14-3-3ζ is overexpressed in more than 40% of breast cancers, but its pathophysiologic relevance to tumorigenesis has not been established. Here, we show that 14-3-3ζ overexpression is sufficient to induce tumorigenesis in a transgenic mouse model of breast cancer. MMTV-LTR promoter-driven HA-14-3-3ζ transgenic mice (MMTV-HA-14-3-3ζ) developed mammary tumors, whereas control mice did not. Whey acidic protein promoter-driven HA-14-3-3ζ transgenic mice (WAP-HA-14-3-3ζ) developed hyperplastic lesions and showed increased susceptibility to carcinogen-induced tumorigenesis. When crossed with MMTV-neu transgenic mice, 14-3-3ζ.neu transgenic mice exhibited accelerated mammary tumorigenesis and metastasis compared with MMTV-neu mice. Mechanistically, 14-3-3ζ overexpression enhanced MAPK/c-Jun signaling, leading to increased miR-221 transcription, which inhibited p27 CDKI translation and, consequently, promoted cell proliferation. Importantly, this 14-3-3ζ-miR-221-p27 proliferation axis is also functioning in breast tumors in patients and is associated with high-grade cancers. Taken together, our findings show that overexpression of 14-3-3ζ has a causal role in mammary tumorigenesis and progression, acting through miR-221 in cooperation with known oncogenic events to drive neoplastic cell proliferation.