Multiple cellular and molecular biological technologies have been used to detect virus infection, replication and variation of intracellular proteins, including western blotting, qRT-PCR, Gene silencing, immunofluorescence, etc. Besides, microarray gene chip technology was applied to analyze the effects of SAMHD1 overexpression on total expressed genes.
Our data show that SAMHD1 down-regulates the expression of genes related to lipid bio-metabolic pathway, accompanied with impaired lipid droplets (LDs) formation, two events important for flaviviruses infection. Mechanic study reveals that SAMHD1 mainly targets on HCV RNA replication, resulting in a broad inhibitory effect on the infectivity of flaviviruses. The C-terminal domain of SAMHD1 is showed to determine its antiviral function, which is regulated by the phosphorylation of T592. Restored lipid level by overexpression of SREBP1 or supplement with LDs counteracts with the antiviral activity of SAMHD1, providing evidence supporting the role of SAMHD1-mediated down-regulation of lipid synthesis in its function to inhibit viral infection.
SAMHD1 plays an important role in IFN-mediated blockade of flaviviruses infection through targeting lipid bio-metabolic pathway.
UNASSIGNED:多种细胞和分子生物学技术已用于检测病毒感染,细胞内蛋白质的复制和变异,包括西方印迹,qRT-PCR,基因沉默,免疫荧光,等。此外,应用微阵列基因芯片技术分析SAMHD1过表达对总表达基因的影响。
UNASSIGNED:我们的数据表明,SAMHD1下调与脂质生物代谢途径相关的基因的表达,伴有脂滴(LDs)形成受损,两个事件对黄病毒感染很重要。机制研究表明,SAMHD1主要针对HCVRNA复制,对黄病毒的感染性产生广泛的抑制作用。SAMHD1的C末端结构域显示确定其抗病毒功能,它受T592的磷酸化调节。通过过表达SREBP1或补充LD来恢复脂质水平抵消SAMHD1的抗病毒活性,提供了支持SAMHD1介导的脂质合成下调在其抑制病毒感染功能中的作用的证据。
UNASSIGNED:SAMHD1通过靶向脂质生物代谢途径在IFN介导的黄病毒感染阻断中起重要作用。