MAPK Signaling Pathway May Directly Regulate the Expression of Hydrophobin Genes in Flammulina filiformis

Fungal hydrophobins reduce the surface tension of hyphae so that hyphae can grow into the air. Reduced expression of hydrophobin genes results in abnormal morphogenesis of both hyphae and the fruiting body of Flammulina filiformis. Previous studies showed that filamentous-growth MAPK signaling pathway directly modulates pseudohyphae formation in budding yeast, so we hypothesized that the specific transcription factor in this pathway may also directly regulate the expression of hydrophobin genes in F. filiformis. Downstream of the G protein, the cAMP/PKA signaling pathway is parallel with the filamentous-growth MAPK signaling pathway in regulating the filamentous growth of fungi. Thus, the cAMP addition test was carried out to exclude the involvement of the PKA/cAMP signaling pathway in aerial-hyphae deficiency of the three mutants used in our previous study. Transcriptomic analysis showed common changes in the MAPK signaling pathway of the three mutants, including 6 downregulated and 3 upregulated genes in common. Transcription factor Tec1 was one of the upregulated genes, and it is a pathway-specific transcription factor for filamentous growth. Motif prediction showed that putative binding sites of Tec1 and Ste12 existed in the promoter region of the three chosen hydrophobin genes mentioned in our previous study, and DAP-seq analysis suggested that putative binding sites of Tec1 and Ste12 were located in 10 hydrophobin genes, respectively, and there were 8 in common for both the transcription factors. These results gave suggestive evidence supporting our hypothesis. We have identified a potential regulatory connection between the filamentous-growth MAPK signaling pathway and hydrophobin genes through Tec1 and Ste12. However, functional validation is required to confirm direct regulation between both the transcription factors and the downstream genes.