Cell migration is a fundamental and precisely regulated event during animal development, immune response and wound healing. It can also lead to detrimental outcomes such as tumor metastasis and atherosclerosis. Hence, unraveling the detailed mechanism of this process is crucial to advance our understanding of development and to improve therapeutics. A cluster of cells termed the border cells in Drosophila egg chambers provides a well-founded and genetically tractable system to study cell migration in vivo. The border cell cluster, originating from the follicular epithelium, is composed of four to six migratory border cells and a pair of non-motile polar cells. Polar cells induce the specification of border cells by secreting the Unpaired (Upd) ligand, which activates the well-conserved Janus Kinase/Signal Transducer and Activator of Transcription (JAK/STAT) signaling pathway in the neighboring follicle cells. Border cells then wrap around polar cells and migrate between nurse cells toward the developing oocyte. In an RNAi screen to identify novel regulators of cell migration, we uncovered a requirement for α- Soluble NSF Attachment Protein (α-SNAP) in border cell cluster specification. α-SNAP is known to function in synaptic transmission, membrane fusion, and vesicle trafficking by facilitating association of N-ethylmalemide-Sensitive Factor (NSF) and SNAP Receptor (SNARE) during vesicle fusion. RNAi-mediated depletion of α-SNAP in the anterior follicle cells, including the polar cells, results in egg chambers that lack both polar and border cells. Mosaic mutant analysis for α-SNAP supports these results, and over-expression of α-SNAP rescues the knockdown phenotype, confirming the specificity of the RNAi reagents. Immunofluorescent labeling reveals the localization of α-SNAP at the periphery of both the polar and border cells. Over-expression of viral anti-apoptotic gene, p35, in the α-SNAP depleted egg chamber verifies that the lack of a border cell cluster is due to impaired signaling, not apoptosis. Through suppression/enhancement assays with various regulators of the STAT signaling pathway and α-SNAP RNAi, we found that α-SNAP functions upstream of upd in the polar cells to regulate the JAK/STAT signaling pathway during border cell cluster formation. Among several SNAREs we examined, only syntaxin 1A recapitulates the lack of border cell cluster phenotype when depleted in the anterior follicle cells, indicating that it likely functions with α-SNAP during border cell cluster formation. Therefore our study identified a specific new role for α-SNAP in STAT-mediated motile cell specification.