Institute of Molecular Virology and Cell Biology, Friedrich-Loeffler-Institut, 17493 Greifswald-Insel Riems, Germany.
Department of Genetics, Cell Biology and Development, University of Minnesota, Minneapolis, MN 55455, USA.
Cells. 2020 Mar 17;9(3):738. doi: 10.3390/cells9030738.
Newly assembled herpesvirus nucleocapsids traverse the intact nuclear envelope by a vesicle-mediated nucleo-cytoplasmic transport for final virion maturation in the cytoplasm. For this, they bud at the inner nuclear membrane resulting in primary enveloped particles in the perinuclear space (PNS) followed by fusion of the primary envelope with the outer nuclear membrane (ONM). While the conserved viral nuclear egress complex orchestrates the first steps, effectors of fusion of the primary virion envelope with the ONM are still mostly enigmatic but might include cellular proteins like SUN2 or ESCRT-III components. Here, we analyzed the influence of the only known AAA+ ATPases located in the endoplasmic reticulum and the PNS, the Torsins (Tor), on nuclear egress of the alphaherpesvirus pseudorabies virus. For this overexpression of wild type and mutant proteins as well as CRISPR/Cas9 genome editing was applied. Neither single overexpression nor gene knockout (KO) of TorA or TorB had a significant impact. However, TorA/B double KO cells showed decreased viral titers at early time points of infection and an accumulation of primary virions in the PNS pointing to a delay in capsid release during nuclear egress.
新组装的疱疹病毒核衣壳通过囊泡介导的核质运输穿过完整的核膜，在细胞质中完成最终的病毒成熟。为此，它们在内核膜上出芽，导致核周空间（PNS）中出现初级包膜颗粒，随后初级包膜与外核膜（ONM）融合。虽然保守的病毒核出芽复合物协调了最初的步骤，但初级病毒包膜与 ONM 融合的效应因子仍然大多是神秘的，但可能包括细胞蛋白，如 SUN2 或 ESCRT-III 成分。在这里，我们分析了唯一已知位于内质网和 PNS 中的 AAA+ATP 酶（Torsin，Tor）对α疱疹病毒伪狂犬病病毒核出芽的影响。为此，我们应用了野生型和突变蛋白的过表达以及 CRISPR/Cas9 基因组编辑。TorA 或 TorB 的过表达或基因敲除（KO）都没有显著影响。然而，TorA/B 双 KO 细胞在感染的早期时间点显示出较低的病毒滴度，并且初级病毒颗粒在 PNS 中积累，表明在核出芽过程中衣壳释放延迟。
