Roles of extensins in cotyledon primordium formation and shoot apical meristem activity in Nicotiana tabacum

Extensins are cell wall basic glycoproteins with a polypeptide backbone that is extremely rich in hydroxyproline. In this paper, the function of extensins in embryo development was studied in Nicotiana tabacum. By using Western blot and immunohistochemistry, the extensin JIM20 epitopes were found to express in different developmental stages of embryos, and specifically in the top of the embryo proper (EP) and the suspensor of the late globular embryos. In order to clarify the functions of extensins, a potent hydroxyproline synthesis inhibitor, 3,4-dehydro-L-proline (3,4-DHP), was used in ovule and embryo culture. The results showed that the addition of 3,4-DHP caused abnormal embryos with single, asymmetry and supernumerary cotyledon primordia, and continuous culture led to cotyledon defects in the germinated seedlings. Histological sections showed that the shoot apical meristem (SAM) of the abnormal seedlings was dissimilar from the controls, especially in the seedlings with cup-shaped cotyledons. Furthermore, the vasculature of the abnormal cotyledons was in an out-of-order format and contained at least two main veins. Finally, both the hydroxyproline assay and fluorescent immunolocalization confirmed that 3,4-DHP treatment reduced the level of extensins in the cultured ovules and embryos. These results indicate that extensins may play important roles in the cotyledon primordium formation, SAM activity, and vasculature differentiation during embryo development

Development of a kinetic metabolic model: application to Catharanthus roseus hairy root

A kinetic metabolic model describing Catharanthus roseus hairy root growth and nutrition was developed. The metabolic network includes glycolysis, pentose-phosphate pathway, TCA cycle and the catabolic reactions leading to cell building blocks such as amino acids, organic acids, organic phosphates, lipids and structural hexoses. The central primary metabolic network was taken at pseudo-steady state and metabolic flux analysis technique allowed reducing from 31 metabolic fluxes to 20 independent pathways. Hairy root specific growth rate was described as a function of intracellular concentration in cell building blocks. Intracellular transport and accumulation kinetics for major nutrients were included. The model uses intracellular nutrients as well as energy shuttles to describe metabolic regulation. Model calibration was performed using experimental data obtained from batch and medium exchange liquid cultures of C. roseus hairy root using a minimal medium in Petri dish. The model is efficient in estimating the growth rate