show Abstracthide AbstractHigh temperature tends to be a contributing factor to the spread of vector-born viral disease via enhancing vector competence. However, the underlying mechanisms of increased virus transmission capacity of vectors under heat stress are still largely unknown. Tomato yellow leaf curl virus (TYLCV) is one of the most devastating plant DNA viruses globally and is exclusively transmitted by Bemisia tabaci. Here we identify a transcription factor of whitefly, heat shock factor (HSF), as a key factor to mediate TYLCV accumulation in their vectors under heat stress. Quantitative analysis show that the amount of virion-sense DNA in whiteflies is gradually increased with temperature rising, but complementary-sense DNA abundance is stable under different temperature treatment. Meanwhile, whitefly HSF is identified to interact with the intergenic region of TYLCV by yeast one-hybrid. Dual luciferase analysis further proves the specificity of this interaction. In addition, the variation trend of HSF transcriptional activity coincides with TYLCV accumulation in whiteflies with the increase of temperature. Knocking down HSF in whiteflies cause a significantly decrease of TYLCV DNA and coat protein. Our results reveal an important role of HSF in mediating the virus accumulation in their insect vectors under high temperature and provide insights into how environmental factors affect plant virus-vector interaction.