Kyung-Tae Lee^1,6, Joohyeon Hong^1,6, Dong-Gi Lee^1,6, Minjae Lee¹, Suyeon Cha¹, Yu-Gyeong Lim¹, Kwang-Woo Jung², Areum Hwangbo¹, Yelin Lee¹, Shang-Jie Yu^3,5, Ying-Lien Chen³, Jong-Seung Lee⁴, Eunji Cheong^1,* & Yong-Sun Bahn^1,*

¹Department of Biotechnology, College of Life Science and Biotechnology, Yonsei University, Seoul 03722, Korea. ²Research Division for Biotechnology, Korea Atomic Energy Research Institute, Jeongeup 56212, Korea. ³Department of Plant Pathology and Microbiology, National Taiwan University, Taipei 10617, Taiwan. ⁴AmtixBio Co., Ltd., Seoul 05836, Korea. ⁵Present address: National Institute of Infectious Diseases and Vaccinology, National Health Research Institute, 35053 Miaoli, Taiwan. ⁶These authors contributed equally: Kyung-Tae Lee, Joohyeon Hong, Dong-Gi Lee.

^*Corresponding authors

Cryptococcus neoformans causes fatal fungal meningoencephalitis. Here, we study the roles played by fungal kinases and transcription factors (TFs) in blood-brain barrier (BBB) crossing and brain infection in mice. We use a brain infectivity assay to screen signature-tagged mutagenesis (STM)-based libraries of mutants defective in kinases and TFs, generated in the C. neoformans H99 strain. We also monitor in vivo transcription profiles of kinases and TFs during host infection using NanoString technology. These analyses identify signalling components involved in BBB adhesion and crossing, or survival in the brain parenchyma. The TFs Pdr802, Hob1, and Sre1 are required for infection under all the conditions tested here. Hob1 controls the expression of several factors involved in brain infection, including inositol transporters, a metalloprotease, PDR802, and SRE1. However, Hob1 is dispensable for most cellular functions in Cryptococcus deuterogattii R265, a strain that does not target the brain during infection. Our results indicate that Hob1 is a master regulator of brain infectivity in C. neoformans.