Cytohistological study of the leaf structures of Panax species, and molecular and functional characterization of ginseng UGT72AL1 in Arabidopsis = 고려인삼 잎의 세포구조학적 특징 및 식물 세포 발달에 영향을 미치는 인삼 UGT72AL1 유전자에 관한 연구

다국어 초록 (Multilingual Abstract)

Ginseng, which belongs to Araliaceae family, is the most popular perennial herb worldwide. For decades, ginseng has gained considerable attention on account of its beneficial health effects. Since the 1900s, ginseng has been commercially utilized to produce bioactive products, which to date have been sold in over 35 countries. However, the 4- to 6-year field cultivation of ginseng is a time-consuming and labor-intensive process. Thus, considerable effort has been made to improve ginseng yield and quality via classical and molecular breeding methods, as well as through improvements in cultivation. To shed light on this process, the cytohistology of the leaf structure of two different ginseng species was studied. In addition, functional characterization of a ginseng glycosytransferase gene, PgUGT72AL1, which might be involved in ginseng saponin biosynthesis, specifically the transfer of a glycosyl group to an acceptor, was performed at the molecular and genetic levels.
Both Panax ginseng Meyer and Panax quinquefolius are obligate shade-loving plants, the natural habitats of which are the broad-leaved forests of Eastern Asia and North America. Panax species are easily damaged by photoinhibition when they are exposed to high temperatures or receive insufficient shade. Thus, a cytohistological study of the leaf structures of two of the most well-known Panax species was performed to gain a better understanding of the physiological processes that limit photosynthesis. The mesostructure of both P. ginseng and P. quinquefolius frequently has one layer of non-cylindrical palisade cells and three to four layers of spongy parenchyma cells. P. quinquefolius was found to contain similar number of stomata in the abaxial leaf surface but more tightly appressed enlarged grana stacks than P. ginseng; however, the adaxial surface of the epidermis in P. quinquefolius shows cuticle ridges with a pattern similar to that of P. ginseng. The anatomical leaf structure of both P. ginseng and P. quinquefolius shows that they are typical shade-loving sciophytes. Slight differences in chloroplast structure suggest that the two species can be authenticated using transmission electron microscopic images, and light-resistant cultivar breeding can be performed via the control of photosynthetic efficiency.
Glycosylation of natural compounds gives rise to a great diversity of secondary metabolites. Glycosylation steps are implicated not only in plant growth and development but also in plant defense responses to various environmental stresses. This glycosylation process is mediated by members of the multigene glycosyltransferase (GT) family, which catalyze the transfer of single or multiple activated sugars to a wide range of substrates, thus influencing their chemical properties and bioactivities. Although their activities have been recognized for a long time and the genes coding for uridine diphosphate (UDP)-dependent glycosyltransferases (UGTs) have been identified in several higher plants, details of the specific functions of GTs remain elusive. Considering that UGTs are key enzymes in the production of numerous different types of ginsenosides in P. ginseng, we selected a UTG gene as a candidate for further functional characterization. Here, we report that the PgUGT72AL1 gene is highly expressed in the upper root, rhizome, and youngest leaf of ginseng. Stress responsiveness against various abiotic stresses, GUS histological expression, and subcellular localization in Arabidopsis were also examined. Exogenous overexpression of PgUGT72AL1 in Arabidopsis resulted in fused organ in the axillary branches, indicating the role of this gene in plant development.

목차 (Table of Contents)

• Abstract 1
• Literature review 4
• Chapter 1. Cytohistological study of the leaf structures of Panax species. 16
• Abstract 16
• 1. Introduction 17
• 2. Materials and methods 19
• 2.1. Plant materials and growth conditions 19
• 2.2. Light microscopy and scanning electron microscopy 19
• 2.3. Transmission electron microscopy 20
• 3. Results and discussion 21
• 3.1. Original cultivation areas and characteristics of P. ginseng and P. quinquefolius 21
• 3.2. Abaxial and adaxial leaf-surface structures 23
• 3.3. Internal leaf structure with spherical palisade cells 25
• 3.4. Chloroplast and thylakoid structures 29
• 4. Conclusion 32
• References 34
• Chapter 2. Molecular and functional characterization of ginseng UGT72AL1 in Arabidopsis 38
• Abstract 38
• 1. Introduction 39
• 2. Materials and methods 42
• 2.1. Plant materials and growth conditions 42
• 2.2. DNA analysis 42
• 2.3. Abiotic stresses and hormone treatment 42
• 2.4. RNA isolation and (quantitative) real-time RT-PCR 43
• 2.5. Vector construction and in planta transformation 44
• 2.6. GUS histochemical staining 45
• 2.7. Confocal microscopy 46
• 3. Results 47
• 3.1. Isolation and identification of ginseng PgUGT72AL1 gene 47
• 3.2. Organ-specific expression patterns of PgUGT72AL1 50
• 3.3. Spatial and temporal expression of PgUGT72AL1 in heterologous Arabidopsis 52
• 3.4. Differential transcript levels of PgUGT72AL1 in response to biotic and abiotic stresses 54
• 3.5. Over expression of PgUGT72AL1 in Arabidopsis resulted in axillary organ fusion 56
• 3.6. Subcellular localization of PgUGT72AL1 in Arabidopsis 58
• 4. Discussion 60
• References 63
• Conclusion 69