Coactivators MBF1 and MBF2 mediate BmFTZ-F1-dependent transcriptional activation in vitro by interconnecting BmFTZ-F1, TATA binding protein TBP, and TFIIA. Here, we analyzed temporal and spatial expression patterns of MBF2 during embryonic and larval development of the silkworm Bombyx mori. MBF2 was detected in unfertilized eggs and embryos until stage 26. In stage 22 embryos, MBF1, MBF2, and BmFTZ-F1 colocalize in neural cells. During the larval stage, MBF2 was not expressed in the fat body and trachea. In the silk gland, MBF2 mRNA was constitutively expressed, but MBF2 protein appeared in the period between the second day and the molting D3 stage in both the third and the fourth instars and then disappeared. MBF2 was also detected on the second and third days of the fifth instar. Immunostaining during the fourth molt showed that MBF1, MBF2, and BmFTZ-F1 localize in the nucleus only at the D3 stage, while the two cofactors are present in the cytoplasm at other stages. Immunoprecipitation experiments suggested that MBF1, MBF2, and BmFTZ-F1 form a complex at the D3 stage. Transient expression of these factors in Schneider cell line 2 revealed that MBF1 and MBF2 localize to the nucleus and enhance BmFTZ-F1-dependent transcription only when all three factors are present. These data illustrate the functional regulation of MBF1 and MBF2 at the step of nuclear transport and implicate MBF2 in tissue- and stage-specific transcription. (C) 2000 Academic Press.