Regulation of synthesis of nitrite reductase in pea leaves: in-vivo and in-vitro studies

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Abstract
Crude protein extracts from leaves of pea (Pisum sativum L.) labeled with an L-14C-amino-acid mixture or [35S]methionine, were treated with antibodies prepared against nitrite reductase (NiR; EC 1.6.6.4). When the immunoprecipitates were subjected to sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE) a polypeptide with the same mobility as that of the native NiR was detected. By using darkened or illuminated leaves in the absence or presence of nitrate, it has been confirmed that nitrate is required in the in-vivo synthesis of NiR and that this synthesis is stimulated by light. Cell-free translation with a wheat-germ extract primed with polysomes from illuminated leaves treated with nitrate yielded polypeptides of a wide range of molecular weights (Mrs). Two polypeptides were immunoprecipitated from the translational products by anti-NiR serum. The mobility of one of them on SDS-PAGE corresponded to that of NiR while the other had a slightly higher Mr. It is concluded that NiR is synthesized as a heavy-molecular-weight precursor. Nitrate appears to regulate NiR synthesis by triggering transcription whereas the light may control the level of transcription or translation.