Induction of nitrate reductase activity and mRNA accumulation in rice

H. B. Hamat

Abstract

Western blot analysis using barley NADH-nitrate reductase antisera revealed that nitrate-induced rice seedling leaf extracts contain a cross reacting polypeptide of 115 000 dalton. Nitrate-induced rice seedling leaf extracts appeared to contain both NADH- and NAD(P)H-nitrate reductase activity. The NADH-nitrate reductase activity was detected 2 h after the addition of nitrate and increased rapidly to 24 h. The activity continued to increase at a slower rate until about 48 h and declined there after. The results on the nitrate content in leaf showed a similar pattern to the NADH-nitrate reductase activity. The NAD(P)H-nitrate reductase activity was detected within t h, after induction with nitrate which increased rapidly reaching a maximum activity within 3 h. Both pHBH1, and pHBH2 clones (accompanying paper) hybridized to a 3.2 kb mRNA, but the appearance of the mRNA following nitrate induction was different' Induction studies using unique sequences of each clone as the probe revealed that clone pHBHI hybridized to a3.2 kb mRNA. Upon induction by nitrate, it showed kinetics similar to NADH-nitrate reductase activity. The 3.2 kb mRNA was detected after 1 h and peaked at 6 h after induction and declined from 12 h to 24 h. The NADH-nitrate reductase activity increased rapidly during the first 4 h and peaked at 48 h after nitrate induction. Clone pHBH2 hybridized to a 3.2 kb mRNA. Upon induction by nitrate, it showed kinetics similar to NAD(P)H-nitrate reductase activity. The 3.2 kb mRNA was detected very early, reached maximum 30 min after nitrate induction' and declined very rapidly. No 3.2 kb mRNA was detected from 6 h to 24 h after induction. The NAD(P)H-nitrate reductase activity was inducible by nitrate. The activity increased very rapidly reaching maximum within 3 h. declined after 5 h, and very little from 10 h to 72 h. These results suggested that clone pHBHI codes for NADH-nitrate reductase while clone pHBH2 codes for NAD(P)H-nitrate reductase. The results also indicated that the regulation at transcriptional levels is different between the NADH and NAD(P)H-nitrate reductase.

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