The Molecular Mechanism Of The Cold Response And Signaling Pathways In Lilium Lancifolium

Lily (Lilium spp.) is an important commodity flower, which was limited and impacted by the cold temperature and other abiotic stresses. Therefore, cultivation applications of strong lily resources are imperative. Lilium lancifolhim, a very important cold-resistant wild flower for lily cold resistance breeding, is widely distributed in southwestern and northeastern China, where the winter temperate can fall as low as -35℃, but it can survive exposure, acclimate to low or freezing temperatures and continuously germinate in the next spring. To gain a better understanding of the cold signaling pathway and the molecular metabolic reactions involved in the cold response, we performed a genome-wide transcriptional analysis using RNA-Seq.1. Approximately 104,703 million clean 90- bp paired-end reads were obtained from three libraries (CK 0h, Cold-treated 2 h and 16 h at 4℃); 18,736 unigenes showed similarity to known proteins in the Swiss-Prot protein database.15,898,13,705 and 1849 unigenes aligned to existing sequences in the KEGG and COG databases (comprising 25 COG categories) and formed 12 SOM clusters, respectively.2. The research identified the sixtheen transcription factors families including NAC, CytochromeP450, Protease, WRKY, MYB, bZIP, ERF, Lipase, AP2/ERF, ARF, RING finger, CIPK, DREB, bHLH, Pathogen-related protein, MADS-box. In each transcription factor family, MYB gene family has the largest stress response genes, followed by Protease, NAC, ERF, bZIP, Cytoclvome P450, CIPK gene family.3.The conduct cold signals to signal transduction genes such as LUCE, LICDPK and transcription factor genes such as L/DREB1/CBF, L1AP2/EREBP, LINAC1, LIR2R3-MYB and L/BZIP, which were expressed highly in bulb. LIFAD3, L1β-amylase, L1P5CS and LICLS responded to cold and enhanced adaptation processes that involve changes in the expression of transcripts related to cellular osmoprotectants and carbohydrate metabolism during cold stress.4. Based on qRT-PCR, GO, COG and KEGG analysis results, we studied three signal regulation pathways -the Ca2+ and ABA independent/dependent pathways. Such regulation of gene expression plays important roles in controlling the physiological cold response, cellular morphology, proliferation, survival, and differentiation of a wide variety of plant cells, as well as being implicated in learning and memory.5. In our present study, the analysis of the structural characteristics of L. lancifolium leaf cells showed that leaf thickness doubles compared with room-temperature controls during cold treatment. Proline may contribute to a remarkable control of cellular electrical conductivity, which reflects the destruction of cell wall and cytoplasm motivation, furthermore regulating outstanding cold tolerance for L. lancifolium. The actual physiology research proves that the L. lancifolium could adapt to low temperature better than Oriental hybrids and Asiatic lily.6. The proteomics technologies were used in the three samples as CKO h, CT16 h and CT16 h+ ABA treatments for L. lancifolium. We found 149 significantly differentially expressed protein spots, and 21 of which were identified by mass spectrometry and caught complete protein peptide fingerprinting.7. During the CT16 h and CT16 h+ABA protein samples comparison, we selected ten significant differences protein with functional characterization, discovering six proteins up-regulated arnd other three proteins down-regulated under ABA treatment.8. In the COG classification for L. lancifolium, the transcriptome function involved the "material metabolism" (21.5%) and "Signal transduction mechanisms" (17.54%), explaining Metabolism process and STM genes were plays an important role in cold stress and cold reaction. This shows that the transcriptome gene function and protein functional classification is basically same. These proteins are involved in different processes in response to cold signal transduction, leading to changes in various physiological indices and plant stress response.9. By RACE technique, transcriptome sequencing LIAPL was full-length cloned, with the full length 1303bp. The gene can encode 278 amino acids. LIAPL was induced by low temperature, high temperature and high salt stress, without ABA and JA hormones. The research was constructed the LIAPL expression vector with pBI121. After using the transformation of Arabidopsis pollen tube pathway, we received the T1 and T2 lines transgenic Arabidopsis. The detection of phenotypic and resistance was observed, showing that the transcription factors LlAPLcould devote to providing the approach of breeding the cold-tolerant plants through genetic manipulation tolerance features. Particularly, for the modern agricultural practices, our comprehensive research of the genes families’ expression and function in the extreme xerophytes, halophytes, cold resistance and heat-resisting plants as L. lancifolium will definitely promote the breeding work for the excellent stress-tolerant flowers.