Sln, a conserved sensor kinase, plays a distinct function in different fungi. The gene AaSln1 in Alternaria alternata was successfully cloned and characterized. Sequence analysis revealed that AaSln1 consists of a transmembrane helix region, two characteristic low complexity regions, and a HisKA, HATPase_c and sesponse reg domain, respectively. Through gene knockout strategy, the results showed that AaSln1 significantly regulated infection structural differentiation, pathogenicity, stress response and melanin synthesis and secretion, while had no effect on growth, sporulation and biomass accumulation of A. alternata. The AaSln1 regulates pathways in A. alternata including ABC transporters, starch and sucrose metabolism according to the transcriptomic analysis. RNA-Seq results revealed that the gene expression levels of the downstream regulators of Hog1-MAPK pathway such as Mss4, Ste12, Fsk2 and Mip1 were significantly reduced in the ΔAaSln1 mutant. These findings demonstrated that AaSln1 has an essential role in the pathogenicity and regulates downstream biological processes of A. alternata by activating the MAPK pathway.