The TFIID transcription initiation complex is composed of TBP and multiple TAFs. Studies in unicellular systems indicate that TAF250 is required for progression through G(1)/S of the cell cycle and repression of apoptosis. Here we extend these in vivo studies by determining the developmental requirements for TAF250 in a multicellular organism, Drosophila. TAF250 mutants were isolated in a genetic screen that also yielded TAF60 and TAF110 mutants, indicating that TAFs function coordinately to regulate transcription. Null alleles of TAF250 are recessive larval lethal. However, combinations of weak loss-of-function TAF250 alleles survive to adulthood and reveal requirements for TAF250 during ovary, eye, ocelli, wing, bristle, and terminalia development as well as overall growth of the fly. These phenotypes suggest roles for TAF250 in regulating the cell cycle, cell differentiation, cell proliferation, and cell survival. Finally, molecular analysis of TAF250 mutants reveals that the observed phenotypes are caused by mutations in a central region of TAF250 that is conserved among metazoan organisms. This region is contained within the TAF250 histone acetyltransferase domain, but the mutations do not alter the histone acetyltransferase activity of TAF250 in vitro, indicating that some other aspect of TAF250 function is affected. Because this region is not conserved in the yeast TAF250 homologue, TAF145, it may define an activity for TAF250 that is unique to higher eukaryotes.