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単純ヘルペスウイルス1型の小胞媒介性核外輸送に関する研究

竹島, 功高 東京大学 DOI:10.15083/0002004982

2022.06.22

概要

単純ヘルペスウイルス1型(HSV-1)感染細胞の核内で形成されたヌクレオカプシドは、小胞媒介性核外輸送と呼ばれる生物学的にとてもユニークな輸送機構によって、核内から核外へ輸送される。この輸送過程では、まず、核内のヌクレオカプシドが核内膜から出芽することで、核膜間に核内膜をエンベロープとして獲得したウイルス粒子が形成される。その後、核膜間のウイルス粒子が核外膜と融合することによって、ウイルス粒子中のヌクレオカプシドが核外へと放出される。HSV-1因子UL31及び、UL34から構成される複合体であるNuclear Egress Complex(NEC)は、小胞媒介性核外輸送において必須の因子であると考えられており、核内膜の膜変性、核膜間粒子へのヌクレオカプシドの取り込み及び、核膜間粒子形成時の核内膜切断など多岐にわたり小胞媒介性核外輸送に関与していると考えられている。HSV-1NECとヌクレオカプシドの結合は、核内で形成されたヌクレオカプシドの出芽と核膜間粒子へのヌクレオカプシド取り込みに関与しているとされるが、この仮説を直接的に証明する実験結果は未だ得られていない。本研究では、NECと核内で形成されたヌクレオカプシドとの結合が核膜間ウイルス粒子形成過程において、どのような役割があるのか解明するために、試験管内でHSV-1NECとヌクレオカプシド及び、カプシドタンパク質との結合を検証できる実験系の構築を行った。

構築した実験系を用いた結果、HSV-1NECは、ヌクレオカプシド及び、特定のカプシドタンパク質(UL25)と結合していた。また、UL31の281番目のアルギニン(UL31R281)及び、282番目のアスパラギン酸が(UL31D282)がNECとヌクレオカプシド及び、UL25との効率的な結合に必要であることが明らかになった。さらに、HSV-1感染細胞におけるHSV-1NECとヌクレオカプシドとの結合の意義を調べるためにHSV-1UL31R281及び、UL31D282をアラニンに置換した変異ウイルスを作製した。HSV-1UL31R281及び、UL31D282に変異を導入したウイルスは、感染細胞におけるウイルス増殖の割合が野生体と比較して低下していた。さらに、HSV-1UL31R281及び、UL31D282に変異を導入したウイルス感染細胞では、野生体と比較して核内にカプシドが蓄積しており、核膜間にヌクレオカプシドを含まない空の粒子が大量に観察された。この空の粒子の大きさは、野生体感染細胞内で通常観察される核膜間ウイルス粒子とほとんど同じであった。これらの結果から、HSV1NECのUL31R281及び、UL31R282を介した、NECとヌクレオカプシドに含まれるUL25との結合は、ヌクレオカプシドの核膜間粒子への効率的な取り込みを促進し、ウイルス増殖に寄与していることが示唆された。

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