ABSTRACT: We used iTRAQ technology to investigate the protein expression characteristics of S-RNases in S-gene homozygous germplasm. By using MaxQuant to search the raw mass spectrometry data files against the protein database, a total of 30,503 identified spectra, 13,568 peptides, and 4,472 proteins were identified in the styles of Huanghua (S1S2) and its homozygous S-genotype offspring (S1S1 and S2S2). These identified proteins were predicted to be located in extracellular regions, cytoplasm, nucleus, mitochondria, Golgi apparatus, endoplasmic reticulum, peroxisomes, vacuoles, nucleoplasm, and other cellular compartments. The number of DEPs among the three varieties was calculated. According to the overall distribution of these proteins, the highest number of DEPs was between S2S2 and S1S2, while the lowest number of DEPs was between S2S2 and S1S1. This indicated that the protein expression patterns of S2S2 and S1S1 were more similar to each other, while both offspring samples showed significant differences in protein expression when compared with the parent S1S2. In this study, we conducted DEP analyses focusing on S-RNases and the proteins that interact with them in the styles of the three pear varieties. Figure 6 shows the heat-map of the expression levels of the 13 identified S-RNase-related proteins, which reflects their differences in expression levels across the three samples. In terms of the overall expression trend, using the offspring sample S1S1 as a normal expression standard, most of the S-RNase-related proteins were down-regulated in the parent S1S2. In contrast, in the offspring sample S2S2, most of the S-RNase-related proteins were up-regulated relative to their levels in the other offspring sample S1S1, with only a few exceptions. Compared with the parent S1S2, both offspring samples, S1S1 and S2S2, showed up-regulated expression of 12 out of the 13 S-RNase-related proteins, except for S1-RNase. Comparing S1S1 and S2S2, the expression levels of F-box/WD-repeat_TBL1XR1, SKP1-like_1A_2, and S2-RNase were higher in S1S1 than in S2S2, while the expression levels of the other 10 S-RNase-related proteins were higher in S2S2 than in S1S1.