The ribosomal proteins of Drosophila melanogaster. II. Comparison of protein patterns of ribosomes from larvae, pupae and adult flies by two-dimensional polyacrylamide gel electrophoresis

Summary The ribosomal proteins from two larval and two pupal stages, within 24 hours before and after pupation, respectively, and adult flies were extracted and compared by two-dimensional polyacrylamide gel electrophoresis. This technique resolved 53 larval, 50 pupal and 52 adult ribosomal proteins, forming a complex pattern. Some proteins were found only in one stage or the other. At present it is not possible, however, to classify these proteins as stage-specific. Some spots showed considerable increase in their staining intensities from one stage to the other, whereas, at the same time, other spots faded. In the ribosomal protein pattern of adult flies 3 proteins showed altered electrophoretic mobilities as compared to earlier developmental stages.Hormones involved in insect development, epigenetic control and non-ribosomal proteins are discussed as possible causes of the variations in the ribosomal protein composition.     

Characterization of N-ethylmaleimide-reactive proteins from human tonsillar ribosomes by two-dimensional polyacrylamide gel electrophoresis.

Human tonsillar 80-S ribosomes were 17% and 43% inactivated by 1 mM N-ethylmaleimide after 12 min at 30 or 37 degrees C, respectively. The ribosomes were unaffected by the reagent during the same period of time at 0 or 20 degrees C. 4, 12, 27 and 59 sulfhydryl groups per 80-S ribosomes were found labeled by 1 mM N-ethyl[14C] maleimide after 12 min at 0, 20, 30 or 37 degrees C, respectively. The analysis of radioactively labeled proteins by two-dimensional gel electrophoresis revealed the following: after 3 min at 37 degrees C only two 40-S proteins, S3 and S7, displayed a significant amount of label. After 12 min at 37 degrees C, there was a several-fold increase in the extent of radioactivity found in each of these proteins and, additionally, S1, S2, S4, S5, S15, S22 and S31 were also found among labeled 40-S proteins. S3 appeared to be the most N-ethylmaleimide-reactive 40S protein. After 3 min at 37 degrees C, L10, L17, L20 (and/or S20), L26, L32 and L33, and after 12 min at 37 degrees C, additionally L1, L2, L7, L9, L11, L15, L16, L18, and L25 were labeled among 60-S proteins. l17 and 32 were the most N-ethylmaleimide-reactive proteins under these conditions. After 12 min at 37 degrees C, approx. 26% and 39% of the radioactivity incorporated into the 80 S or 60 S ribosomal protein, respectively, was found in these two proteins. After 12 min at 0 degrees C, S3, L17, L32 and L33 were the only labeled proteins.     

Lectin activity in adult and larval Drosophila melanogaster

A soluble lectin activity is described in extracts prepared from adult and third instar larval stages of Drosophila melanogaster. Based on erythrocyte specificity and hapten inhibition studies the lectin from both sources appears to be identical. Extracts from both sources were found to contain an endogenous inhibitor of lectin activity which may serve as the natural receptor for the lectin.     

Calcium homeostasis in larval and adult Drosophila melanogaster

Calcium homeostasis in Drosophila melanogaster was examined in response to the challenges imposed by growth, reproduction and variations in dietary calcium content. Turnover time for calcium, calculated as the time for (45)Ca(2+)to accumulate to half the steady state value of 3.46 nmol/fly, was 3.3 days. Although larvae weighed 2x as much as adults, they contained 3-4x as much calcium. Anterior Malpighian tubules (Mts) contain much more calcium than posterior Mts, accounting for 25-30% of the calcium content of the whole fly. In response to a 6.2-fold increase in dietary calcium level, calcium content of whole flies increased only 10%. Hemolymph calcium concentration ( approximately 0.5 mM) was similar in males and females and in animals raised on diets differing in calcium content. Fluid secretion rate, secreted fluid calcium concentration, and transepithelial calcium flux in tubules isolated from flies raised on high and low calcium diets did not differ significantly. Malpighian tubules secrete calcium at rates sufficient to eliminate whole body calcium content in 0.5 and 3 days for tubules secreting fluid at basal and maximal rates, respectively. It is suggested that flies absorb high quantities of calcium from the diet and maintain homeostasis through the combined effects of elimination of calcium in fluid secreted by the Malpighian tubules and the sequestration of calcium in granules, especially within the distal segment of the anterior pair of Malpighian tubules.     

Characterization of ribosomal proteins from different tissues and species of animals by electrophoresis on polyacrylamide gel

Summary Proteins from ribosomes of different tissues and animals were characterized by polyacrylamide disc electrophoresis. The proteins from ribosomes of different tissues from the same animal are qualitatively similar. The results of the experiments with ribosomes from the livers of different species of animals exhibit clear differences, in the electrophoretic patterns of the proteins.     

Pupal and larval cuticle proteins of Drosophila melanogaster

Proteins, soluble in 7 M urea, were extracted from third-instar larval and pupal cuticles of Drosophila melanogaster. Both extracts contain a limited number of polypeptides resolved by one- or two-dimensional electrophoresis. The five major larval proteins have low molecular weights (less than 20000) and are not glycosylated. The major pupal cuticle proteins fall into two size classes: two with apparent molecular weights of 56K and 82K and four with molecular weights between 15K and 25K. The proteins with high apparent molecular weights are glycosylated. In nondenaturing gels, no components of the larval and pupal cuticle extracts comigrate. One-dimensional "fingerprints" indicate that cuticle proteins from these two stages have unique primary structures. Immunological results indicate that the major low molecular weight larval and pupal cuticle proteins are comprised of two families of proteins that share antigenic determinants. The high molecular weight pupal cuticle proteins are immunologically unrelated to the low molecular weight components. We conclude that the pupal and larval proteins are encoded in part by multigene families that have arisen by gene duplication and evolutionary divergence.     

Regulation of synthesis of larval serum proteins after transplantation of larval fat body into adult Drosophila melanogaster

The larval serum proteins, LSP1 and LSP2, of Drosophila melanogaster are synthesized by the fat body during the third instar. We examined the potential for LSP synthesis by fat body implants in adult flies. Fat body from third instar donors will continue to synthesize LSPs in both males and females. Implants from late second instar larvae will start synthesizing LSP1 and LSP2 in females but only LSP1 in males, suggesting that regulation of these proteins is not the same and that the physiological milieu in the two sexes differs. The newly synthesized LSPs are secreted into the hemolymph for approximately 48 hr when secretion stops but synthesis continues. This sequence follows the pattern for LSP secretion in situ. Fat body from mid second instar larvae is variable in its ability to synthesize LSPs. LSPs are not detected in implants from first instar larvae despite there being a high level of protein synthesis in the implant and considerable growth of the fat body cells. We conclude that there is a critical stage of differentiation during the latter half of the second instar when the fat body becomes independent of the larval milieu and can synthesize LSPs in the adult.     

The ribosomal proteins of Drosophila melanogaster. Localization of stage specific proteins on the small and large ribosomal subunit

Summary By means of SDS-polyacrylamide gel electrophoresis the protein patterns of untreated, 0.5M and 1M KCl treated ribosomes of Drosophila melanogaster 4d larvae and adults are compared. 0.5M KCl treatment does not change the stage specific pattern (Fig. 1 b, c). 1 M KCl treatment removes 2 stage specific proteins from adult ribosomes (Fig. 1d, band No. 1 and 3). One protein found only in the adults is localized on the small ribosomal subunit and removed by 1 M KCl (band No. 3). The possible significance of these results is discussed.This work was supported by the Georges und Antoine Claraz-Schenkung.