Bochum, Universiẗat, Diss., 2000.

In part I of this thesis are described the structural determinations of sitsirikine, dihydrositsirikine and isositsirikine, three new alkaloids from Vinca rosea Linn. Sitsirikine, C₂₁H₂₆O₃N₂, and dihydrositsirikine, C₂₁H₂₈ O₃N₂, were isolated as an inseparable mixture, which was shown by hydrogenation studies to be comprised of an olefin and its dihydro derivative. The formation of formaldehyde upon ozonisation of the mixture, and of propionic acid in a modified Kuhn-Roth oxidation of dihydrositsirikine demonstrated that, sitsirikine possessed a vinyl group. Both sitsirikine and dihydrositsirikine gave mono-acetates, and the N.M.R. data indicated that primary hydroxyl groups were present in the original alkaloids. A methyl ester function suggested by spectral evidence was established by hydride reduction of dihydrositsirikine to a diol. Since the diol, yielded an acetonide, it was inferred that dihydrositsirikine possessed a β-hydroxy-ester unit. The U.V. spectrum of dihydrositsirikine was characteristic of an indole chromophore, which the mass spectrum showed to be part of a tetrahydro-β-carboline system. Dehydrogenatioh afforded a compound with a flavocoryline-type U.V. spectrum, and this suggested that sitsirikine was a relative of the tetracyclic corynantheine class of alkaloids. This was confirmed by conversion of dihydro-corynantheine into dihydrositsirikine. The structure of the related indole alkaloid isositsirikine, C₂₁H₂₆O₃N₂, was determined by a similar series of reactions. Ozonolysis yielded acetaldehyde, which authenticated the ethylidene group indicated by the N.M.R. spectrum. Acetylation afforded a mono-acetate, whose N.M.R. spectrum suggested that isositsirikine had a primary hydroxyl function. A methyl ester was established by hydride reduction to a diol, which formed an acetonide and hence showed the presence of a β-hydroxy-ester unit in the original alkaloid. Since dehydrogena-tion of dihydro-isositsirikine yielded flavocoryline, a tetracyclic structure very similar to that of sitsirikine could be postulated for isositsirikine. Part II is concerned with the chemistry of cleavamine, a scission product, of the Vinca alkaloid catharanthine. Treatment of catharanthine with aqueous acid in the presence of a reducing agent, led to the isolation of descar-bomethoxycatharanthine, cleavamine and two epimeric dihydro-cleavamines. A tentative mechanism for the reaction is proposed, which can account for the formation of these compounds. Reduction of catharanthine in glacial acetic acid provided carbomethoxy-dihydrocleavamine. Mercuric acetate oxidised this compound to a mixture of two immonium ions, both of which underwent transannular cyclisations. One of the ions gave the known Iboga alkaloids coronaridine and dihydrocathafanthine, whereas the other afforded pseudo-vincadifformine - a synthetic analogue of the known Vinca alkaloid vincadifformine. The structure of pseudo-vincadifformine was determined by conversion into compounds which had U.V., I.R., N.M.R. and mass spectra completely analogous to the corresponding derivatives of vincadifformine. Similar transannular cyclisations to the above are postulated in the scheme advanced by Wenkert for the biogenesis of Iboga and Aspidosperma alkaloids, and the significance of our results with regard to this theory is duscussed. The formation of coronaridine and dihydrocatharanthine in the reaction constituted partial syntheses of these alkaloids, and the potential use of transannular cyclisations in laboratory syntheses of Iboga and Aspidosperma alkaloids is also considered. / Science, Faculty of / Chemistry, Department of / Graduate

In part A, a much sought synthesis of the calycanthaceous alkaloids is described. Oxidative dimerization of N- methyltrytamine afforded dl- chimonanthine and meso-chimonanthine, and dl- calycanthine was produced by subsequent acid-catalyzed rearrangement of the carbon skeleton through a tetraminodialdehyde. As the suggested biosynthesis of these alkaloids is represented as occurring by an oxidative dimerization of N- methyltryptamine (itself a natural product) the very direct synthesis described is a biosynthetic model. The discovery of meso-chimonanthine in an extract of Calycanthus f1oridus is also reported and is the first natural calycanthaceous alkaloid with this stereochemistry to be discovered. As folicanthine and calycanthidine are N-methyl chimonanthines and this methylation has been reported, a synthesis of chimonanthine also represents a formal synthesis of these alkaloids. A proposal for the structure of hodgkinsine, the remaining calycanthaceous alkaloid, is made. A number of synthetic by-products are also described. In part B evidence for the mono-terpenoid origin of the nine or ten carbon non-tryptophan derived portion of the indole alkaloids is presented. The monoterpene, geraniol- 2- ¹⁴C, was administered to Vinca rosea cuttings and the Aspidosperma- type alkaloid, vindoline, was isolated and shown by Kuhn-Roth degradation to be labelled at C-5 as predicted by theory. Feeding experiments with mevalonic acid- 2- ¹⁴C are also described. / Science, Faculty of / Chemistry, Department of / Graduate

Thesis (M. Phil.)--University of Hong Kong, 1981.

Catharanthus roseus est une plante médicinale produisant divers types d’alcaloïdes indoliques monoterpéniques (AIM) d’intérêt en santé humaine. Ainsi, les AIM dimères comme la vinblastine et la vincristine sont utilisés en chimiothérapie anticancéreuse et les alcaloïdes monomères de type hétéroyohimbine présentent diverses activités pharmacologiques. La fabrication de ces molécules dans la plante est fort complexe. Elle requiert un haut niveau de compartimentation tissulaire et subcellulaire et met en jeu plus d’une trentaine d’étapes enzymatiques, dont certaines sont encore très mal connues. Dans ce contexte, l’objectif de la thèse a consisté à élucider plusieurs étapes enzymatiques de la voie de biosynthèse des AIM. Nos travaux ont permis de caractériser de nouvelles isoformes enzymatiques de la famille des cytochromes P450 ainsi que les réductases qui leur sont associées. Ils ont abouti à l’identification de nouvelles déshydrogénases et mis en évidence, in planta, leurs interactions avec la strictosidine synthase suggérant une biosynthèse orientée vers les divers alcaloïdes de type hétéroyohimbine. Enfin, en ayant recours à l’ingénierie métabolique, un segment de la voie de biosynthèse a été transféré dans la levure Saccharomyces cerevisiae, lui conférant la capacité de bio-transformer la tabersonine en vindoline, l’un des deux précurseurs finaux des alcaloïdes dimères. / Catharanthus roseus is a medicinal plant producing various types of monoterpene indole alkaloids (MIA) with a great interest in human health. Dimeric alkaloids such as vinblastine and vincristine are used in cancer chemotherapy and monomeric heteroyohimbine alkaloids exhibit various pharmacological activities. The production of these molecules in the plant is very complex. It requires a high level of tissular and subcellular compartmentalization and involves more than thirty enzymatic steps, some of which are largely unknown. In this context, the aim of this thesis was to elucidate several enzymatic steps of the MIA biosynthetic pathway. Our work allowed us to characterize new enzyme isoforms of cytochrome P450 and their associated reductases. They also resulted in the identification of new dehydrogenases and highlighted their interactions with the strictosidine synthase suggesting a directed biosynthesis towards various heteroyohimbine type of alkaloids. Finally, engineered yeast containing a segment of the MIA biosynthetic pathway was able to convert tabersonine into vindoline, one of the two final precursors of the dimeric alkaloids.

Thesis (M.S.)--Mississippi State University. Department of Plant and Soil Sciences. / Title from title screen. Includes bibliographical references.

Cancer has become a major health problem due to its high rate of morbidity and mortality. Severe side effects associated with most of the available anticancer drugs and the development of the drug resistant cells are the major hurdles limiting their application and therapeutic success. Much attention has thus been focused on natural compounds with minimal or no toxicity in humans and having capacity to suppress proliferation and/or induce apoptosis in cancer cells. Therefore, it is imperative to discover and develop new anticancer drugs. Endophytic fungi though underexploited have remained a rich source of structurally novel and biologically active secondary metabolites. That they are a prolific resource for new compounds in drug discovery is evident from a plethora of reported research findings of the recent past. Those of endophytic fungi especially isolated from medicinal plants with known therapeutic value have especially caught the attention of several research groups worldwide for the production of host associated or novel lead molecules. In this backdrop, the focus of my research work has been centered on exploring the endophytic fungal population associated with Catharanthus roseus in search of production of novel anticancer compounds. Twenty endophytic fungi were isolated from different parts of Catharanthus roseus plant. The ethyl acetate (EA) extracts of three week grown liquid cultures of the individual endophytes were assessed in vitro for their cytotoxic activities in HeLa and HepG2 cancer cells using MTT assay. Of all the isolates thus screened, the EA extract from an isolate identified as Chaetomium nigricolor exhibited significant cytotoxicity in the tested cell lines with a better activity profile against HeLa cells. To understand if the liquid culture filtrate or the mycelium harbored the active cytotoxic principle, the C. nigricolor culture filtrate and mycelia was extracted separately using organic solvents with varying polarity namely ethyl acetate, dichloromethane, chloroform and hexane. These extracts were further tested for cytotoxicity induction in HeLa and MCF-7 cells by MTT assay. The ethyl acetate culture filtrate and mycelial extracts showed best cytotoxic activity on HeLa cells compared to other extracts; therefore, the mycelial and culture filtrate ethyl acetate extracts of C. nigricolor were pooled and was used for further work. Hexane culture filtrate extract of C. nigricolor showed best antiproliferative activity against MCF-7 cells. Thus, the compounds with anticancer potential from both ethyl acetate pooled extracts and hexane culture filtrate extract of C. nigricolor were purified using bioassay guided isolation method. This finally resulted in the isolation of two different anticancer compounds, one each from EA pooled extract and Hexane culture filtrate extracts of C. nigricolor named as compound 1 and compound 2, respectively. Based on various analyses including mass spectrometry (MS and MS-MS) and UV-visible, Fourier transform infrared (FT-IR) and nuclear magnetic resonance (NMR) spectroscopy, the compounds 1 and 2 were identified as 1, 2 Bis (diethylamino) ethene-1, 2 diol (BDED) and Nonacos-4-enoic acid (NA), respectively An improved understanding of the cellular responses to chemotherapeutic agents in cancer cells and the underlying molecular mechanisms gains particular relevance in the efforts to improve the clinical outcome of chemotherapeutic agents. Arresting the growth of cancer cells in one way or the other and induction of apoptosis in a drug specific manner is the hallmark of almost all the clinically used anticancer drugs. Towards this end, evaluation of the cytotoxic effects of the newly identified compounds on various human cancer cells was carried out. BDED was examined for in vitro cytotoxicity of against a panel of five human cancer cell lines namely HeLa, A-431, A549, COLO 205, and MCF-7. Among the cell lines screened, HeLa cells were most vulnerable to BDED treatment with an IC50 value of 27 μM. This cytotoxicity was later identified as resulting from apoptosis induction by BDED which was scored by the characteristic events of membrane externalization of phosphatidylserine, cytoplasmic shrinkage and chromatin condensation which were observed in a large majority of the BDED-treated cells. Further detailed studies were performed to delineate the cellular mechanisms of BDED-induced apoptosis in HeLa cells. Analysis of cell cycle progression by propidium iodide (PI) staining revealed BDED-induced cell cycle arrest in the G1 phase of cell cycle. The results indicated that BDED induced a significant ROS generation and a significant loss of mitochondrial membrane potential which were abrogated upon pretreatment of the cells with an antioxidant, N-acetyl cysteine (NAC). These observations suggested the probable involvement of the initial events of BDED-induced ROS production in causing cell death via mitochondria-based intrinsic pathway of apoptosis. In addition, the results from the expression profile of pro- and anti- apoptotic proteins by western blot analysis in the BDED-treated HeLa cells further corroborated this hypothesis. These included an abnormally deregulated cellular abundance of BAX 1 and BCL 2 proteins, elevated levels of APAF-1, and activated cleaved species of procaspases 9 and 3. In addition, a pronounced abundance of cleaved PARP protein was observed in these experiments. Thus, the results suggest that BDED induced apoptosis in HeLa cells via ROS mediated mitochondrial dependent pathway. In parallel, the in vitro cytotoxic effects of NA was also studied, the second antiproliferative compound identified from hexane extracts of C. nigricolor using a panel of four human cancer cell lines - A-431, A549, COLO 205, and MCF 7. NA was thus found to be most potent against MCF 7 (breast cancer) cells. PI staining-based viability assays and microscopic observations showed a dose dependent cytotoxicity of NA on MCF 7 cells. Our data reveled an IC 50 of 40 μM for NA The NA-induced apoptosis was confirmed by flow cytometric detection of membrane externalization of phosphatidylserine using Annexin V FITC/PI dual staining. NA was also observed to induce cytotoxicity best at 72 h this could be attributed to the fact that it is a derivative of a Nonocosane-(a naturally prevalent molecule found in several vegetables). Similar to our results from BDED-treated HeLa cells, ROS generation and loss of mitochondrial membrane potential which were abrogated on pretreatment with NAC, were also observed in NA-treated MCF-7cells. Similar to several other fatty acids, NA was also observed to induce cell cycle arrest in the G1 phase of cell cycle. Further, apoptotic signature of an altered expression of anti-apoptotic BCL-2 and pro-apoptotic BAX, APAF-1 and procaspase 9 and 3 and a cleaved PARP were observed in NA-treated MCF 7 cells. In conclusion, the results indicate that BDED and NA have cytotoxic and apoptotic effects on HeLa (cervical cancer) and MCF 7 (breast cancer) cell lines, respectively. This leaves open further avenues to evaluate their potential application as anticancer agents for treatment of human cervical and breast cancers.

OLIVEIRA, Francisco Ítalo Fernandes de. Influência do modo de irrigação na tolerância de plantas ornamentais à salinidade da água. 2016. 103 f. Dissertação (Mestrado em Solos e Nutrição de Plantas)-Universidade Federal do Ceará, Fortaleza, 2016. / Submitted by Anderson Silva Pereira (anderson.pereiraaa@gmail.com) on 2017-01-26T16:59:49Z No. of bitstreams: 1 2016_dis_fifoliveira.pdf: 2741362 bytes, checksum: 0461d0245af9c6ece1c9a5111667008f (MD5) / Approved for entry into archive by Jairo Viana (jairo@ufc.br) on 2017-01-26T20:32:19Z (GMT) No. of bitstreams: 1 2016_dis_fifoliveira.pdf: 2741362 bytes, checksum: 0461d0245af9c6ece1c9a5111667008f (MD5) / Made available in DSpace on 2017-01-26T20:32:19Z (GMT). No. of bitstreams: 1 2016_dis_fifoliveira.pdf: 2741362 bytes, checksum: 0461d0245af9c6ece1c9a5111667008f (MD5) Previous issue date: 2016 / World trade of flowers and ornamental plants, of which the most significant branch is cut flowers, followed by the live plants, is an important segment economically to Brazil especially for the Northeast. Although the Northeast region has favorable climatic conditions for the cultivation of flowers and ornamental plants, low quality irrigation water, it is revealed as a problem. The plant species exhibit different mechanisms of tolerance to salt, in some cases, allowing their survival and production. Thus, the research aimed to evaluate the tolerance of four ornamental species to increasing levels of salinity of irrigation water and different modes of supply of water. The research was conducted in a protected environment, in the period from September to November 2015, in the experimental area of the weather station at the University Campus of Pici, the Federal University of Ceará, Fortaleza - Ceará. The treatments were distributed in randomized blocks, arranged in a split plot design with four replications with six levels of salinity of irrigation water to the plots (A1 - 0.6; A2 - 1.2; A3 - 1.8; A4 - 2 4; A5 - 3.0 and A6 - 3.6 dS/m), two modes of irrigation in the subplots (M1 - localized irrigation and M2 - not localized irrigation) and four ornamental species in subsubplots (E1 - Catharanthus roseus; E2 - Allamanda cathartica; E3 - Ixora coccinea and E4 - Duranta erecta), with 48 plants in each block, totaling one hundred ninety-two experimental units. The seedlings of ornamentals were transferred to plastic pots with a volume of 7 liters filled with a gravel layer and substrate, comprising a mixture of sand and earthworm humus, in the ratio 2:1 respectively. They were evaluated biometric, physiological, nutritional and sensorial parameters in plants. The most appropriate variables to classify the species as tolerance to salinity, are related to biomass production. The species are more sensitive to the irrigation method not located than the localized mode. The species that are more tolerant to salinity of irrigation water were Catharanthus roseus and Ixora coccinea. / O comércio mundial de flores e plantas ornamentais, do qual o ramo mais expressivo é o de flores de corte, seguido pelo de plantas vivas, é um segmento importante economicamente para o Brasil, sobretudo, para a região Nordeste. Embora a região Nordeste apresente condições climáticas favoráveis ao cultivo de flores e plantas ornamentais, a água de irrigação de baixa qualidade, revela-se como um problema. As espécies vegetais apresentam diferentes mecanismos de tolerância aos sais, em alguns casos, permitindo sua sobrevivência e produção. Dessa forma, a pesquisa objetivou avaliar a tolerância de quatro espécies ornamentais a níveis crescentes de salinidade da água de irrigação e diferentes modos de fornecimento da água. A pesquisa foi desenvolvida em ambiente protegido, no período de setembro a novembro de 2015, na área experimental da Estação Agrometeorológica, no Campus Universitário do Pici, da Universidade Federal do Ceará, município de Fortaleza – Ceará. Os tratamentos foram distribuídos em blocos casualizados, arranjados em parcelas subsubdivididas, com quatro repetições, com seis níveis de salinidade da água de irrigação nas parcelas (A1 - 0,6; A2 – 1,2; A3 – 1,8; A4 – 2,4; A5 – 3,0 e A6 – 3,6 dS/m), dois modos de irrigação nas subparcelas (M1 - irrigação localizada e M2 - irrigação não localizada) e quatro espécies ornamentais nas subsubparcelas (E1 - Catharanthus roseus; E2 - Allamanda cathartica; E3 - Ixora coccinea e E4 - Duranta erecta), com 48 plantas em cada bloco, totalizando cento e noventa e duas unidades experimentais. As mudas das espécies ornamentais foram transplantadas para vasos de material plástico, com volume de 7 litros, preenchidos com uma camada de brita e com substrato, composto por uma mistura de areia e húmus de minhoca, na proporção de 2:1, respectivamente. Foram avaliados parâmetros biométricos, fisiológicos, nutricionais e sensoriais nas plantas. As variáveis mais adequadas para classificar as espécies, quanto a tolerância a salinidade, são as relacionadas a produção de biomassa. As espécies são mais sensíveis à irrigação pelo modo não localizado do que ao modo localizado. As espécies que se mostraram mais tolerantes a salinidade da água de irrigação foram a Catharanthus roseus e Ixora coccinea.

Thesis (MSc (Biomedical Technology))--Cape Peninsula University of Technology, 2018 / Diabetes mellitus is defined as a group of metabolic disorders characterised by chronic hyperglycaemia due to insufficient production and/or action of insulin and is regarded as one of the major sources of morbidity, mortality and economic burden to the modern society. A large body of scientific evidence support the fact that oxidative stress is elevated in diabetic conditions. Oxidative stress plays a significant role in the development of secondary complications of diabetes including diabetes-linked male sexual dysfunction. The management of sexual dysfunction as a secondary complication of diabetes relies on the management of the underlying diabetic condition. Glycaemic control and increased antioxidant protection are therefore necessary in the management of diabetes-induced oxidative stress male infertility. Pharmacological management of diabetes in form of various antihyperglycaemic, synthetic drugs has improved the outlook of diabetic patients; however, they are expensive, harbour unfavourable adverse effects and some have done little to prevent secondary complications of diabetes including diabetes-induced male sexual dysfunction. In addition to this, access to basic technologies for the management of diabetes mellitus and its secondary complications is still a challenge in low resource areas. Because of these challenges, there is a need to search for alternative remedies such as medication from natural products which are more affordable, well tolerated by the human body and are easily accessible. Medicinal plants are therefore viewed as an easily accessible and potent source of antioxidants capable of scavenging free radicals and fighting diabetes-induced oxidative stress. This study therefore investigated the effects of vindoline; an alkaloid extractable from Cantharanthus roseus in ameliorating diabetes-induced oxidative stress effects in testicular and epididymal tissues using male Wistar rats. Forty-eight (48), 6-week old male Wistar rats weighing between 190-230g with a conventional microbial status were divided into 6 groups, n=8, and used for this research project. Group 1 was the normal control, group 2 comprised non-diabetic rats treated with vindoline, and group 3 was the non-diabetic group of rats treated with glibenclamide- the standard drug for the treatment of diabetes. Group 4 was the diabetic control, group 5 comprised diabetic rats treated with vindoline and group 6 was the diabetic group of rats treated with glibenclamide. Diabetes was induced in group 4, group 5 and group 6 rats by subjecting them to 10% fructose water over a period of 2 weeks and thereafter, administering a single intraperitoneal injection of 40 mg/kg b.w streptozotocin (STZ). Fasting blood glucose levels were measured 72 hours after STZ injection and hyperglycaemia was confirmed where fasting blood glucose levels were more than 18mmol/l. The diabetic control (group 4) had higher fasting blood glucose levels, lower body weights as well as lower testicular and epididymal weights in comparison to the normal control (group 1). Additionally, the extent of lipid peroxidation in testicular and epididymal tissues of the diabetic control (group 4) was higher in comparison to that of the normal control (group 1). The diabetic control had lower testicular and epididymal antioxidant enzyme activities (superoxide dismutase and catalase) and lower oxygen radical absorption capacity (ORAC) in comparison to the normal control. Ferric reducing antioxidant power (FRAP) in testicular and epididymal tissues of the diabetic control (group 4) were not significantly different from those of the normal control (group 1). Treatment of diabetic rats with vindoline (group 5) for 5 weeks significantly reduced fasting blood glucose levels although the extent of reduction could not restore diabetic blood glucose levels to near-normal levels. Overall, treatment of diabetic rats with vindoline was able to minimise testicular oxidative stress as reflected by reduction in testicular malondialdehyde (MDA) levels. Furthermore, results of this study showed an increase in both testicular and epididymal catalase activities, an increase in epididymal SOD, an increase in testicular ORAC as well as an increase in both testicular and epididymal FRAP levels after 5 weeks of treating diabetic rats with vindoline (group 5). Epididymal lipid peroxidation levels, epididymal ORAC levels and testicular SOD levels of diabetic rats treated with vindoline (group 5) were however not significantly different from those of the diabetic control (group 4). Treatment of diabetic rats with vindoline or glibenclamide could not restore total body weights and testicular weights of group 5 and group 6 rats respectively, to near-normal levels. Furthermore, epididymal weights and testicular SOD activity of diabetic rats treated with vindoline (group 5) were not significantly different from those of the normal control (group 1). In conclusion, findings from this study demonstrated that treatment with vindoline could have protective effects against diabetes-induced oxidative stress in both testicular and epididymal tissues of male Wistar rats. Vindoline can therefore be considered a potential agent for the management of diabetes-induced oxidative stress male sexual dysfunction. Further studies with advanced technologies are however recommended to study the possible efficacy of vindoline in ameliorating diabetes-induced oxidative stress male sexual dysfunction. Furthermore, studies on the dose-dependent effects and long-term effects of vindoline administration on male reproductive function as well as the overall safety of treatment with vindoline are necessary.

Catharanthus roseus (Madagascar periwinkle), is a well-known medicinal plant that produces a vast array of terpenoid indole alkaloids (TIAs), including two anticancer compounds vinblastine and vincristine. Industrial scale production of TIAs is hampered by the difficulties of total chemical synthesis of these compounds and the fragmented knowledge on TIA pathway. Transcriptional regulation of the TIA biosynthetic pathway has not been thoroughly investigated in Catharanthus and only a few structural genes have been identified as the targets of two master regulators: the basic helix-loop-helix (bHLH) transcription factor (TF) CrMYC2 and APETALA2/ETHYLENE RESPONSE FACTOR (AP2/ERF), ORCA3. Next generation sequencing (NGS) has been used as a tool to isolate novel genes encoding enzymes and regulators of TIA pathway in Catharanthus. In this dissertation, I have performed the transcriptomic analysis of transgenic Catharanthus hairy roots ectopically expressing a dominant repressive form of CrMYC2 or ORCA3 in order to understand their potential impact on the TIA transcriptional regulatory network and to identify and characterize novel target(s) of these two key TFs. MYC2 acts as regulatory hub involved in diverse aspects of plant growth, development, and specialized metabolite biosynthesis by coordinating the crosstalk among different phytohormone signals. CrMYC2 was initially identified in Catharanthus as a regulator of ORCA3. CrMYC2 transactivates ORCA3 by binding to the T/G-box in jasmonate-responsive element (JRE) of ORCA3 promoter. RNA interference (RNAi) mediated knockdown of CrMYC2 strongly reduced TIA accumulation in Catharanthus cell suspension culture. However, the potential influence of CrMYC2 on the expression of other regulatory and structural genes in the TIA pathway remains poorly understood. Transcriptomic analyses revealed that CrMYC2 plays an essential role in JA-induced gene expression and the differentially expressed genes are involved in diverse aspects of growth and development as well as abiotic and biotic stress responses in Catharanthus. Additionally, the expression of genes related to auxin, ethylene, and abscisic acid signaling cascades were affected in hairy roots with modified CrMYC2 expression, suggesting this TF mediates cross-talk between JA and other phytohormones. Surprisingly, overexpression of CrMYC2 resulted in repressed expression of TIA pathway genes in transgenic hairy roots. Expressions of key activators of indole and iridoid pathway were downregulated whereas expression of repressors were upregulated in CrMYC2 hairy roots. Activators (i.e. CrMYC2 and ORCA3) and repressors (i.e. G-box binding factors; GBFs) have been isolated and characterized for their role in regulation of TIA pathway. However, the interconnection between those regulators and the underlying molecular mechanism has not been throughly studied. I identified (i) the interaction of CrMYC2 with CrGBFs and (ii) how this cross-family transcription factor interactions fine-tunes TIA biosynthesis in Catharanthus. The expression profiles of CrMYC2 and CrGBFs were highly correlated in different tissues and in response JA. Moreover, CrMYC2 interacted with CrGBF1 and CrGBF2 in both yeast and plant cells. CrGBF1 and CrGBF2 could form homo- and hetero-dimer which bound T/G-box elements of TIA pathway gene promoters. In plant cells, CrGBF1 antagonizes the activity of CrMYC2 on target promoters in a dosage dependent-manner. Similarly, CrMYC2 can overcome CrGBF1-mediated repression of target promoters in a dosage dependent manner. ORCA3 is another major regulator of TIA biosynthesis in Catharanthus. The transcriptomic analysis of ORCA3 transgenic hairy roots revealed (i) the effect of ORCA3 on newly identified TIA pathway biosynthetic enzymes; (ii) identify the potential effect of ORCA3 on three biological processes: abiotic stress response, plant secondary metabolic process, and response to hormonal stimulus; and (iii) the identification of potential regulator(s) of TIA biosynthesis using ORCA3 based co-expression analysis.