| Summary: | F-box protein is a key protein of the SCF E3 ubiquitin ligase complex, responsible for substrate recognition and degradation through specific interactions. Previous studies have shown that F-box proteins play crucial roles in <i>Cryptococcus</i> sexual reproduction. However, the molecular mechanism by which F-box proteins regulate sexual reproduction in <i>C. neoformans</i> is unclear. In the study, we discovered the AICAR transformylase/IMP cyclohydrolase Ade16 as a substrate of Fbp1. Through protein interaction and stability experiments, we demonstrated that Ade16 is a substrate for Fbp1. To examine the role of <i>ADE16</i> in <i>C. neoformans</i>, we constructed the i<i>ADE16</i> strains and <i>ADE16</i><sup>OE</sup> strains to analyze the function of Ade16. Our results revealed that the i<i>ADE16</i> strains had a smaller capsule and showed growth defects under NaCl, while the <i>ADE16</i><sup>OE</sup> strains were sensitive to SDS but not to Congo red, which is consistent with the stress phenotype of the <i>fbp1</i>Δ strains, indicating that the intracellular protein expression level after <i>ADE16</i> overexpression was similar to that after <i>FBP1</i> deletion. Interestingly, although i<i>ADE16</i> strains can produce basidiospores normally, <i>ADE16</i><sup>OE</sup> strains can produce mating mycelia but not basidiospores after mating, which is consistent with the <i>fbp1</i>Δmutant strains, suggesting that Fbp1 is likely to regulate the sexual reproduction of <i>C. neoformans</i> through the modulation of Ade16. A fungal nuclei development assay showed that the nuclei of the <i>ADE16</i><sup>OE</sup> strains failed to fuse in the bilateral mating, indicating that Ade16 plays a crucial role in the regulation of meiosis during mating. In summary, our findings have revealed a new determinant factor involved in fungal development related to the post-translational regulation of AICAR transformylase/IMP cyclohydrolase.
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