Proper lymph node (LN) development requires tumor necrosis factor–related activation-induced cytokine (TRANCE) expression. Here we demonstrate that the defective LN development in TRANCE^−/− mice correlates with a significant reduction in lymphotoxin (LT)αβ⁺α₄β₇⁺CD45⁺CD4⁺CD3⁻ cells and their failure to form clusters in rudimentary mesenteric LNs. Transgenic TRANCE overexpression in TRANCE^−/− mice results in selective restoration of this cell population into clusters, and results in full LN development. Transgenic TRANCE-mediated restoration of LN development requires LTαβ expression on CD45⁺ CD4⁺CD3⁻ cells, as LNs could not be induced in LTα^−/− mice. LTα^−/− mice also showed defects in the fate of CD45⁺CD4⁺CD3⁻ cells similar to TRANCE^−/− mice. Thus, we propose that both TRANCE and LTαβ regulate the colonization and cluster formation by CD45⁺ CD4⁺CD3⁻ cells in developing LNs, the degree of which appears to correlate with the state of LN organogenesis.