Elucidating rotaviral condensate structure and function for future antiviral targeting

几乎所有已知的人类病毒都在称为病毒质的细胞中的特殊隔间内复制。这些结构的相分离性质使它们有望成为轮状病毒的抗病毒靶标；将高通量微流体学方法与生物物理学和分子病毒学相结合，将为研究它们的行为提供独特的见解。例如，与凝析油形成相关的NSP5在整个感染过程中都会磷酸化。微流体学将使研究磷酸化对相边界的影响成为可能；除了体内研究，这可能揭示为什么病毒体随着时间的推移而固化。病毒质相关RNA在相分离和颗粒组装中的作用也尚不清楚。虽然这些分子的无序状态使结构预测具有挑战性，但反向遗传学可以用来拯救病毒及其RNA。通过检测液滴内差异FRET的发射并与已知结构进行比较，可以识别不同条件下病毒RNA的结构变化。此外，高通量筛选可以识别特定聚集并破坏病毒凝聚物的潜在抗病毒药物。更好地理解病毒相分离，以及我将从这个项目中获得的技能，然后可以应用于其他感染系统。在面对由抗病毒和抗菌素耐药性造成的全球危机时，这一点尤为重要。

Nearly all known human viruses replicate within specialised compartments in cells called viroplasms. The phase-separating nature of these structures makes them promising antiviral targets in rotaviruses; combining high-throughputmicrofluidics approaches with biophysics and molecular virology will offer unique insights into their behaviour. For example, NSP5 associated with condensate formation phosphorylates throughout infection. Microfluidics would allowinvestigations into the effects of phosphorylation on phase boundaries; alongside in vivo studies, this could reveal why viroplasms solidify over time. The role of viroplasm-associated RNA in phase separation and particle assembly alsoremains unclear. While the disordered state of these molecules makes structure prediction challenging, reverse genetics can be used to rescue viruses and their RNA. By detecting differential FRET emissions inside droplets and comparingwith known structures, structural changes in viral RNA under various conditions could be identified. Additionally high-throughput screening can identify potential antivirals that specifically aggregate into and disrupt viral condensates. Abetter understanding of viral phase separation, along with the skillset I will have acquired from this project, could then be applied to other infectious systems. This is of particular importance when facing of a global crisis caused by antiviral and antimicrobial resistance.