Purification and characterization of a biliverdin-associated protein from the hemolymph of Manduca sexta

Goodman, Walter G.; Adams, Bruce G.; Trost, Jeffrey T.

doi:10.1021/bi00326a017

Cited by 58 publications

(26 citation statements)

References 27 publications

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“…The same molecule is found in the hemolymph [l- 3,6]. Figure 1 shows that the antibody to insecticyanin purified from pupal hemolymph [I] selectively stained the pigment granules in the epidermis.…”

Section: Bidirectional Secretion Of Insecticyaninmentioning

confidence: 86%

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Developmental expression, synthesis, and secretion of insecticyanin by the epidermis of the tobacco hornworm, Manduca sexta

Riddiford

Palli

Hiruma

et al. 1990

Arch Insect Biochem Physiol

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Insecticyanin was found to be synthesized in several isoelectric forms and stored in the pigment granules in the epidermis. Both major epidermal forms (INS-a, pl 5.5; INS-b, pl 5.7) were found in the cuticle, but only the most basic form, INS-b, was present in the hemolymph. In vitro the epidermis synthesized and secreted both forms into both the cuticle and the medium. Isolation of two cDNA clones for insecticyanin followed by hybridization to epidermal mRNA showed the presence of only one 1.1 kb mRNA, but transcription of the longer cDNA yielded a RNA which produced INS-a but no INS-b. Insecticyanin mRNA was present during the intermolt feeding stages of the 4th and 5th instars and absent during the larval molt and after the onset of metamorphosis. Exposure of either day 2 4th-instar or day 1 5th-instar larval epidermis to 20-hydroxyecdysone (20HE) in vitro caused a dose-dependent decline in this mRNA that was not prevented by simultaneous exposure to JH. When synthesis resumes just before ecdysis, INS-b appears before INS-a; then on the final day of feeding, synthesis of INS-a ceases before that of INS-b. Culture experiments showed that exposure of day 15th epidermis to a small pulse of 20HE followed by an ecdysteroid-free period was best to mimic the in vivo situation, indicating that the small rise of ecdysteroid in the absence of JH on day-2 was responsible for this differential cessation.

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Section: Bidirectional Secretion Of Insecticyaninmentioning

confidence: 86%

“…Even during the prepupal period when the insecticyanin is lost from the epidermis, only INS-b was present in the hemolymph (data not shown). This INS-b spot comigrated with crystalline insecticyanin purified from either larval [3] or pupal [l] hemolymph.…”

Section: Bidirectional Secretion Of Insecticyaninmentioning

confidence: 99%

Developmental expression, synthesis, and secretion of insecticyanin by the epidermis of the tobacco hornworm, Manduca sexta

Riddiford

Palli

Hiruma

et al. 1990

Arch Insect Biochem Physiol

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“…to provide camouflage for protecting the larval development (2 -3). Biochemical studies have revealed that insecticyanin is a biliverdin IXy-associated protein (1,(4)(5) and belongs to the lipocalin family of proteins recently described by Pervaiz and Brew (6). Two-dimensional gel electrophoresis resolved two different isoelectric forms: INS-a (pl 5.5) and INS-b (pI 5.7) (7)(8).…”

Section: Introductionmentioning

confidence: 99%

The two duplicated insecticyanin genes,ins-aandins-bare differentially expressed in the tobacco hornworm,Manduca sexta

Riddiford

1994

Nucl Acids Res

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Two gene-specific probes were generated from the unique sequences in the 3' non-coding regions of the two insecticyanin genes, ins-a and ins-b to study the developmental expression of these genes in Manduca sexta. Both genes were initially transcribed in the freshly hatched first instar larvae and then expressed in the epidermis and to a lesser degree in the fat body during every larval feeding stage. In the epidermis of the 4th and 5th instar larvae, both mRNAs appeared shortly before ecdysis and accumulated to maximal levels within a day. As the larval epidermis became pupally committed on day 3 of the 5th (final) instar, INSa mRNA quickly decreased, whereas INS-b mRNA showed a second peak of accumulation. In the fat body, both genes showed a similar expression pattern within the 4th instar to that of the epidermis except that levels were lower and ins-b mRNA dominated. In the final instar, only ins-b mRNA was present in significant amount. These findings not only reveal that the two duplicated insecticyanin genes have diverged in their expression pattern but also demonstrate, for the first time, that fat body also expresses insecticyanin genes.

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“…Blue pigments are believed to function as camouflage with yellow carotenoids (Cherbas, 1973;Riley et al, 1984), and are usually associated with proteins as pigment protein complexes. sexta INS is 21.4 kDa (Riley et al, 1984;Goodman et al, 1985;Li and Riddiford, 1992), and P. brassicae BBP is composed of 19.7 kDa subunit (Suter et al, 1988). The second group is cyanoprotein (CP) from Locusta migratoria and Riptortus clavatus, composed of 76 kDa (Chino et al, 1983) and 83 kDa subunits (Chinzei et al, 1990a(Chinzei et al, ,b, 1991.…”

Section: Introductionmentioning

confidence: 99%

Comparative analysis of two biliproteins, BP1 and BP2, from haemolymph of cabbage white butterfly,Pieris rapae

Choi

Nam

Seo

et al. 2006

Arch. Insect Biochem. Physiol.

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Two blue-pigment binding proteins, BP1 and BP2, are present in larval and pupal haemolymph of cabbage white butterfly, Pieris rapae, and fluctuate in expression during development. Both BP1 and BP2 are found in pupal haemolymph in varying proportions as well as in adult haemolymph, while only small amounts of BP2 are found in larval haemolymph. BPs are separated by 75% ammonium sulfate, and then purified effectively by ion exchange column chromatography and preparative gel electrophoresis. It was shown that BP1 and BP2 have molecular masses of 20,244 and 19,878 Da, and isoelectric points of 7.0 and 6.8, respectively. Considering their amino acid compositions and N-terminal amino acid sequences, the two proteins are almost identical except the first N-terminal amino acid. The first amino acid of BP1 is asparagine, whereas the initial residue of BP2 is aspartic acid. Anti-BP1 cross-reacts with BP2, indicating that they have immunological homogeneity. Western blotting analyses revealed that only BP1 was present in the larval tissues such as fat body, integument, muscle, and hindgut. However, BP1 was not found in midgut, Malphigian tubules, and silk gland. BP1 was also present in the protein bodies, and both cuticle and hemocoel sides of larval epidermis cells by the transmission electron microscopic observation. The information in this report will facilitate studies on the molecular biology and biological significance of insect BPs.

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Purification and characterization of a biliverdin-associated protein from the hemolymph of Manduca sexta

Cited by 58 publications

References 27 publications

Developmental expression, synthesis, and secretion of insecticyanin by the epidermis of the tobacco hornworm, Manduca sexta

Developmental expression, synthesis, and secretion of insecticyanin by the epidermis of the tobacco hornworm, Manduca sexta

The two duplicated insecticyanin genes,ins-aandins-bare differentially expressed in the tobacco hornworm,Manduca sexta

Comparative analysis of two biliproteins, BP1 and BP2, from haemolymph of cabbage white butterfly,Pieris rapae

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