No abstract

Induction of nitrate reductase activity and mRNA by nitrate and light is prevented if chloroplasts are destroyed by photooxidation in norflurazon-treated squash (Cucurbita maxima L.) cotyledons. The enzyme activity and mRNA can be induced if norflurazontreated squash seedlings are kept in low-intensity red light, which minimizes photodamage to the plastids. It is concluded that induction of nitrate reductase activity and nitrate reductase mRNA requires intact plastids. If squash seedlings grown in low-intensity red light are transferred to photooxidative white light, nitrate reductase activity accumulates during the first 12 hours after the shift and declines thereafter. Thus photodamage to the plastids and the disappearance of nitrate reductase activity and mRNA are events separable in time, and disappearance of the enzyme activity is a consequence of the damage to the plastids.

We compared the response of NO 3 (-) -induced nitrate-reductase (NR) and nitrite-reductase (NIR) levels in virtually carotenoid-free far-red-light-grown mustard (Sinapis alba L.) cotyledons following a photooxidative treatment of the plastids. The cytosolic localization of NR and the plastidic localization of NIR were confirmed with this approach. Emphasis was on a plastidic factor previously postulated to be involved obligatorily in the transcriptional control of nuclear genes coding for proteins destined for the chloroplast. Photooxidative damage of the plastid would be to destroy the ability of the organelle to send off this signal. Dependency of NIR and NR induction by NO 3 (-) on the plastidic factor is described in detail, and it is concluded that requirement for the plastidic factor is relatively high in the case of NR while factor requirement to allow induction is low in the case of NIR. The data indicate that in the case of NIR the photooxidative damage done to the plastid also affects accumulation of the enzyme directly. Since this effect is absent in the case of cytosolic NR, induction of NR is a particularly suitable system for further molecular studies of the plastidic factor and its mode of action.

We have measured levels of ribulose-1,5-bisphosphate carboxylase (RuBPCase) and levels of in-vitro-translatable mRNA for the small subunit (SSU) of RuBPCase up to 96 h after sowing in mustard (Sinapis alba L.) cotyledons, in order to investigate to what extent the rate of enzyme synthesis is related to the level of SSU-mRNA. Both enzyme and mRNA level are controlled strongly by phytochrome, but the rate of RuBPCase accumulation was found to be unrelated to the level of translatable SSU-mRNA. As an example, it was found that the amount of SSU-mRNA in far-red light (FR)-grown mustard seedlings doubles between 54 and 84 h after sowing while the rate of RuBPCase accumulation remains constant over this period. Since the holoenzyme shows zero turnover during this period it is concluded that the rate of enzyme synthesis remains constant although the level of SSU-mRNA increases strongly. Following an FR→dark transition, with different levels of physiologically active phytochrome (Pfr) established at the end of the light period, no correlation was found between the time course of mRNA levels in darkness and the rate of enzyme synthesis. Rather, the data indicate that there is at least one translational or post-translational regulatory step which is also phytochrome-dependent. It is concluded that coarse control of the appearance of translatable SSU-mRNA is essential for RuBPCase to appear at a high rate but that fine tuning by phytochrome of the actual appearance of RuBPCase is not transcriptional.