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Abstract：
Baculoviruses enter insect midgut epithelial cells via a set of occlusion-derived virion (ODV) envelope proteins called per os infectivity factors (PIFs). P74 of Autographa californica multicapsid nucleopolyhedrovirus (AcMNPV), which was the first identified PIF, is cleaved by an endogenous proteinase embedded within the occlusion body during per os infection, but the target site(s) and function of the cleavage have not yet been ascertained. Here, based on bioinformatics analyses, we report that cleavage was predicted at an arginine and lysine-rich region in the middle of P74. A series of recombinant viruses with site-directed mutants in this region of P74 were generated. R325 or R334 was identified as primary cleavage site. In addition, we showed that P74 is also cleaved by brush border membrane vesicles (BBMV) of the host insect at R325 or R334, instead of R195, R196, and R199, as previously reported. Simultaneous mutations in R195, R196, and R199 lead to instability of P74 during ODV release. Bioassays showed that mutations at both R325 and R334 significantly affected oral infectivity. Taken together, our data show that both R325 and R334 of AcMNPV P74 are the primary cleavage site for both occlusion body endogenous proteinase and BBMV proteinase during ODV release and are critical for oral infection.
OBJECTIVE: Cleavage of viral envelope proteins is usually an important trigger for viral entry into host cells. Baculoviruses are insect-specific viruses that infect host insects via the oral route. P74, a per os infectivity factor of baculoviruses, is cleaved during viral entry. However, the function and precise cleavage sites of P74 remain unknown. In this study, we found that R325 or R334 between the N- and C-conserved domains of P74 was the primary cleavage site by proteinase either from the occlusion body or host midgut. The biological significance of cleavage seems to be the release of the potential fusion peptide at the N-terminus of the cleaved C-terminal P74. Our results shed light on the cleavage model of P74 and imply its role in membrane fusion in baculovirus per os infection.
OBJECTIVE: Cleavage of viral envelope proteins is usually an important trigger for viral entry into host cells. Baculoviruses are insect-specific viruses that infect host insects via the oral route. P74, a per os infectivity factor of baculoviruses, is cleaved during viral entry. However, the function and precise cleavage sites of P74 remain unknown. In this study, we found that R325 or R334 between the N- and C-conserved domains of P74 was the primary cleavage site by proteinase either from the occlusion body or host midgut. The biological significance of cleavage seems to be the release of the potential fusion peptide at the N-terminus of the cleaved C-terminal P74. Our results shed light on the cleavage model of P74 and imply its role in membrane fusion in baculovirus per os infection.
摘要：
杆状病毒通过一组闭塞衍生的病毒粒子（ODV）包膜蛋白进入昆虫中肠上皮细胞，这些包膜蛋白称为peros感染因子（PIF）。Californica多衣壳核型多角体病毒（AcMNPV）的P74，这是第一个确定的PIF，在每个操作系统感染期间被嵌入闭塞体内的内源性蛋白酶切割，但切割的靶位点和功能尚未确定。这里,基于生物信息学分析，我们报道了在P74中部的精氨酸和赖氨酸丰富区域预测切割。产生了一系列在P74的该区域中具有定点突变体的重组病毒。R325或R334被鉴定为主要切割位点。此外,我们表明，P74也被宿主昆虫的刷状缘膜囊泡（BBMV）在R325或R334处裂解，而不是先前报道的R195，R196和R199。R195、R196和R199中的同时突变导致在ODV释放期间P74的不稳定性。生物测定显示R325和R334处的突变显著影响口腔感染性。一起来看,我们的数据显示,AcMNPVP74的R325和R334都是ODV释放过程中闭塞体内源性蛋白酶和BBMV蛋白酶的主要切割位点,对口腔感染至关重要.
目的：病毒包膜蛋白的裂解通常是病毒进入宿主细胞的重要触发因素。杆状病毒是通过口服途径感染宿主昆虫的昆虫特异性病毒。P74，一种杆状病毒的个人感染因子，在病毒进入过程中被切割。然而,P74的功能和精确的切割位点仍然未知。在这项研究中,我们发现P74的N-和C-保守域之间的R325或R334是来自闭塞体或宿主中肠的蛋白酶的主要切割位点。切割的生物学意义似乎是在切割的C末端P74的N末端释放潜在的融合肽。我们的结果揭示了P74的裂解模型，并暗示了其在杆状病毒每次感染中的膜融合中的作用。
目的：病毒包膜蛋白的裂解通常是病毒进入宿主细胞的重要触发因素。杆状病毒是通过口服途径感染宿主昆虫的昆虫特异性病毒。P74，一种杆状病毒的个人感染因子，在病毒进入过程中被切割。然而,P74的功能和精确的切割位点仍然未知。在这项研究中,我们发现P74的N-和C-保守域之间的R325或R334是来自闭塞体或宿主中肠的蛋白酶的主要切割位点。切割的生物学意义似乎是在切割的C末端P74的N末端释放潜在的融合肽。我们的结果揭示了P74的裂解模型，并暗示了其在杆状病毒每次感染中的膜融合中的作用。
