Towards understanding the molecular regulation of somatic embryogenesis in cotton

Functional Genomics

poster

Yang, Xiyan; Zhang, Xianlong

Yang, Xiyan

Zhang, Xianlong

Somatic embryogenesis (SE) is the developmental process by which somatic cells of higher plants can dedifferentiate and reorganize into new plants. It is a notable illustration of cell totipotency, representing a unique developmental pathway of characteristic events, during which cells dedifferentiate, activate their cell division, and reprogram their physiology, metabolism, and gene expression patterns. However, the precise molecular mechanisms regulating SE remain unclear. The Digital Gene Expression (DGE) system, in combination with small RNA sequencing, parallel analysis of RNA ends (PARE) and comparative transcriptome analysis using materials from different development time-point/stages of somatic embryogenesis and zygotic embryos (ZEs) from Gossypium hirsutum cv. YZ1, were conducted in cotton, a typical plant species in SE. Genome-wide profiling of gene expression allowed the identification of novel molecular markers characteristic of this developmental process. DGE profiling and functional assignments of the genes differentially expressed during cotton somatic embryogenesis indicated significant transcriptional complexity during this process. Bioinformatics analysis showed that the genes were enriched for basic processes such as metabolic pathways and biosynthesis of secondary metabolites. Transcription factor–encoding genes were found to be differentially regulated during SE. The complex pathways of auxin abundance, transport and response with differentially regulated genes revealed that the auxin-related transcripts belonged to IAA biosynthesis, indole-3-butyric acid (IBA) metabolism, IAA conjugate metabolism, auxin transport, auxin-responsive protein / indoleacetic acid-induced protein (Aux/IAA), auxin response factor (ARF), small auxin-up RNA (SAUR), Aux/IAA degradation, and other auxin-related proteins, which allow an intricate system of auxin utilization to achieve multiple purposes in SE. A total of 4242 differentially expressed genes were identified in at least one developmental stage by comparative transcriptome analysis using three parallel development stages of the two types of embryos from Gossypium hirsutum cv. YZ1. Expression pattern and functional classification analysis based on these differentially expressed genes revealed that SE development exhibited remarkable and broad stress responses. RT-PCR for selected stress-related genes with different expression levels further confirmed that the higher expression levels of stress-related genes in SEs than in ZEs. Moreover, stress treatment induced by NaCl and ABA accelerated SE development and increased the transcription of genes related to stress response, in parallel with decelerated proliferation of embryogenic calli under stress treatment. Our data reveal that SE development is the process of plant cells adapting to stress conditions, through genetic adjustment of the balance between cell proliferation and differentiation. In the small RNA profiling, A total of 36 known miRNA families were found to be differentially expressed, by comparing seedling hypocotyl and embryogenic callus (EC) of G. hirsutum YZ1, of which 19 miRNA families were represented by 29 precursors. 25 novel miRNAs were identified. 234 transcripts in EC and 322 transcripts in control (CK) were found to be the targets of 23 and 30 known miRNA families respectively, and 16 transcripts were targeted by 8 novel miRNAs. Several targets, which were enriched in auxin signaling pathway, were further validated via RNA ligase-mediated 5′ rapid amplification of cDNA ends (RLM 5′-RACE). The comprehensive transcriptome dynamics, together with biochemical and histological approaches, present a new insight into the molecular mechanisms of somatic embryogenesis and provide strategies that can be used for regulating the developmental processes of somatic embryogenesis in plants.