The Wnt signaling pathway is crucial for development and disease. The regulation of Wnt protein trafficking is one of the pivotal issues in the Wnt research field. Here we performed a genetic screen in Drosophila melanogaster for genes involved in Wingless/Wnt secretion, and identified the p24 protein family members Baiser, CHOp24, Eclair and a v-SNARE protein Sec22, which are involved in the early secretory pathway of Wingless/Wnt. We provided genetic evidence demonstrating that loss of p24 proteins or Sec22 impedes Wingless (Wg) secretion in Drosophila wing imaginal discs. We found that Baiser cannot replace other p24 proteins (CH0p24 or Eclair) in escorting Wg, and only Baiser and CH0p24 interact with Wg. Moreover, we showed that the v-SNARE protein 5ec22 and Wg are packaged together with p24 proteins. Taken together, our data provide important insights into the early secretory pathway of Wg/Wnt. (C) 2015 Elsevier Inc. All rights reserved.