Institute of Molecular Biophysics, Florida State University, Tallahassee, Florida, USA.
Department of Biological Science, Florida State University, Tallahassee, Florida, USA.
J Biol Chem. 2023 Mar;299(3):102986. doi: 10.1016/j.jbc.2023.102986. Epub 2023 Feb 7.
Dengue virus (DENV) is one of the most prevalent mosquito-transmitted human viruses that causes significant morbidity and mortality worldwide. To persist in the cell and consequently cause disease, DENV is evolved with mechanisms to suppress the induction of type I interferons by antagonizing cGAS-STING signaling. Using recombinant proteins and in vitro cleavage assays, we have shown that the DENV protease NS2B3 is capable of cleaving cGAS in the N-terminal region without disrupting the C-terminal catalytic center. This generates two major cleavage products: cleavage product N-terminal (CP-N) and cleavage product C-terminal (CP-C). We observed reduction in DNA-binding affinity of CP-C as compared to full-length cGAS. Reduction in DNA-binding affinity is also correlated with the decrease in enzymatic activity of CP-C. CP-N, on the other hand, has almost comparable DNA-binding ability as that of the full-length cGAS. In fact, CP-N competitively inhibits cyclic GMP-AMP production by both full-length cGAS and CP-C. We hypothesize that high DNA-binding affinity of CP-N enables it to sequester the DNA from CP-C and noncleaved full-length cGAS and thus reduces the rate of enzyme activation and cyclic GMP-AMP synthesis. Furthermore, we found that NS2B3 physically interacts with full-length cGAS and CP-C, laying the basis for their shuttling to and eventual degradation in the autophagosome. Overall, our study highlights a multifaceted and effective strategy by which an RNA virus antagonizes cGAS-STING signaling which may be useful for the design of antivirals targeting viral proteases.
登革热病毒（DENV）是一种最常见的通过蚊子传播的人类病毒，它在全球范围内造成了大量的发病率和死亡率。为了在细胞中持续存在并因此引起疾病，DENV 进化出了抑制 I 型干扰素诱导的机制，通过拮抗 cGAS-STING 信号。我们使用重组蛋白和体外切割实验表明，DENV 蛋白酶 NS2B3 能够在不破坏 C 末端催化中心的情况下，在 N 末端区域切割 cGAS。这产生了两个主要的切割产物：切割产物 N 末端（CP-N）和切割产物 C 末端（CP-C）。与全长 cGAS 相比，我们观察到 CP-C 的 DNA 结合亲和力降低。DNA 结合亲和力的降低也与 CP-C 的酶活性降低相关。另一方面，CP-N 的 DNA 结合能力几乎与全长 cGAS 相当。事实上，CP-N 竞争性地抑制全长 cGAS 和 CP-C 产生的环鸟苷酸-腺苷酸。我们假设 CP-N 的高 DNA 结合亲和力使其能够将 DNA 与 CP-C 和未切割的全长 cGAS 隔离，从而降低酶的激活和环鸟苷酸-腺苷酸合成的速度。此外，我们发现 NS2B3 与全长 cGAS 和 CP-C 发生物理相互作用，为它们在自噬体中的转运和最终降解奠定了基础。总的来说，我们的研究强调了一种多方面的、有效的策略，即 RNA 病毒拮抗 cGAS-STING 信号，这可能有助于设计针对病毒蛋白酶的抗病毒药物。
