Study on the regulation of broad-spectrum re-sistance is an active area in plant biology. RE-SISTANCE TO POWDERY MILDEW 8.1 (RPW8.1) is one of a few broad-spectrum resistance genes triggering the hypersensitive response (HR) to re-strict multiple pathogenic infections. To address the question how RPW8.1 signaling is regulated, we performed a genetic screen and tried to iden-tify mutations enhancing RPW8.1-mediated HR. Here, we provided evidence to connect an annexin protein with RPW8.1-mediated resistance in Arabidopsis against powdery mildew. We iso-lated and characterized Arabidopsis b7-6 mutant. A point mutation in b7-6 at the At5g12380 locus resulted in an amino acid substitution in ANNEXIN 8 (AtANN8). Loss-of-function or RNA-silencing of AtANN8 led to enhanced expression of RPW8.1, RPW8.1-dependent necrotic lesions in leaves, and defense against powdery mildew. Conversely, over-expression of AtANN8 compromised RPW8.1-mediated disease resistance and cell death. Interestingly, the mutation in AtANN8 en-hanced RPW8.1-triggered H2O2. In addition, mu-tation in AtANN8 led to hypersensitivity to salt stress. Together, our data indicate that AtANN8 is involved in multiple stress signaling pathways and negatively regulates RPW8.1-mediated resistance against powdery mildew and cell death, thus linking ANNEXIN's function with plant immunity.