Pupal cuticle proteins of Manduca sexta: characterization and profiles during sclerotization

Muscle potentials were recorded extracellularly from developing pupae and adults of the saturniid moths Antheraea polyphemus and A. pernyi and the sphingid moth Manduca sexta. During the week prior to the terminal ecdysis, developing moths still enclosed within the pupal cuticle produced motor patterns similar to those recorded from adults during flight and shivering. The pupal patterns had a longer cycle time and were more variable than the adult motor patterns. Characteristic inter-family differences in adult motor patterns were apparent in pupal motor patterns. Development of motor patterns was followed over several days by observing individuals with chronically implanted leads. Early in the pupal period potentials were small and infrequent. The amount of activity gradually increased and became more patterned. As development proceeded adult patterns were produced for increasing lengths of time, although the patterns changed quickly and spontaneously. Restricting the wing movements of A. polyphemus adults increased the cycle time, increased the number of spikes per burst in muscles opposing the restraint, and did not alter the interspike interval within a burst. The flight patterns produced by pharate moths, in which the wings are also immobile, also have a longer cycle time than that of adult flight, but the number of spikes per burst the same and the interspike interval is longer than in adult flight. These observations suggest that the differences between pupal and adult patterns are not necessarily due to the confinement of the wings by the pupal cuticle.

Download Full-text

Properties of tyrosinase and dopa quinone imine conversion factor from pharate pupal cuticle of Manduca sexta L.

Insect Biochemistry ◽

10.1016/0020-1790(84)90103-3 ◽

1984 ◽

Vol 14 (4) ◽

pp. 463-472 ◽

Cited By ~ 55

Author(s):

Yoichi Aso ◽

Karl J. Kramer ◽

Theodore L. Hopkins ◽

Steven Z. Whetzel

Keyword(s):

Manduca Sexta ◽

Conversion Factor ◽

Quinone Imine ◽

Pupal Cuticle

Download Full-text

Incorporation of Radiolabeled Tyrosine, N-Acetyldopamine, N-β-Alanyldopamine, and the Arylphorin Manducin into the Sclerotized Cuticle of Tobacco Hornworm (Manduca sexta) Pupae

Zeitschrift für Naturforschung C ◽

10.1515/znc-1984-11-1212 ◽

1984 ◽

Vol 39 (11-12) ◽

pp. 1066-1074 ◽

Cited By ~ 40

Author(s):

Lothar Grün ◽

Martin G. Peter

Keyword(s):

Manduca Sexta ◽

Larval Instar ◽

Tobacco Hornworm ◽

Insoluble Fraction ◽

High Loss ◽

Tyrosine Residues ◽

Cuticle Sclerotization ◽

Larval Serum Protein ◽

Pupal Cuticle

Abstract Radiolabeled manducin, which is the arylphorin (i.e. tyrosine-rich larval serum protein) of the tobacco hornworm Manduca sexta was prepared by in vivo biosynthesis from [U-14C]-ʟ-tyrosine. In order to see whether manducin is incorporated into the cuticle of developing pupae and participates in cuticle sclerotization, the radiolabeled protein was injected into late last larval instar larvae. Fractionation of the sclerotized pupal cuticle into a buffer soluble, acid soluble and acid insoluble fraction shows that up to 2.6% of peptidic tyrosine residues end up in the acid soluble portion. Another 0.5% are recovered from the acid insoluble fraction by combustion. Only 30% of peptidic tyrosine residues of manducin incorporated into the acid soluble fraction are recovered as tyrosine. The presence of radioactivity in the acid insoluble fraction suggests that peptidic tyrosine residues are transformed partly into melanin-like material. The incorporation of manducin into the cuticle of pupae is also evident from immunological studies. Relative large quantities of radiolabeled acid insoluble melanine like material is also recovered from sclerotized cuticle after incorporation of radiolabeled tyrosine as well as tanning substrates N-acetyldopamine and N-β-alanyldopamine. Application of doubly labeled [7-3H,8-14C]-N-acetyldopamine shows a high loss of 3H in the acid insoluble fraction. It is suggested that tanning agents form lignine-like polymers and that sclerotization results from copolymerization with peptidic tyrosine residues in the cuticle. Thus, the arylphorin manducin appears to be an important constituent of the sclerotization system in Manduca sexta.

Download Full-text

Prepupal differentiation in Drosophila: distinct cell types elaborate a shared structure, the pupal cuticle, but accumulate transcripts in unique patterns

Development ◽

10.1242/dev.106.4.649 ◽

1989 ◽

Vol 106 (4) ◽

pp. 649-656 ◽

Cited By ~ 1

Author(s):

K. Fechtel ◽

D.K. Fristrom ◽

J.W. Fristrom

Keyword(s):

Cell Types ◽

Imaginal Discs ◽

Specific Expression ◽

Distinct Cell ◽

Cuticle Proteins ◽

Shared Structure ◽

Pupal Cuticle

The components of the pupal cuticle are the main differentiation products synthesized by both the larval and adult epidermis during the prepupal period of Drosophila development. The pupal cuticle is formed in vitro by imaginal discs in response to a 6 h pulse of 20-hydroxyecdysone (20-HE). We previously described the isolation and initial characterization of four ecdysone-dependent genes (EDGs) whose expression in imaginal discs occurs only in response to a pulse of 20-HE. In this report, we demonstrate that the pattern of temporal and tissue-specific expression of these EDGs in vivo is like that expected for genes that encode pupal cuticle proteins. Transcripts of these genes are detected in prepupae only in the epidermis and only when cuticle components are synthesized and secreted. Nonetheless, their temporal and spatial patterns of accumulation differ. EDG-84A-1 transcripts accumulate only in prepupae and only in imaginal cells. EDG-78E and EDG-64CD transcripts accumulate at the same time in both larval and imaginal cells. EDG42-A transcripts appear first in prepupae in imaginal cells and then, after a 2–4 h lag, in larval cells. It is evident that some genes are not restricted in their expression to only larval or imaginal epidermis.

Download Full-text

Metabolism and protein transport of a possible pupal cuticle tanning agent in Manduca sexta

Journal of Insect Physiology ◽

10.1016/0022-1910(74)90140-1 ◽

1974 ◽

Vol 20 (6) ◽

pp. 981-992 ◽

Cited By ~ 23

Author(s):

John K. Koeppe ◽

Lawrence I. Gilbert

Keyword(s):

Manduca Sexta ◽

Protein Transport ◽

Pupal Cuticle

Download Full-text

Physiology of Insect Ecdysis

Journal of Experimental Biology ◽

10.1242/jeb.58.3.821 ◽

1973 ◽

Vol 58 (3) ◽

pp. 821-829

Author(s):

JAMES W. TRUMAN

Keyword(s):

Nervous System ◽

Manduca Sexta ◽

Hormone Activity ◽

Hormonal Factors ◽

Corpora Cardiaca ◽

Eclosion Hormone ◽

Neural Input ◽

The Brain ◽

Pupal Cuticle

1. In pharate Manduca sexta moths eclosion hormone activity was present in the brain and corpora cardiaca. Bursicon activity was confined to the abdominal nervous system, and was most concentrated in the abdominal perivisceral organs (PVOs). 2. When newly emerged moths were given access to suitable wing-spreading sites, bursicon activity was depleted from the PVOs and appeared in the blood within 15 min after eclosion. This hormone was responsible for the tanning and hardening of the wings. 3. Bursicon release could be delayed for at least 24 h by forcing the newly emerged moth to dig. Secretion then occurred swiftly upon giving the moth a suitable wing-spreading site. 4. The pupal cuticle was removed from pharate Manduca approximately 7 h before their normal eclosion gate, and the peeled moths were provided with a wing-spreading site. These moths did not then secrete bursicon until after their normal time of eclosion. 5. Injection of the eclosion hormone into pharate moths caused early eclosion followed by precocious bursicon secretion. 6. It was concluded that bursicon release is regulated by both neural and hormonal factors. The eclosion hormone triggers a program of neural output which includes the secretion of bursicon. This release, however, can be delayed by neural input which is associated with the digging behaviour of the moth.

Download Full-text

The pupal cuticle of Drosophila: differential ultrastructural immunolocalization of cuticle proteins.

The Journal of Cell Biology ◽

10.1083/jcb.102.1.306 ◽

1986 ◽

Vol 102 (1) ◽

pp. 306-311 ◽

Cited By ~ 35

Author(s):

W J Wolfgang ◽

D Fristrom ◽

J W Fristrom

Keyword(s):

Molecular Weight ◽

Protein Composition ◽

Ultrastructural Localization ◽

Low Molecular Weight ◽

Thin Sections ◽

Lamellar Morphology ◽

Cuticle Proteins ◽

Antigenic Domains ◽

Intracellular Vesicles ◽

Pupal Cuticle

Precise ultrastructural localization of Drosophila melanogaster pupal cuticle proteins (PCPs) was achieved by the immunogold labeling of frozen thin sections. PCPs were found in lamellate cuticle and intracellular vesicles but, curiously, were absent from the assembly zone of the cuticle. Antibodies that distinguish between the two classes of PCPs--low molecular weight (L-PCPs) and high molecular weight (H-PCPs)--revealed that the morphologically distinct outer lamellae contained L-PCPs and the inner lamellae contained H-PCPs. The sharp boundary between these two antigenic domains coincides with the transition from the outer to the inner lamellae, which in turn is correlated with the cessation of L-PCP synthesis and the initiation of H-PCP synthesis in response to 20-hydroxyecdysone (Doctor, J., D. Fristrom, and J.W. Fristrom, 1985, J. Cell Biol. 101:189-200). Hence, differences in protein composition are associated with differences in lamellar morphology.

Download Full-text

The pupal cuticle of Drosophila: biphasic synthesis of pupal cuticle proteins in vivo and in vitro in response to 20-hydroxyecdysone.

The Journal of Cell Biology ◽

10.1083/jcb.101.1.189 ◽

1985 ◽

Vol 101 (1) ◽

pp. 189-200 ◽

Cited By ~ 65

Author(s):

J Doctor ◽

D Fristrom ◽

J W Fristrom

Keyword(s):

Molecular Weight ◽

High Molecular Weight ◽

Ultrastructural Changes ◽

Low Molecular Weight ◽

Before And After ◽

Cuticle Protein ◽

Cuticle Proteins ◽

Pupal Cuticle

We investigated the synthesis and localization of Drosophila pupal cuticle proteins by immunochemical techniques using both a complex antiserum and monoclonal antibodies. A set of low molecular weight (15,000-25,000) pupal cuticle proteins are synthesized by the imaginal disk epithelium before pupation. After pupation, synthesis of the low molecular weight proteins ceases and a set of unrelated high molecular weight proteins (40,000-82,000) are synthesized and incorporated into the pupal cuticle. Ultrastructural changes in the cuticle deposited before and after pupation correlate with the switch in cuticle protein synthesis. A similar biphasic accumulation of low and high molecular weight pupal cuticle proteins is also seen in imaginal discs cultured in vitro. The low molecular weight pupal cuticle proteins accumulate in response to a pulse of the insect steroid hormone 20-hydroxyecdysone and begin to appear 6 h after the withdrawal of the hormone from the culture medium. The high molecular weight pupal cuticle proteins accumulate later in culture; a second pulse of hormone appears to be necessary for the accumulation of two of these proteins.

Download Full-text