It is known that receptor proteins that bind Gemcitabine molecular weight elicitors generate signals that are transmitted to the sites of gene expression via different components, such as Ca2 ion fluxes, medium alkalinization and cytoplasmic acidification, oxi dative burst, jasmonate and nitric oxide etc. Many CDPKs and MAPKs have been identified to play a role in defense responses and also secondary metabolite produc tion. The effect of UV B irradiation on expression of TIA biosyn thetic genes, Tdc and Str, and catharanthine production has been reported previously in C. roseus leaves. The transcription factor GT 1 binds to the promoter region of Tdc in vitro. The functional importance of GT 1 in the induction of Tdc expression by UV light has been demonstrated by point mutations in the GT 1 binding site.

However, the molecular basis of UV B signaling cas cades leading to the induction of expression of Tdc and Str genes and the production of TIAs is largely unknown. It has been observed that the polypeptide wound signal, sys temin specific cell surface receptors initiate a signal trans duction cascade upon UV B irradiation in L. peruvianum cell suspension cultures. In the present study, the sig naling pathways mediating UV B induced catharanthine accumulation in C. roseus suspension cultures were inves tigated. UV B induced alkalinization of the culture medium, generation of hydrogen peroxide, activation of CDPK and MBPK as well as accumulation of catharan thine and stimulation of transcription of Tdc and Str genes were studied.

Inhibitors of binding of ligand cell surface receptors, protein kinases and phosphatases, calcium fluxes and H2O2 were used to dissect the UV B signaling cascade. Results Alkalinization of C. roseus cell suspension medium in response to UV B irradiation and its inhibition by suramin Medium alkalinization an early event occurring in elici tor treated plant cell cultures, has been used as a marker of elicitor responses in studying elicitor binding sites in plant cells. Medium alkalinization is thought to result from elicitor stress induced depolarization of the plasma membrane and subsequent K H exchange with Ca2 influx Cl efflux. To determine whether medium alkalinization is involved in UV B signal transduction as an early event, six day old cells were exposed to UV B irra diation for various time periods and extracellular pH changes were measured in the cell sus pension medium for 120 min.

As shown in Figure 1a, the effect of UV B on medium alkalinization was not dose dependent. However, the kinetics and intensity of this response were Brefeldin_A dependent on their respective exposure times. C. roseus cells showed a rapid increase in the medium pH after UV B irradiation peaking at 10 min with an increase of about 0. 7 units in 5 min irradiated cells.