As one of the largest and most diverse classes of specialized metabolites in plants, terpenoids (oprenoid compounds, a type of bio-based material) are widely used in the fields of medicine and light chemical products. They are the most important secondary metabolites in coniferous species and play an important role in the defense system of conifers. Terpene synthesis can be promoted by regulating the expressions of terpene synthase genes, and the terpene biosynthesis pathway has basically been clarified in Pinus massoniana, in which there are multiple rate-limiting enzymes and the rate-limiting steps are difficult to determine, so the terpene synthase gene regulation mechanism has become a hot spot in research. Herein, we amplified a PmDXR gene (GenBank accession no. MK969119.1) of the MEP pathway (methyl-erythritol 4-phosphate) from Pinus massoniana. The DXR enzyme activity and chlorophyll a, chlorophyll b and carotenoid contents of overexpressed Arabidopsis showed positive regulation. The PmDXR gene promoter was a tissue-specific promoter and can respond to ABA, MeJA and GA stresses to drive the expression of the GUS reporter gene in N. benthamiana. The DXR enzyme was identified as a key rate-limiting enzyme in the MEP pathway and an effective target for terpene synthesis regulation in coniferous species, which can further lay the theoretical foundation for the molecularly assisted selection of high-yielding lipid germplasm of P. massoniana, as well as provide help in the pathogenesis of pine wood nematode disease.

Pinus massoniana is a major fast-growing timber tree species planted in arid areas of south China, which has a certain drought-resistant ability. However, severe drought and long-term water shortage limit its normal growth and development. Therefore, in this study, physiological indices, and the transcriptome sequencing and cloning of AP2/ERF transcription factor of P. massonsiana were determined to clarify its molecular mechanism of drought stress. The results showed that stomatal conductance (Gs) content was significantly decreased, and superoxide dismutase (SOD) activity, and malondialdehyde (MDA) and abscisic acid (ABA) content were significantly increased under drought stress. Transcriptomic analysis revealed that compared to the control, 9, 3550, and 4142 unigenes with differential expression were identified by comparing plants subjected to light, moderate or severe drought. AP2/ERF with high expression was screened out for cloning. To investigate the biological functions of ERF1, it was over-expressed in wild-type Populus davdianaand × P. bolleana via the leaf disc method. Under drought stress, compared to wild-type plants, ERF1 over-expressing poplar lines (OE) maintained a higher photosynthetic rate and growth, while the transpiration rate and stomatal conductance significantly decreased and water use efficiency was improved, indicating that drought tolerance was enhanced. This study provides an insight into the molecular mechanism of drought stress adaptation in P. massoniana.

Pinus massoniana Lamb. is the timber species with the widest distribution and the largest afforestation area in China, providing a large amount of timber, turpentine and ecological products. but low temperature limits its growth and geographical distribution. Physiological and molecular studies can well explain the mechanism of P. massoniana response to low temperature. In this study, physiological and biochemical indexes, cell morphology, lignin content, gene regulatory networks, and gene expression patterns of different P. massoniana varieties (cold-tolerant and cold-sensitive) were studied from physiological, biochemical, and molecular perspectives. The results indicated that under low-temperature stress, the cold-tolerant cultivar maintained high contents of osmoregulatory substances, and the root morphology and structure remained intact. In the initial stage of low-temperature stress, the number of differentially expressed genes was 7148, and with the extension of stress time, the number of differentially expressed genes decreased to 1991. P. massoniana might direct its responses to low temperature by regulating phenylpropane metabolism, starch and sucrose metabolism, hormone signaling pathways, and transcription factors. BAM, 4CL, CCoAOMT, PRX5, WRKYs, and hormone synthesis related genes play important roles. P. massoniana cultivars may vary in response mechanisms. In this study, physiological and analytical techniques were used to study the root tip response mechanism of Masson\'s pine to low temperature stress. The results of this study lay a foundation for in-depth research on the molecular functions of P. massoniana under low-temperature stress conditions.

BACKGROUND: Pinus massoniana Lamb. is the timber species with the widest distribution and the largest afforestation area in China, providing a large amount of timber, turpentine and ecological products. Seasonal drought caused by climate warming severely constrains the quality and growth of P. massoniana forests. WRKY transcription factors play an important role in plant responses to abiotic stress. In this study, the molecular mechanisms by which P. massoniana responds to drought stress were analysed based on the P. massoniana WRKY (PmWRKY) family of genes.
RESULTS: Forty-three PmWRKYs are divided into three major families, 7 sub-families, and the conserved motifs are essentially the same. Among these 43 PmWRKYs express under drought stress but with different expression patterns in response to stress. PmWRKYs respond to drought stress induced by exogenous hormones of SA, ABA, and MeJA. The expression of PmWRKY6, PmWRKY10, and PmWRKY30 up-regulate in different families and tissues under drought stress, while PmWRKY22 down-regulate. Transgenetic tobaccos of PmWRKY31 are with lower malondialdehyde (MDA) content and higher proline (Pro) content than wild type (WT) tobaccos. In transgenic tobaccos of PmWRKY31, expression levels of related genes significantly improve, and drought tolerance enhance.
CONCLUSIONS: This study analysed the molecular biological characteristics of PmWRKYs and investigated the expression patterns and functions of PmWRKYs in response to drought stress in P. massoniana. The results of this study provide a basis for in-depth research of the molecular functions of PmWRKYs in response to drought stress.

The normal megastrobilli and microstrobilli before and after the sexual reversal in Pinus massoniana Lamb. were studied and classified using a transcriptomic approach. In the analysis, a total of 190,023 unigenes were obtained with an average length of 595 bp. The annotated unigenes were divided into 56 functional groups and 130 metabolic pathways involved in the physiological and biochemical processes related to ribosome biogenesis, carbon metabolism, and amino acid biosynthesis. Analysis revealed 4,758 differentially expressed genes (DEGs) between the mega- and microstrobili from the polycone twig. The DEGs between the mega- and microstrobili from the normal twig were 5,550. In the polycone twig, 1,188 DEGs were identified between the microstrobili and the sexually reversed megastrobili. Concerning plant hormone signal transduction pathways, the DEGs from both the normal and polycone twigs displayed distinct male or female associated expression patterns. There were 36 common hormone-related DEGs from the two types of twigs of P. massoniana. Interestingly, expression of these DEGs was up-regulated in the bisexual strobili, which underwent the sexual reversal. A portion of MADS-box genes in the bisexual strobili were up-regulated relative to expression in microstrobili.

N and S contents in Cinnamomum camphora leaves, Pinus massoniana Lamb. leaves, epilithic Haplocladium microphyllum(Hedw.) Broth. and rhizosphere soil collected along 3 directions from urban area to rural area at Guiyang city in a total of 296 samples were investigated systematically. The level of atmospheric N deposition and atmospheric SO2 concentrations at each sampling site were calculated according to the quantitative relationship between moss N content and atmospheric N deposition and the quantitative relationship between moss S content and atmospheric SO2 concentration. Leaves N content in Cinnamomum camphora(1.01%-2.37%) and Pinus massoniana Lamb.(0.99%-2.42%)showed significant decrease from urban area with the highest atmospheric nitrogen deposition to suburb, while slightly higher leaves N reemerged at rural area beyond 24 km, suggesting increased N deposition occurred in rural area. Leaves S content in Cinnamomum camphora(0.16%-0.43%) and Pinus massoniana Lamb.(0.18%-0.32%) showed significant decrease from urban area to suburb, the highest level at urban was mainly contributed by the high sulfur released from the production and living of urban areas into the atmosphere, and the lowest level occurred at rural area(30-36km). No significant difference was seen for soil N and S contents. The relationships between the estimated input of atmospheric N deposition and the leaves N content of Cinnamomum camphora and Pinus massoniana Lamb. at the sites investigated were found to be significant linear correlations, and the relationships between the estimated atmospheric SO2 concentration and the leaves S content of Cinnamomum camphora and Pinus massoniana Lamb. were also significant linear correlations(P<0.05). The results indicated that N and S contents in Cinnamomum camphora and Pinus massoniana Lamb. leaves can be used to show the spatial variation of atmospheric N and S deposition.

Prostatitis is a common prostate disease that could be promoted by bacterial or non-bacterial infectious agents. In addition, inflammatory pathways involved in prostatitis have been increasingly studied, and herbal extracts endowed with anti-inflammatory effects are under investigation, individually or in combination, for their efficacy in alleviating the burden of inflammation, with possible improvements in symptoms. Serenoa repens (Serenoa), in combination with Crocus sativus (Crocus) and Pinus massoniana (Pinus), has previously shown to improve sexual function and limit urinary symptoms in patients suffering from concomitant erectile dysfunction and lower urinary tract symptoms. In this context, the aim of the present study is to evaluate the efficacy of Serenoa, Crocus and Pinus extracts, either alone or in combination, on immortalized prostate cells (PC3) and in an experimental model of bacterial prostatitis constituted by ex vivo prostate specimens challenged with lipopolysaccharide (LPS). We found that the tested extracts were able to reduce ROS production by PC3 cells and NFkB and PGE2 activity in prostate specimens challenged with LPS. In addition, the pharmacological association of the extracts displayed synergistic effects indicating a rational use of the mixture of the tested extracts as a novel anti-oxidant and anti-inflammatory formulation in bacterial prostatitis. Finally, we performed analytical and in vitro evaluation to better characterize the phytochemical profile and the mechanism of action of selected secondary metabolites.

Two new sesquiterpene glycosides, namely massonside A (1) and massonside B (2), were isolated from the n-Bu extract of the fresh needles of Pinus massoniana Lamb. Their structures were established by 1D, 2D nuclear magnetic resonance and high-resolution mass spectrometry. Their biological activities were profiled by the anti-HBV and anti-HCV assays.

High-resolution winter temperature reconstructions in China are rare, yet vital for the comprehensive understanding of past climate change. In the present work, the first winter-half year minimum mean temperature from previous November to current April in northwestern Yichang, South Central China, was reconstructed back to 1875 based on tree-ring material. The reconstruction can explain 55 % of the variance over the calibration period during 1955-2011. The temperature maintained at comparatively low level before 1958, and an abnormal warming was seen since 1959. However, the warming trend stagnated after 2000 AD. 2001-2010 was the warmest decade not only during the instrumental period but also during the whole reconstructed period. The reconstruction indicates good spatial resemblance to other temperatures series in adjacent areas and Northern Hemisphere, yet the recent warming in this study is earlier and more prominent than that of Southeast China. This work also manifests that the winter-half year minimum temperature in study area has good agreement with summer (June-September) maximum temperature variation in Southeast China at decadal scale, except that the winter-half year warming in recent decades is more evident than summer. This reconstruction is not only useful in improving our knowledge of long-term temperature variation but also useful in predicting the tree growth dynamics in the future in the study area.