Syne2b/nesprin-2 is a giant protein implicated in tethering the nucleus to the cytoskeleton and plays an important role in maintaining cellular architecture. Epiboly is a conserved morphogenetic movement that involves extensive spreading and thinning of the epithelial blastoderm to shape the embryo and organize the three germ layers. Dynamic cytoskeletal organization is critical for this process, but how it is regulated remains elusive. Here we generated a zebrafish syne2b mutant line and analyzed the effects of impaired Syne2b function during early development. By CRISPR/Cas9mediated genome editing, we obtained a large deletion in the syne2b locus, predicted to cause truncation of the nuclear localization KASH domain in the translated protein. Maternal and zygotic syne2b embryos showed delayed epiboly initiation and progression without defects in embryonic patterning. Remarkably, disruption of Syne2b function severely impaired cytoskeletal organization across the embryo, leading to aberrant clustering of F-actin at multiple cell contact regions and abnormal cell shape changes. These caused disintegration of the epithelial blastoderm before the end of gastrulation in most severely affected embryos. Moreover, the migration of yolk nuclear syncytium also became defective, likely due to disorganized cytoskeletal networks at the blastoderm margin and in the yolk cell. These findings demonstrate an essential function of Syne2b in maintaining cytoskeletal architecture and epithelial integrity during epiboly movement.