Modulation of key nitrogen assimilating enzymes by NAA and in vitro culture in Cuscuta reflexa

Abstract

Nitrogen assimilation in the callus of an angiosperm holoparasitic plant, Cuscuta reflexa, has been investigated by studying the level of key enzymes of the nitrogen assimilation pathway, namely nitrate reductase (NR), glutamine synthetase (GS), glutamate synthase (GOGAT) and glutamate dehydrogenase (GDH), during its growth in the absence and presence of NAA. The activity of all these enzymes in culture exhibited a developmental profile of an initial increase followed by a decrease. The presence of NAA increased the activity of all the enzymes throughout the culture period without altering their developmental profiles. Isozyme profiles of GS and GDH in the callus of C. reflexa were analyzed by PAGE and direct in gel activity staining. In the absence of NAA, the callus exhibited one isozyme of GS and two isozymes of GDH. NAA treatment led to the development of one additional isozyme of GS. Further stimulating effect of NAA on the activity of each of these enzymes was also evident by in gel activity staining of the isozymes. A comparison of the levels of NR, GS, GOGAT and GDH in field vines of C. reflexa, leaves of its host plant, Catheranthus with those of Cuscuta callus, led to the observation that all the nitrogen assimilating enzymes except GDH, were absent in the field vines of C. reflexa. Callus and field vines revealed a preponderance of GDH as compared to GS activity, while a reverse trend was observed in the host plant. The data are suggestive of ammonia assimilation through GDH pathway in this parasite.