The association of changes in cerebral ganglion dopamine metabolism with ootheca development in Periplaneta americana

• 1.1. Dopamine levels and DOPA-decarboxylase activity were measured in cerebral ganglia and haemolymph of female Periplaneta americana.
• 2.2. Measurements were made at four points in the oothecal cycle of cockroaches known to drop oothecae at regular three day intervals.
• 3.3. Dopamine levels and DOPA-decarboxylase activity in haemocytes and plasma cycle in phase with ootheca formation; their levels in haemolymph are maximal when a half visible, untanned ootheca is present.
• 4.4. In the cerebral ganglia dopamine levels and DOPA-decarboxylase also cycle in phase with ootheca formation suggesting that cerebral ganglion dopamine metabolism is under the same controls as dopamine metabolism associated with oothecal tanning.

     

Chitin synthesis in larval and pupal epidermis of the Indian meal moth,Plodia interpunctella (Hübner), and the greater wax moth,Galleria mellonella (L.)

The level of chitin synthesis was determined in whole last-instar larvae and in pupae of Plodia interpunctella, and in epidermal tissue from similar stages of Galleria mellonella. The incorporation of radiolabelled N-acetyl-d-glucosamine into chitin was used to measure synthesis. Chitin production was similar in both species with peaks of synthesis occurring at the beginning of the last larval instar, in prepupae, in white pupae and prior to adult emergence. P. interpunctella also exhibited an additional small increase at mid-instar. Exposure of larval epidermis of P. interpunctella to 20-hydroxyecdysone in vitro stimulated chitin synthesis, but only after a 24 hr lag period subsequent to exposure to the hormone. This hormonal stimulation of chitin synthesis was inhibited by actinomycin-D and cycloheximide which suggested that 20-hydroxyecdysone-stimulated production of chitin depended on synthesis of RNA and protein. Comparison of the synthesis of chitin in epidermis of G. mellonella with previously published ecdysone titres, indicated that chitin production in vivo is preceded by an elevated ecdysone titre.     

PROTEIN SYNTHESIS IN PANCREAS OF FASTED PIGEONS

The regulation of protein synthesis in the pigeon has been studied by comparing the capability of cell-free amino acid incorporating systems of membrane-bound and membrane-free polysomes prepared from fasted and fed birds. New methods were developed for isolating polysomes since techniques used for other tissues did not provide quantitative recovery of polysomal RNA. The sucrose gradient profile of polysomes from pigeon pancreas showed a predominance of trisome species. Although initiation factors are present on polysomes, it was found that polysomes in cell-free systems would not initiate protein synthesis without exogenous initiation factors. This suggested the presence of an inhibitor or regulator of protein synthesis. These studies show that fasting resulted in: (a) decreased amounts of polysomes; (b) disaggregation of polysomes to monosomes; (c) decreased capability of polysomes to synthesize nascent peptides and to initiate additional synthesis, apparently not related to concentration of initiation factors.     

The termination of the synthesis of proteins in vitro with extracts from the undeveloped cyst of Artemia salina

Soluble fractions (S-100) from both undeveloped cysts and developing embryos of Artemia salina promoted elongation of polypeptides initiated in vivo on polysomes of developing embryos or nauplius larvae. The ability of the extract from the undeveloped cyst to terminate correctly the synthesis of polypeptides has been determined indirectly from the distribution of polysomes before and after in vitro translation and, more directly, from the nature of the protein product released from rabbit reticulocyte polysomes. The extract from the undeveloped cyst and also, as expected, that from the developing embryo catalyzed a reduction in the amount of the polysomes of larger size and an increase in the amount of 80 S ribosomes. The soluble extract from the undeveloped cyst can terminate the synthesis of rabbit globin on reticulocyte polysomes. The major polypeptide product released from the polysomes had an electrophoretic mobility identical with that of the subunit of isolated rabbit globin. This indicated that the cyst contained the components necessary to complete and terminate the synthesis of polypeptides correctly and that the released protein product was not predominantly as a result of premature chain termination. The size distribution of Artemia salina proteins released from polysomes from developing embryos was similar when the synthesis was directed by the S-100 at each stage of development.     

Further studies on eukaryote DNA stimulation of amino acid incorporation in E. coli extracts

DNA from adenovirus-2 and mouse myeloma tumors stimulate RNA synthesis and amino acid incorporation into protein in a cell-free extract from Escherichia coli. The RNA synthesis is dependent on exogenous DNA, and the RNA can be hybridized to respective template DNA. A major part of this RNA is also found attached to E. coli polysomes suggesting that RNA with messenger-like activity has been synthesized. However, the in vitro-synthesized polypeptides using adenovirus DNA or myeloma DNA do not correspond in size or antigenic activity to either the virion proteins or immunoglobulins, respectively.     

Effects of juvenile hormone on polysome integrity

Juvenile hormone inhibits protein and RNA synthesis in cell cultures from Trichoplusia ni and in the testicular germinal cysts of Hyalophora cecropia pupae in vitro. Sucrose gradient analyses revealed that the polysomes of both the T. ni cells and the germinal cysts were disaggregated almost immediately after the addition of juvenile hormone in vitro with a corresponding dose-dependent increase in monosomes. It is suggested that previous reports revealing juvenile hormone inhibition of ecdysone stimulated RNA and protein synthesis may be due to polysome disaggregation. Further studies demonstrated that the effect is not restricted to insect cells and can be elicited by several other lipids devoid of juvenile hormone morphogenetic activity. Experiments with broken cell preparations and isolated polysomes suggest the necessity of cell membrane integrity for the effect on the polysomes. Several probing studies utilizing cycloheximide, ribonuclease, and high K⁺ concentrations were conducted on the means by which juvenile hormone and other lipids may elicit polysome disaggregation.     

Untersuchungen zur Veränderung der Unkrautflora bei mehrmaligem Getreidenachbau auf einer Tonschwarzerde des Thüringer Beckens

The latent effects of precocenes I and II (PI and PH) and juvenile hormone I (JHI) when topically applied to the last three instars of Spodoptera littoralis (Boisd.) larvae have been studied. Application of both PI or PII resulted in morphogenetic abnormalities resemble some effects induced by administration of JHI, e.g., larval‐pupal intermediate, partial or severe cases of untanned pupae and imperfect moths. In PII‐treatments, the effect was instar‐dose‐dependent. The intermediate dose (55 μg) was more effective on S. littoralis larvae than other doses. The effectiveness of both doses of 40 or 70 μg in production of deformed larvae and pupae decreased when applied as repeated doses instead of single ones. In Pi‐treatments, the lower dose (25 μg) was more harmful to Spodoptera larvae than the higher dose (70 μg). Repeated application by either lower or higher doses did not enhance the production of imperfect insects. Application of JHI induced symptoms ranging from supernumerary instars, larval‐pupal intermediate, untanned pupae and deformed adults. The effect was dose‐dependent. In all tested compounds, there apparently was a latent or delayed effects. Although the insects were treated while they were larvae the complete effects were not apparent until after the insect had become a pupa or an adult. More efforts will be needed to understand how does precocene interferes with the process of tanning or sclerotization?     

Comparison between the sclerotization of adult and larval cuticle in Schistocerca gregaria

Femur cuticle from fifth instar larvae of the desert locust, Schistocerca gregaria, has been characterized with respect to composition, rate of deposition, and rate of sclerotization. The results are compared with those from adult cuticle of the same species. The protein compositions of the two types of cuticle are very similar, but the rates of deposition of both protein and chitin are different. The main difference is, however, that sclerotization is restricted to the first day after ecdysis in larval cuticle, whereas in adult cuticle sclerotization continues for at least a couple of weeks. The result is that the endocuticle remains untanned in the larvae, whereas in the adults the whole cuticle becomes tanned.     

The cuticular proteins of Tenebrio molitor: II. Patterns of synthesis during postembryonic development

The program of synthesis for the soluble cuticular proteins of Tenebrio molitor was determined by following the incorporation of labeled leucine after a 4-hr pulse in vivo. Soluble proteins were extracted from labeled cuticles and separated on SDS-polyacrylamide slab gels; individual gel slices were counted. The synthetic patterns of larvae and pupae were similar to one another but distinct from the adult pattern. At each stage, the preecdysial pattern was unlike that of postecdysial animals. Distinct periods of synthesis were detected for different proteins. One protein was synthesized and deposited throughout cuticle formation in all three metamorphic stages. One group was synthesized only after ecdysis, while synthesis and secretion of other proteins were restricted to the preecdysial period. Some cuticular proteins never acquired detectable label.     

Characterization of the major haemolymph proteins in Ceratitis capitata

There are four major proteins species in the haemolymph of Ceratitis capitata. Their immunological relationship to calliphorin has been shown by immunoprecipitation and Ouchterlony double diffusion test. Using SDS electrophoresis these proteins are separated into three polypeptide bands differing in molecular weight maximally by ± 2000–3000 and showing similar migration to calliphorin. From visual inspection of the stained gels it appeared that the amounts of these proteins increase from that in the three day old larvae, reaching maximal levels in the four day old larvae and white pupae and then decrease gradually throughout the pupal and early adult stages.     

A possible therapy by juvenile hormone I to Spodoptera littoralis (Boisd.) larvae previously treated by precocene II

An effective constant dose (55 μg) of precocene II (PII) was topically tested against the last three instars of Spodoptera littoralis (Boisd.) larvae. Application of PII induced morphogenetic abnormalities typical of juvenile hormone (JH) excess. The resultant imperfect insects included larval‐pupal mosaic and partial or severe cases of untanned pupae. The sixth‐instar larvae were more sensitive to PII administration than the two preceding instars. However, sensitivity of the last larval instar to PII varied with the timing of dose application relative to the developmental status of the larvae. Whereas the newly ecdysed (0‐day old) larvae were more sensitive, the older larvae of the same sixth‐instar showed sharp decrease in their sensitivity to PII with a concomitant increase of their age. Application of a single dose (5 μg) of JH I to PII pre‐treated larvae significantly (P < 0.001) reduced the production of imperfect insects where many PH‐treated larvae developed successfully to apparent normal pupae. Although a single dose of PII was more effective on S. littoralis larvae than repeated daily doses, the effectiveness of JHI‐therapy to PH pre‐treated larvae by repeated doses was less effective in producing perfect insects than JH‐therapy to PII pre‐treated larvae by single doses. The reversal of any of these by applied JHI is quite interesting but the mechanisms remain to be unraveled.     

Movement of ribosomes over messenger RNA in polysomes of rel + and rel - Escherichia coli strains

The in vitro movement of ribosomes over messenger RNA was studied in both the presence and the absence of protein synthesis. For this purpose, labeled polysomes were extracted from rel⁺ and rel^? strains of Escherichia coli grown in the presence of radioactive uracil and incubated in a cell-free system containing tRNA, amino acids, soluble enzymes and a source of energy. The gradual conversion of the labeled polysomes into monosomes and ribosomal subunits was followed by subjecting the reaction mixture to sucrose gradient sedimentation after various incubation times and measuring the radioactivity present in the three relevant ribosomal fractions.It was found that when the conditions of incubation allow protein synthesis to occur, polysomes extracted from rel⁺ and rel^? cells are converted mainly into free monosomes, which can be made to dissociate into subunits by high-sodium or low-magnesium ion concentrations. Under conditions in which protein synthesis cannot occur because a mutant aminoacyl-tRNA synthetase has been rendered inactive, polysome conversion still occurs, though to a reduced extent. When the products of such residual run-off are examined, however, a difference is manifest between polysomes extracted from rel⁺ and from rel^? strains: whereas the polysomes from the rel^? strain are still converted into free monosomes even in the absence of protein synthesis, the polysomes from the rel⁺ strain are now converted mainly into subunits. It can be inferred, therefore, that ribosomes from rel^? bacteria, but not those from rel⁺ bacteria, continue movement over messenger RNA in the absence of protein synthesis.Studies of mixed extracts from rel^? and rel⁺ bacteria have shown that the character of the run-off process does not depend on the source of tRNA and soluble enzymes; the proportions of monosomes and subunits among the run-off products formed in the absence of protein synthesis depend only on the source of the polysomes. It is suggested that the mutation of the rel gene alters the functional architecture of ribosomes.     

Major haemolymph proteins inCeratitis capitata: Biosynthesis and secretion during development

Summary The accumulation of major haemolymph proteins (a group of proteins immunologically related to Calliphorin) their biosynthesis in vivo and in organ culture as well as their secretion, has been studied during the late larval stages and white pupae of the Mediterranean fruit flyCeratitis capitata. The accumulation of major haemolymph proteins in the haemolymph, shows a twenty fold increase from the 4-day old larvae to the white pupae stage, while in the fat body there is only a seven fold increase. It is evident from the in vivo and organ culture studies, that the major haemolymph proteins are synthesized during the late larval stage and their synthesis declines abruptly during the stage of white pupae. It seems also that each polypeptide has its own characteristic developmental kinetics of synthesis. The major haemolymph proteins are synthesized in the fat body and are very quickly secreted into the haemolymph.     

Factors influencing juvenile hormone esterase activity in the wax moth, Galleria mellonella

The effects of juvenile hormone, antiallatotropins, selected surgical procedures and starvation on the juvenile hormone esterase levels in Galleria larvae and pupae were investigated. JH reduced JH esterase activity in larvae but induced the enzyme in 1-day-old pupae. In vitro studies confirmed that the peak of synthesis and/or release of JH esterase from the fat body of last instar larvae occurred 4 days after ecdysis. These studies also showed that fat body from JH-treated larvae released much less enzyme than controls. Antiallatotropins, precocene 2 and ZR 2646 also reduced JH esterase levels in larvae, but ZR 2646 induced JH esterase in pupae. In starved larvae, JH esterase did not increase during the first five days. A minimum of 36 hr of feeding was necessary for the larval esterase activity to increase on schedule on day 4 of the last larval stadium. When day-l larvae were ligated behind the head or the prothorax, they had lower JH esterase levels and yet showed a slight increase in the enzyme when the larvae reached the age of 4 days. The significance of these results is discussed in relation to the possible control of esterase activity during metamorphosis.     

Effects of methoprene,a juvenile hormone analogue,on the larval and pupal stages of the yellow fever mosquito, Aedes aegypti

Exposure of early fourth-instar larvae of Aedes aegypti to the juvenile hormone analogue Altosid ZR15^® (methoprene) significantly increased the concentration of carbohydrates in the haemolymph of late fourth-instar larvae and reduced the haemolymph carbohydrate concentration of 24-h-old pupae relative to controls. Such treatment also effected a decline in haemolymph amino nitrogen levels of the pupal stage and a depletion of haemolymph proteins in late fourth-instar larvae as well as pupae. Two of nine protein fractions in the haemolymph of larvae were significantly depleted following methoprene treatment. Fourteen soluble protein fractions were present in the haemolymph of control pupae; two of these were missing from the pupae which were treated as larvae with methoprene. A further protein fraction, common to the haemolymph of both treated and control pupae, was significantly reduced in concentration as a consequence of exposure to methoprene. The juvenile hormone analogue impaired the capacity of the fat bodies of late fourth-instar larvae and pupae to synthesise proteins, resulting in a lowered concentration of fat body proteins. Glycogen levels in the fat bodies of treated larvae were significantly lower than in controls and glycogenolysis was suppressed due to an overall depletion of glycogen phosphorylase and, in pupae, a lowered ratio of active: inactive enzyme. The data are consistent with the proposition that the juvenile hormone analogue elicits neuroendocrinological changes in the target insect.     

Protein synthesis during salivary gland development,ageing, and cell death in Chironomus tentans larvae

Salivary gland protein synthesis in Chironomus tentans larvae was analyzed from the mid-third instar to larval pupation. Correcting for stage specific variations in the specific activity of the amino acid pool revealed a 30–40% reduction in the rate of protein synthesis during the larval moult. Except for a transient increase early in the fourth instar, this low rate of protein synthesis was maintained until the pharate pupal period when protein synthesis dramatically increased: maximum synthesis occurred in mid-pharate pupae with a subsequent decline correlating with gland autolysis and cell death at pupation. Each developmental period was characterized by a particular pattern of secretory protein synthesis: high and 35,000 daltons peptides were maximally synthesized only at particular larval stages, being reduced or absent in post-ecdysis, diapause and autolysing salivary glands.Except for the ecdysone puffs, and as otherwise previously noted (Clever, 1961, 1962), puffing activity during the peri-moult period remained relatively constant and did not decrease by the 30–40% predicted from the decreased rate of protein synthesis. The nearly complete loss in synthesis of the 35,000 daltons peptide was not accompanied by regression in any puff.     

Microbial synthesis of spherical nanosilver and nanogold for mosquito control

Biosynthesis of metal nanoparticles using microorganisms is an important area of research in nanobiotechnology, which is an emerging eco-friendly science of well-defined sizes, shapes and controlled monodispersity. The present study proposed a green process for the extracellular production of silver (Ag) and gold (Au) nanoparticles (NPs) using the soil fungi Chrysosporium keratinophilum and Verticillium lecanii. The synthesized NPs were formed fairly uniform with spherical shape determined by Transmission Electron Microscope (TEM) and confirmed by Scanning Electron Microscope (SEM). Elemental analysis on single particle was carried by EDX analysis. The results were further supported by UV-vis spectrophotometry. In addition, we have also investigated the effect of synthesized AgNPs and AuNPs against the larvae and pupae of Anopheles stephensi, Culex quinquefasciatus and Aedes aegypti. The efficacy test was performed at different concentrations for periods of different lengths by the probit analysis. The larvae and pupae of Cx. quinquefasciatus, An. stephensi and Ae. aegypti were found highly susceptible to the synthesized AgNPs than the AuNPs. The larvae of Cx. quinquefasciatus and Ae. aegypti were found to be more susceptible to the AgNPs and AuNPs synthesized using the C. keratinophilum and V. lecanii compared with the larvae of An. stephensi. The pupae of Ae. aegypti have shown higher mortality against the synthesized AgNPs than the pupa of Cx. quinquefasciatus, while no adverse effects could be observed in the pupa of An. stephensi. By this approach, it is suggested that this rapid synthesis of nanoparticles would be useful for developing a biological process for mosquito control.     

Cell-free Synthesis of Globulin by Developing Oat (Avena sativa L.) Seeds

The primary storage protein synthesized during oat (Avena sativa L.) groat development is a globulin. Polysomes were isolated from oat groats 12 days after anthesis. These polysomes directed the incorporation of radioactive amino acids into protein in a cell-free protein synthesis system containing wheat germ supernatant. The Mg(2+) optimum was 4 mm, the pH optimum was 6-8, and the amount of amino acid incorporation depended on polysome concentration. Incorporation of amino acids was linear for about 10 min and approached a maximum after 20 min. Using the initiation inhibitor, T-2 toxin, it was determined that about 36% of the amino acid incorporation was due to the initiation of new polypeptide chains. The in vitro product co-electrophoresed with authentic oat groat globulin on polyacrylamide-sodium dodecyl sulfate (SDS) gels. The cyanogen bromide peptides of the in vitro product partially corresponded with those from authentic globulin when electrophoresed on polyacrylamide-SDS gels. These data suggest that the in vitro product is primarily oat globulin. The polysome population was separated into membrane-bound and free polysomes. Membrane-bound polysomes synthesized about twice the amount of protein as did free polysomes. Products synthesized in vitro on both types of polysomes were essentially the same.