Transcriptional regulation of the Nia1 gene encoding nitrate reductase in Chlamydomonas reinhardtii: effects of various environmental factors on the expression of a reporter gene under the control of the Nia1 promoter

The NAD(P)H nitrate reductase (NR) from Chlamydomonas reinhardtii is encode d by the structural gene Nia1. Numerous data from the literature indicate t hat this enzyme is submitted to complex regulation mechanisms involving mul tiple controls at transcriptional and post-transcriptional levels. To speci fically investigate the regulation of the Nia1 gene at the transcriptional level, NR+ and NR- transformed cells harbouring the Nia1:Ars construct (Nia 1 promoter fused to the arylsulfatase (ARS)-encoding Ars reporter gene) wer e cultivated under various experimental conditions and the ARS activities w ere recorded. ARS levels were very low in cells grown in the presence of NH 4Cl and dramatically increased on agar medium deprived of any nitrogen sour ce or containing nitrate, nitrite, urea, arginine or glutamine. Compared to nitrogen-free medium, a slight positive effect of nitrate in the NR+ strai n and a significant negative effect of nitrite in both NR+ and NR- strains were observed. The ARS activities were high in the light and very low in th e dark or in the light in the presence of DCMU, indicating that Nia1 transc ription is strikingly dependent on photosynthetic activity. Acetate used as a carbon source in the dark did not substitute for light in stimulating Ni a1:Ars expression. Inactivation of NR by tungstate treatment of the NR+ str ain resulted in a dramatic increase of ARS level suggesting that in Chlamyd omonas, like in higher plants, active NR negatively regulates the transcrip tion of the NR structural gene. Deleting the major part of the Nia1 leader sequence still present in the chimeric gene resulted in a decrease of ARS l evel but did not modify the regulation pattern.