provides a further understanding of the expansin gene family in Populus and forms a basis for future functional research studies.

Fasciclin-like arabinogalactan proteins (FLAs) are a subclass of arabinogalactan proteins (AGPs) containing both AGP-like glycated domains and fasciclin (FAS) domains, which are involved in plant growth and development and synthesis of the cell wall. However, these proteins have not been identified or analyzed in willow, Salix, the sister genus of Populus. In this study, we performed a whole genome study of the FLA gene family of Salix suchowensis and compared it with the FLA gene family of Populus deltoides. The results showed the presence of 40 and 46 FLA genes in P. deltoides and S. suchowensis, distributed on 17 and 16 chromosomes, respectively. Four pairs of tandem repeat genes were found in willow, while poplar had no tandem repeat genes. Twelve and thirteen pairs of duplicated gene fragments were identified in poplar and willow, respectively. The multispecies phylogenetic tree showed that the FLA gene family could be divided into four groups (I–IV), with Group 1 showing significant expansion in woody plants. A gene expression analysis showed that PdeFLA19/27 in Group I of poplar was highly expressed, specifically during the secondary growth period of the stem and the rapid elongation of seed hairs. In the Group I genes of S. suchowensis, SsuFLA25/26/28 was also highly expressed during the secondary growth period, whereas increased expression of SsuFLA35 was associated with seed hair tissue. These results provide important clues about the differences in the FLA gene family during the evolution of herbs and woody plants, and suggest that the FLA gene family may play an essential role in regulating the secondary growth of woody plants. It also provides a reference for further studies on the regulation of secondary growth and seed hair development by FLA genes in poplar and willow.

Salix is a dioecious plant. Research on the molecular regulation mechanism of male and female inflorescence differentiation and development is necessary to analyze sex differentiation in the willow and the underlying mechanisms of unisexual flower development. However, at present, there are no reference genes suitable for stable expression in the process of willow inflorescence development. In this study, Salix suchowensis was used as the research material, nine candidate reference genes (α-TUB1, α-TUB2, ACT, H2A, DnaJ, CDC2, GAPDH, TIP41, β-TUB) were selected, and qRT-PCR technology was used to detect the expression of each candidate reference gene in female and male flowers at different developmental stages and using five algorithms (geNorm, Normfinder, Delta Ct, BestKeeper, and RefFinder) to comprehensively evaluate the stability of candidate reference genes. The results showed that ACT and DnaJ were stably expressed in all samples and could be used as reference genes. In addition, the reliability of the screening results was further verified via an expression pattern analysis of the CFS gene that encodes flower specific transcription factor in different samples. The stable reference genes selected in this study provide the basis for future research on the expression analysis of functional genes related to the development of male and female flowers of S. suchowensis.