阿拉伯芥 NLP1/2 在硝酸鹽與銨鹽反應中的角色

Abstract

硝酸鹽和銨鹽是植物生長發育的氮源與訊息分子。在硝酸鹽和銨鹽所誘發的轉錄反應中，轉錄因子 NLP (NIN-LIKE PROTEIN) 扮演重要角色。最近研究發現序列相近的 NLP6 和 NLP7 在硝酸鹽反應是功能部分重疊的正調控因子，且在銨鹽反應是功能完全重疊的負調控因子。然而，其他 7 個 NLP 蛋白之間的關係尚待釐清。在序列相近的 nlp 雙突變株中，只有 nlp1nlp2 和 nlp6nlp7 在硝酸鹽有生長遲緩，而 nlp4nlp5 和 nlp8nlp9 的生長和野生型相近，顯示 NLP1 與 NLP2 的關係可能和 NLP6 與 NLP7 相似。因此，本研究的目的是探討 NLP1 和 NLP2 在硝酸鹽訊息傳遞和銨鹽訊息傳遞的功能。對於調控生長，NLP2 在低硝酸鹽時的功能比 NLP1 更重要，而 NLP1 和 NLP2 在高硝酸鹽時有些相加 (additive) 的關係；NLP2 和 NLP7 在低硝酸鹽和高硝酸鹽時都有相乘 (synergistic) 的關係，而 NLP2 和 NLP7 在銨鹽時的功能重疊。對於調控基因表現，NLP2 正向調控 HYH、BEE2、ERF60、LBD38、MAPKKK14、ZAT1、UMAMIT36、At1g22170 和 PRS2 的高硝酸鹽誘導轉錄，而 NLP1 對這些基因的貢獻較少或是沒有貢獻。在銨鹽時，nlp2nlp7 的轉錄反應和 nlp6nlp7 雙基因剔除突變株 (double knockout mutant) 相似，顯示 NLP2/7 和 NLP6/7 負向調控一樣的銨鹽反應基因的轉錄。對於細胞內的分布，NLP1 和 NLP2 在硝酸鹽的條件下可能分布於細胞質，而 NLP1 和 NLP2 在銨鹽的條件下都分布於細胞質和細胞核。有趣的是，NLP2/7 和 NLP6/7 在高硝酸鹽負向調控地上部的硝酸鹽濃度，並正向調控地上部硝酸還原酶的活性。總結來說，NLP2 在硝酸鹽反應是正調控因子，而 NLP1 的貢獻比 NLP2 微弱。NLP2 和 NLP7 在銨鹽反應是功能可互相取代的負調控因子，而 NLP1 的功能並不明顯。

Nitrate and ammonium are two major nitrogen sources and signal molecules for plant growth and development. Transcription factors NLPs (NIN-LIKE PROTEINs) play important roles in the transcriptional response which is induced by nitrate and ammonium. Recent studies have showed that NLP6 and NLP7 in the same clade were partially functionally redundant activators in the nitrate response and completely functionally redundant repressors in the ammonium response. However, the relationship of other 7 NLPs remains to be elucidated. Among the nlp double mutants in the same clade, only nlp1nlp2 and nlp6nlp7 displayed growth retardation under nitrate, but the growth of nlp4nlp5 and nlp8nlp9 was similar to the wild type, indicating that the relationship between NLP1 and NLP2 may be similar to NLP6 and NLP7. Therefore, the function of NLP1 and NLP2 is investigated in the nitrate signaling and ammonium signaling in this study. For growth regulation, the function of NLP2 is more important than NLP1 under low nitrate. There are some additive effects between NLP1 and NLP2 under high nitrate. The relationship of NLP2 and NLP7 is synergistic under low nitrate and high nitrate conditions, but is functionally redundant under ammonium. For gene regulation, NLP2 positively regulates the high nitrate-induced transcription of HYH, BEE2, ERF60, LBD38, MAPKKK14, ZAT1, UMAMIT36, At1g22170 and PRS2, but NLP1 contributes little or no to these genes. Under ammonium, the transcriptional response of nlp2nlp7 is similar to nlp6nlp7 double knockout mutant. NLP2/7 and NLP6/7 negatively regulate transcription of the same ammonium-responsive genes. For intracellular localization, NLP1 and NLP2 may localize to the cytosol under nitrate condition, and localize to the cytosol and nucleus under ammonium condition. Interestingly, NLP2/7 and NLP6/7 negatively regulate shoot nitrate concentration and positively regulate shoot nitrate reductase activity under high nitrate. In summary, NLP2 is an activator in the nitrate response, but the contribution of NLP1 is weaker than NLP2. NLP2 and NLP7 are repressors which can functionally replace each other in the ammonium response, but the function of NLP1 is not clear.