Pruksarojanakul Patthasarun, Atsumi Kazune, Oh Youngjun, Matoba Nobuyuki, Matsuda Ryo
Department of Biological and Environmental Engineering, Graduate School of Agricultural and Life Sciences, The University of Tokyo, Tokyo, Japan.
Department of Pharmacology and Toxicology, University of Louisville School of Medicine, Louisville, Kentucky, USA.
Biotechnol Bioeng. 2025 May;122(5):1142-1152. doi: 10.1002/bit.28942. Epub 2025 Jan 31.
Postinfiltration air temperature is known to affect the accumulation of recombinant protein in Agrobacterium-mediated transient expression in Nicotiana benthamiana plants, including the number of days needed to reach maximum content and the rate of reduction thereafter. This study aimed to clarify whether the transcript levels of the transgenes and those of plant stress response markers (i.e., hypersensitive response and endoplasmic reticulum [ER] stress) could be primary determinants of the accumulation of recombinant influenza hemagglutinin (HA) at 21 or 26°C. We found no correlation between the transgene expression levels (HA, RdRP, and MP) and the number of days needed to reach the maximum HA protein content at both temperatures. Regardless of the accumulation compartment, HA protein content peaked earlier at 26°C than at 21°C. The rapid reduction of HA content after reaching the maximum, observed only in the ER at 26°C, correlated with severe necrosis and high transcript levels of two representative ER stress markers, bZIP60 and BiP. This correlation suggests that high postinfiltration air temperature affects HA accumulation primarily through ER stress, a key factor in the rapid reduction of HA content after the peak.
已知浸润后空气温度会影响本氏烟草植株中农杆菌介导的瞬时表达过程中重组蛋白的积累，包括达到最大含量所需的天数以及此后的减少速率。本研究旨在阐明转基因的转录水平以及植物应激反应标记物（即过敏反应和内质网[ER]应激）的转录水平是否可能是21或26°C下重组流感血凝素（HA）积累的主要决定因素。我们发现在这两个温度下，转基因表达水平（HA、RdRP和MP）与达到最大HA蛋白含量所需的天数之间没有相关性。无论积累区室如何，HA蛋白含量在26°C时比在21°C时更早达到峰值。仅在26°C的内质网中观察到达到最大值后HA含量的快速下降，这与两种代表性ER应激标记物bZIP60和BiP的严重坏死和高转录水平相关。这种相关性表明，浸润后空气温度较高主要通过ER应激影响HA积累，ER应激是峰值后HA含量快速下降的关键因素。
