Summary
Wing imaginal discs isolated from last instar larvae of the Indian meal moth,Plodia interpunctella, produced chitin when incubated in vitro with ≧2×10−7 M 20-hydroxyecdysone. Chitin biosynthesis was initiated 8 h after the conclusion of a 24-h treatment with hormone. Simulataneous incubation of wing discs with 20-hydroxyecdysone and either inhibitors of RNA synthesis (alpha-amanitin, actinomycin-D) or inhibitors of protein systhesis (cycloheximide, puromycin) prevented chitin biosynthesis. We conclude from our results that RNA and protein synthesis must continue undiminished during the hormone-contact period, and that synthesis of protein, but not of new RNA is required during the posthormone culture period. Our findings are consistent with the hypothesis that ecdysteroids stimulate insect metamorphosis by promoting the synthesis of new RNA and protein during a hormone-dependent phase followed by hormone-independent protein synthesis.
Similar content being viewed by others
References
Ashburner M (1974) Sequential gene activation by ecdysone in polytene chromosomes ofDrosophila melanogaster II. The effects of inhibitors of protein synthesis. Dev Biol 39:141–157
Ashburner M, Chihara C, Meltzer P, Richards G (1974) On the temporal control of puffing activity in polytene chromosomes. Cold Spring Harbor Symp Quant Biol 38:655–662
Baliga BS, Pronczuk AW, Munro HN (1969) Mechanism of cycloheximide inhibition of protein synthesis in a cell-free system prepared from rat liver. J Biol Chem 244:4480–4489
Benson J, Oberlander H (1974) Protein synthesis in wing discs ofGalleria mellonella: effects of ecdysone and fat body in vitro. Insect Biochem 4:423–428
Bergtrom G, Oberlander H (1975) RNA synthesis in imaginal disks ofGalleria mellonella: effects of alpha-and beta-ecdysone and fat body in vitro. J Insect Physiol 21:39–52
Campbell AJ, Birt LM (1975) Effect of actinomycin D on soluble α-glycero phosphate dehydrogenase activity in the sheep blowfly,Lucilia cuprina. Insect Biochem 5:113–117
Clever U, Romball CG (1966) RNA and protein synthesis in the cellular response to a hermone, ecdysone. Proc Natl Acad Sci USA 56:1470–1476
Dutkowski AB, Oberlander H (1974) Interactions between betaecdysone and fat body during wing disk development in vitro. J Insect Physiol 20:743–749
Ferkovich SM, Oberlander H, Leach CE, Essen F Van (In press 1979) Hormonal control of chitin biosynthesis in imaginal discs. Proc Inter Conf on Invertebrate Tissue Culture, Rigi, Switzerland
Fristrom JW, Logan WR, Murphy C (1973) The synthetic and minimal culture requirements for evagination of imaginal dises ofDrosophila metanogaster in vitro. Dev Biol 33:441–456
Grayson S, Berry J (1973) Estimation of the half-life of a secretory protein message. Science 180:1071–1072
Jacob ST, Sagdel EM, Munro HN (1970) Different responses of soluble whole nuclear RNA polymerase and soluble nucleolar RNA polymerase to divalent cations and to inhibition by α-amanitine. Biochem Biophys Res Commun 38:765–770
Kafatos FC, Reich J (1968) Stability of differentiation-specific and non-specific messenger RNA in insect cells. Proc Natl Acad Sci USA 60:1458–1465
Lindell TJ (1976) Evidence for an extra nucleolar mechanism of actinomycin D action. Nature 263:347–350
Marks EP, Sowa BA (1976) Cuticle formation in vitro. In: Hepburn HR (ed.) The Insect Integument. Elsevier, Amsterdam, pp 339–357
Nowock J, Sridhara S, Gilbert I (1978) Evidence for the existence of two forms of RNA polymerase I and II in insect wing epidermis. Biochim Biophys Acta 520:393–403
Oberlander H (1976) Hormonal control of growth and differentiation of insect tissues cultured in vitro. In Vitro 12:225–235
Oberlander H, Ferkovich SM, Essen F Van, Leach CE (1978) Chitin biosynthesis in imaginal discs cultured in vitro. Wilhelm Roux' Archiv Entwicklungsmech Org 185:95–98
Perry RP, Kelley DE (1970) Inhibition of RNA synthesis by actinomycin D: characteristic dose-response of different RNA species. J Cell Physiol 76:127–140
Phillips DR, Loughton BG (1979) The effect of inhibition of RNA and protein synthesis on the development of larvae ofLocusta migratoria. Insect Biochem 9:241–245
Shaaya E, Clever U (1972) In vitro effects of α-amanitin on RNA synthesis inCalliphora erythrocephala. Biochem. Biophys Acta 272:373–381
Silhacek DL, Miller GL (1972) Growth and development of the Indian meal moth,Plodia interpunctella, under laboratory massrearing conditions. Ann Entomol Soc Am 65:1084–1087
Stevenson JR, Tung DA (1971) Inhibition by actinomycin D of the initiation of chitin biosynthesis in the crayfish. Comp Biochem Physiol 39B:559–567
Traut RR, Monro RE (1964) The puromycin reaction and its relation to protein synthesis. J Mol Biol 10:63–72
Vardanis A (1976) An in vitro assay system for chitin synthesis in insect tissue. Life Sci 19:1949–1956
Author information
Authors and Affiliations
Rights and permissions
About this article
Cite this article
Oberlander, H., Ferkovich, S., Leach, E. et al. Inhibition of chitin biosynthesis in cultured imaginal discs: Effects of alpha-amanitin, actinomycin-D, cycloheximide, and puromycin. Wilhelm Roux' Archiv 188, 84–86 (1980). https://doi.org/10.1007/BF00848613
Received:
Accepted:
Issue Date:
DOI: https://doi.org/10.1007/BF00848613