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植物の全身的な窒素吸収を制御するシグナル伝達経路

大久保, 祐里 名古屋大学

2022.06.03

概要

植物にとって窒素は最も要求量の多い栄養素であり,土壌中のNO3−を主な窒素源として利用している.植物は根の細胞膜に存在するNO3−トランスポーターを介して土壌中のNO3−を体内へと吸収しており,高親和性のNO3−トランスポーターであるNRT2.1がNO3−吸収において主要な役割を果たす.自然界における土壌中のNO3−の分布は,他の植物体による吸収や雨水による流出によって不均一となることが多いため,植物は,一部の根が土壌中のNO3−欠乏を感知すると,NO3−が周囲に高濃度で存在する根でNRT2.1の発現を誘導し,相補的にNO3−吸収量を増大させることで,個体全体として生育に必要な窒素量を維持する仕組みを進化させてきた.この仕組みは全身的な窒素要求シグナル伝達と呼ばれ,一部の根における局所的なNO3−欠乏の情報は地上部の葉から師管を介して他の根へ伝達することが示唆されていたものの,その長距離移行因子の分子実体は長年に渡って未解明のままだった.

その解明の突破口となったのは,ペプチドホルモンのCEPとその受容体のCEPRが,全身的な窒素要求シグナル伝達に関わることを見出した2014年の報告である.NO3−欠乏を感知した根ではペプチドホルモンのCEPが産生され,道管を移行して地上部の師管に存在する受容体CEPRに認識される.これを引き金として,おそらく再び根に向かう未知の2次シグナルが産生され,NRT2.1などの発現量を上昇させ,離れた根でのNO3−吸収を相補的に増大させているものと考えられた.

そこで第1章では,CEP受容体下流2次シグナルを同定するため,葉の維管束でCEP依存的に発現変動する遺伝子群をトランスクリプトーム解析によって探索し,師管を移行し得る非分泌型の比較的低分子のタンパク質を2次シグナル候補とした.この候補遺伝子群について過剰発現株を作製して,根におけるNRT2.1誘導活性を指標としたスクリーニングを実施した結果,新規ポリペプチドCEPDownstream1/2(CEPD1/2)を見出した.CEPD1/2の二重欠損株は葉の黄化など典型的な窒素欠乏症状を示し,不均一なNO3−環境下での相補的なNO3−吸収が行なわれないことから,CEPD1/2が全身的な窒素要求シグナル伝達に必須であることが明らかとなった.CEPD遺伝子は葉の師管で特異的に発現する一方,GFP-CEPD1タンパク質のシグナルは根で明瞭に検出され,根の師管から表皮・皮層細胞の核へと拡散している様子が観察された.また接ぎ木実験によって,葉で産生されたCEPDが師管を通って根へ長距離移行することを証明した.葉で発現したCEPDは周囲のNO3−量に関係なくどの根にも等しく移行するが,周囲にNO3−が存在する根においてのみNRT2.1の発現を誘導しており,土壌中のNO3−量という局所的な情報と,CEP-CEPR-CEPDと長距離伝達された植物体の窒素欠乏の情報が統合され,NRT2.1の発現を制御していると考えられる.このように,第1章では葉から根へ長距離移行して根におけるNO3−の吸収効率を上昇させるシグナル分子を世界で初めて同定した.

CEPDの主な機能は,根におけるNO3−トランスポーターの転写促進であることが第1章およびその後の研究において明らかとなった.しかし,CEPD経路によって発現制御される遺伝子群は他にも数多く存在しており,詳細な解析はこれまで行なわれていない.そこで第2章では,CEPD過剰発現株のトランスクリプトーム解析によって,NRT2.1を含むNO3−トランスポーターと同程度に発現が上昇した機能未知のタンパク質脱リン酸化酵素CEPD-inducedPhosphatase(CEPH)に着目した.CEPHの発現は窒素欠乏時に葉で産生されるCEPD依存的に誘導され,根の主に表皮・皮層細胞の細胞質で発現する.CEPHを欠損した植物は地上部の矮化や葉の黄化といった窒素欠乏症状を示し,根におけるNO3−吸収活性が低下していたことから,CEPHがNO3-吸収活性を翻訳後レベルで活性化する可能性が示された.そこで,CEPHの基質を網羅的に探索するため15N代謝標識を用いた定量リン酸化プロテオミクスを実施したところ,CEPH欠損株では主要なNO3−トランスポーターNRT2.1の501番目のSer残基(Ser501)のリン酸化レベルが増加していた.NRT2.1のSer501がリン酸化を受けるとNRT2.1のNO3-吸収活性が著しく低下したことから,CEPHはNRT2.1のSer501を直接脱リン酸化することでNO3−吸収を活性化していることが明らかとなった.この発見により,これまで知られていなかったNRT2.1の脱リン酸化を介した活性制御機構の存在が示され,窒素要求シグナルCEPDはNO3−トランスポーターNRT2.1の転写活性化だけでなく,脱リン酸化による活性化の両面からNO3−吸収を増大させることが明らかになった.窒素欠乏となった植物はNO3−吸収を促進しようするが、窒素欠乏時はアミノ酸合成ができないためNRT2.1を大量に新規合成することは難しい.そこで,植物は窒素が豊富にあるうちにNRT2.1を余分に合成して不活性型でストックしておき,窒素不足になった時にCEPHを介してNRT2.1を活性化するというシステムを獲得したと考えられる.

本研究で同定した葉からの窒素要求シグナルCEPDと脱リン酸化酵素CEPHは,いずれも単子葉類・双子葉類問わず多くの植物で保存されていることから,植物の窒素吸収において普遍的かつ重要な役割を担う新しい分子群である.これらの研究により,刻々と変動する土壌窒素環境に対する植物の適応メカニズムの全容解明が大きく前進した.

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