Constitutive and inducible aspects of nitrate‐nitrogen uptake by Chlamydomonas reinhardtii

Abstract

Similarly to higher plant root systems, Chlamydomonas reinhardtii Dangeard (UTEX 90) cells exhibited biphasic NO3− uptake kinetics. The uptake pattern was similar in cells cultured in 10 mM NO3− (NO3−‐grown), 0.25 mM NO3− (N‐limited) or 10 mM NO3− followed by an 18‐h period of N‐deprivation (N‐starved). In all cell types there was an apparent phase transition in uptake at 1.1 mM NO3−, although there were variations in the uptake Vmax of both isotherms. The rate of uptake via isotherm 0 ([NO3−]<1.1 mM) in N‐limited cells was higher than that of either NO3−‐grown or N‐starved cells. In contrast, NO3−‐grown and N‐limited cells exhibited comparable Vmax values when supplied with 1.1 to 1.8 mM NO3− (isotherm 1). When supplied with 1.6 mM NO3−, both N‐limited and N‐starved cells exhibited enhanced linear uptake after 60 min of incubation. We ascribed this to an induction phenomenon. This trend was not observed when NO3−‐grown cells were supplied with 1.6 mM NO3−, or when N‐limited and N‐starved cells were supplied with 0.6 mM NO3−. The ‘inducible’ aspect of uptake by N‐limited cells was blocked by cycloheximide (10 mg l−1), but not by actinomycin D (5 mg l−1), thus indicating the involvement of a translational or post‐translational event. To investigate this phenomenon further, we analysed the cell proteins of N‐limited cells supplied with either 0.6 or 1.6 mM NO3− for 90 min, using two‐dimensional gel electrophoresis. Comparison of protein profiles enabled the identification of a single cell membrane‐associated polypeptide (21 kDa, pI ca 5.5) and ten soluble fraction polypeptides (17–73 kDa, pI ca 5.0 to 7.1) unique to the high NO3− treatment. We propose that the ‘inducible’ portion of NO3− uptake may provide the means by which C. reinhardtii cells regulate uptake in accordance with assimilatory capacity.