Many of the early ecdysteroid-induced transcription factors appear during both larval molting and metamorphosis, but the Broad Complex (BRC) only appears in most tissues in the absence of juvenile hormone (JH) at metamorphosis in both Drosophila and the tobacco hornworm larva, Manduca sexta. In Manduca abdominal epidermis, pupal commitment occurs at the end of the larval feeding period in response to low 20-hydroxyecdysone (20E) in the absence of JH. At this time 20E activates BRC transcription, but it can be suppressed by JH. Immunocytochemistry shows that BRC expression occurs first in the areas of the segment that first become committed to pupal differentiation and hence will form pupal cuticle in the subsequent molt. Once BRC expression is turned on, its subsequent regulation by ecdysteroids is unaffected by JH and it persists through the pupal molt. Hence, BRC is one of the first molecular markers of pupal commitment in the abdominal epidermis. The wing disc and the eye primordium however respond differently to both 20E and JH in the final instar. The wing disc forms early in larval life and proliferates as the larva grows. It first acquires ecdysone receptors (EcR) at onset of the final instar and becomes able to form pupal cuticle during a larval molt although it continues to proliferate until the onset of pupal development. Coincidentally BRC mRNA appears, but its expression cannot be suppressed, only delayed, by JH given during the fourth instar molt. Preliminary experiments suggest that BRC protein at this time is present only in the peripodial membrane and later appears in the disc cell themselves at the ecdysteroid-induced onset of pupal development. The eye primordium makes larval cuticle through the penultimate molt, then early in the final instar as the JH titer falls, begins to proliferate to form the eye disc. In vitro studies show that only the absence of JH is necessary for this proliferation to occur. Once this proliferation has begun, this disc is committed to pupal differentiation. Yet BRC does not appear until the disc is exposed to low ecdysteroid in the absence of JH. In the latter state the eye disc begins organizing its ommatidial clusters, a process dependent on low ecdysteroid and unaffected by JH. Thus, BRC seems to be important in all three tissues for ecdysteroid-induced pupal differentiation, but its role in pupal commitment of imaginal tissues remains to be fully resolved and will be discussed. Supported by NIH and NSF.

Index words: Manduca sexta, Broad Complex