Background: Breast cancer remains a major health issue, with bone metastases negatively impacting patient outcomes. The biochemical and biological functions of the exon 4-splice isoform (ZNF217-ΔE4) of the oncogenic transcription factor ZNF217 have been poorly investigated. Methods/Results: This study, for the first time, elucidates through advanced live-cell single-molecule tracking microscopy that the C-terminus of ZNF217 influences chromatin engagement and binding stability. ZNF217-ΔE4 retains its ability to be recruited and to promote positive transcriptional activity. CRISPR/Cas9-mediated silencing of the ZNF217 gene in MDA-MB-231 breast cancer cells impairs cell aggressiveness, while reintroduction of the ZNF217-ΔE4 isoform is sufficient to restore increased cell proliferation, migration, invasion, and stemness features. In vivo, ZNF217 ΔE4—although less potent than the wild-type isoform—accelerates the formation of bone marrow micrometastases. A retrospective analysis of primary breast tumors revealed that patients with high ZNF217-ΔE4 mRNA levels had a higher risk of developing bone metastases. Conclusions: Overall, this study identifies ZNF217-ΔE4 as a novel functional isoform that mediates breast cancer cell aggressiveness and bone marrow homing. It also highlights this isoform as a promising biomarker and potential therapeutic target for breast cancers at elevated risk of bone metastasis.