Temporal changes in DNA synthesis of prothoracic gland cells during larval development and their correlation with ecdysteroidogenic activity in the silkworm, Bombyx mori

Abstract

DNA synthesis in prothoracic gland cells of the silkworm, Bombyx mori, was studied immunocytochemically after in vivo labeling with 5-bromo-2'-deoxyuridine (BrdU), and its developmental changes during the 3rd, 4th, and last larval instars were examined. During the early stages of both the 3rd and 4th larval instars, a dramatic increase in the number of DNA-synthesizing cells of the prothoracic glands was detected. However, during the latter stages of each instar, the number of DNA-synthesizing cells greatly decreased. The determination of glandular protein content showed that dramatic increases occurred during the latter stages of each larval instar. Comparison of changes in prothoracic gland cell DNA synthesis with ecdysteroidogenic activity showed that the increase in DNA synthesis precedes ecdysteroidogenesis. The cellular mechanism underlying changes in prothoracic gland cell DNA synthesis during the last two larval instars was further analyzed by determining the in vitro DNA synthesis of the glands, their responsiveness to hemolymph growth factors, and changes in the growth-promoting activity of hemolymph during development. It was found that both growth factors and the responsiveness of the prothoracic gland cells to growth factors from hemolymph may play roles in regulating DNA synthesis of gland cells.

Inhibitory effects of starvation on prothoracic gland cell DNA synthesis during the last larval instar of the silkworm, Bombyx mori

“…Our previous results indicated that it is not changes in the growth-promoting activity of hemolymph, but changes in the responsiveness of the prothoracic gland cells to the growth factors which may, at least in part, contribute to alterations in DNA synthesis during both the fourth and last larval instars (Gu and Chow, 2005). The exact nature of the growth factor that activates prothoracic gland cell DNA synthesis remains to be explored, but the nutrition-dependent response to the growth factor in prothoracic gland cells is similar to endoreplicating cells in Drosophila (Britton et al, 2002).…”

“…because our previous study showed that hemolymph from this stage showed the highest growthpromoting effects (Gu and Chow, 2005). As shown in Figure. 4A, when prothoracic glands from control day 4 and day 5 last instar larvae were incubated in medium containing 10% hemolymph, higher levels of DNA synthesis were observed compared with those in control medium.…”

“…Glands from both control and starved larvae were incubated in medium containing BrdU with or without 10% hemolymph. Our previous study showed that the addition of 10% hemolymph from both the fourth and last larval instar larvae into incubation medium greatly increased DNA synthesis of the prothoracic glands (Gu and Chow, 2005). In this study, the hemolymph was derived from day 3 fourth instar larvae.…”

“…This result clearly indicates that continued feeding is necessary for prothoracic glands to maintain high levels of DNA synthesis. Temporal changes in DNA synthesis during the fourth larval instar showed that the activity of DNA synthesis reached a peak on day 1 and then gradually decreased until day 4 (Gu and Chow, 2005). The effects of starvation on fourth instar Fig.…”

“…The prothoracic glands from day 4 last instar larvae were selected to examine changes in the responsiveness to the hemolymph growth factor because we previously found that basal levels of DNA synthesis were low at this stage; however, they showed sensitive responses to stimulatory factors from hemolymph of day 3 fourth instar larvae (Gu and Chow, 2005). Figure 4B shows that when the glands were incubated in control medium, the level of DNA synthesis was very low.…”