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.

Pinus massoniana is an important timber tree species in southern China, and acid aluminum stress seriously endangers its growth. This study focuses on physiology, gene regulation and root exudates. Aluminum stress increased the activity of malondialdehyde (MDA), proline (PRO), peroxidase (POD), soluble proteins (SP), soluble sugars (SS) and superoxide dismutase (SOD) in P. massoniana seedlings, and led to changes in growth. We identified hub genes (UCHL3, TCP1, SEC27, GluRS and ACTF) responding to aluminum stress of low concentration and hub genes (RGP, MPT, RPL24, RPL7A and EC3.2.1.58) responding to aluminum stress of high concentration. Aluminum stress mainly affected phenylpropanoid biosynthesis and flavonoid biosynthesis, and it may alleviate aluminum toxicity by inducing the upregulation of genes such as CHS, COMT, DFR and LAR to enhance root exudation of catechin. These results lay the foundation for in-depth studying the molecular mechanism of P. massoniana aluminum stress.

Background: Pinus massonianaLamb. 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 ofP. massonianaforests. WRKY transcription factors play an important role in plant responses to abiotic stress. In this study, the molecular mechanisms by whichP. massonianaresponds to drought stress were analysed based on theP. massonianaWRKY (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 43PmWRKYsexpress under drought stress but with different expression patterns in response to stress.PmWRKYsrespond to drought stress induced by exogenous hormones of SA, ABA, and MeJA. The expression ofPmWRKY6,PmWRKY10, andPmWRKY30up-regulate in different families and tissues under drought stress, whilePmWRKY22down-regulate. Transgenetic tobaccos ofPmWRKY31are with lower malondialdehyde (MDA) content and higher proline (Pro) content than wild type (WT) tobaccos. In transgenic tobaccos ofPmWRKY31, 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 ofPmWRKYsin response to drought stress inP. massoniana. The results of this study provide a basis for in-depth research of the molecular functions ofPmWRKYsin response to drought stress.

Xylem is required for the growth and development of higher plants to provide water and mineral elements. The thickening of the xylem secondary cell wall (SCW) not only improves plant survival, but also provides raw materials for industrial production. Numerous studies have found that transcription factors and non-coding RNAs regulate the process of SCW thickening. Pinus massoniana is an important woody tree species in China and is widely used to produce materials for construction, furniture, and packaging. However, the target genes of microRNAs (miRNAs) in the developing xylem of P. massoniana are not known. In this study, a total of 25 conserved miRNAs and 173 novel miRNAs were identified via small RNA sequencing, and 58 differentially expressed miRNAs were identified between the developing xylem (PM_X) and protoplasts isolated from the developing xylem (PM_XP); 26 of these miRNAs were significantly up-regulated in PM_XP compared with PM_X, and 32 were significantly down-regulated. A total of 153 target genes of 20 conserved miRNAs and 712 target genes of 113 novel miRNAs were verified by degradome sequencing. There may be conserved miRNA-mRNA modules (miRNA-MYB, miRNA-ARF, and miRNA-LAC) involved in softwood and hardwood formation. The results of qRT-PCR-based parallel validation were in relatively high agreement. This study explored the potential regulatory network of miRNAs in the developing xylem of P. massoniana and provides new insights into wood formation in coniferous species.