The spleen tyrosine kinase (Syk) and ζ-associated protein of 70 kD (ZAP-70) tyrosine kinases are both expressed during early thymocyte development, but their unique thymic functions have remained obscure. No specific role for Syk during β-selection has been established, and no role has been described for ZAP-70 before positive selection. We show that Syk and ZAP-70 provide thymocytes with unique and separable fitness advantages during early development. Syk-deficient, but not ZAP-70–deficient, thymocytes are specifically impaired in initial pre-TCR signaling at the double-negative (DN) 3 β selection stage and show reduced cell-cycle entry. Surprisingly, and despite overlapping expression of both kinases, only ZAP-70 appears to promote sustained pre-TCR/TCR signaling during the DN4, immature single-positive, and double-positive stages of development before thymic selection occurs. ZAP-70 promotes survival and cell-cycle progression of developing thymocytes before positive selection, as also shown by in vivo anti-CD3 treatment of recombinase-activating gene 1–deficient mice. Our results establish a temporal separation of Syk family kinase function during early thymocyte development and a novel role for ZAP-70. We propose that pre-TCR signaling continues during DN4 and later stages, with ZAP-70 dynamically replacing Syk for continued pre-TCR signaling.