scholarly journals Fat body: a site of hemoglobin synthesis in Chironomus thummi (diptera).

1976 ◽  
Vol 69 (2) ◽  
pp. 264-274 ◽  
Author(s):  
G Bergtrom ◽  
H Laufer ◽  
R Rogers
Keyword(s):  
Amino Acids ◽  
Salivary Glands ◽  
Culture Medium ◽  
Aminolevulinic Acid ◽  
Fat Body ◽  
Hemoglobin Synthesis ◽  
A Site ◽  
Coomassie Brilliant ◽  
Chironomus Thummi

Fourth instar larvae of Chironomus thummi were permitted to incorporate labeled amino acids and/or sigma-aminolevulinic acid (sigma-ALA) in vivo and in organ culture. The products secreted into the hemolymph or into the culture medium were examined by acrylamide gel electrophoresis. Nine electrophoretic bands can be resolved as hemoglobins without staining. When gels are sliced for scintillation counting, incorporated amino acids and sigma-ALA are shown to be associated primarily with the same nine hemoglobin bands, suggesting that hemoglobins are assembled and secreted. Staining of gels with Coomassie brilliant blue reveals that there are several bands in addition to the visible hemoglobins. These bands incorporate amino acids, but not sigma-ALA, suggesting that they are non-heme proteins. The results of culturing isolated salivary glands, gut, and fat body demonstrate that the fat body is the major site of hemoglobin synthesis and secretion. Labeled products of the gut represent about 5% of the total hemoglobins produced by the tissues, while no hemoglobins are produced by the salivary glands. Although nine hemoglobins are visibly resolved on gels, labeling techniques reveal as many as 14 hemoglobins. This is the first demonstration of hemoglobin synthesis by specific tissues in culture in an invertebrate.

scholarly journals In Vivo Identification of Intermediate Stages of the DNA Inversion Reaction Catalyzed by the Salmonella Hin Recombinase

Genetics ◽  
1998 ◽  
Vol 149 (4) ◽  
pp. 1649-1663
Author(s):  
Oliver Z Nanassy ◽  
Kelly T Hughes
Keyword(s):  
Amino Acids ◽  
Biochemical Analysis ◽  
Mutational Analysis ◽  
Recombination Reaction ◽  
Recombination Site ◽  
Dna Inversion ◽  
Hin Recombinase ◽  
One Step ◽  
A Site

Abstract The Hin recombinase catalyzes a site-specific recombination reaction that results in the reversible inversion of a 1-kbp segment of the Salmonella chromosome. The DNA inversion reaction catalyzed by the Salmonella Hin recombinase is a dynamic process proceeding through many intermediate stages, requiring multiple DNA sites and the Fis accessory protein. Biochemical analysis of this reaction has identified intermediate steps in the inversion reaction but has not yet revealed the process by which transition from one step to another occurs. Because transition from one reaction step to another proceeds through interactions between specific amino acids, and between amino acids and DNA bases, it is possible to study these transitions through mutational analysis of the proteins involved. We isolated a large number of mutants in the Hin recombinase that failed to carry out the DNA exchange reaction. We generated genetic tools that allowed the assignment of these mutants to specific transition steps in the recombination reaction. This genetic analysis, combined with further biochemical analysis, allowed us to define contributions by specific amino acids to individual steps in the DNA inversion reaction. Evidence is also presented in support of a model that Fis protein enhances the binding of Hin to the hixR recombination site. These studies identified regions within the Hin recombinase involved in specific transition steps of the reaction and provided new insights into the molecular details of the reaction mechanism.

Drosophila transcriptional repressor protein that binds specifically to negative control elements in fat body enhancers

1992 ◽  
Vol 12 (9) ◽  
pp. 4093-4103
Author(s):  
D Falb ◽  
T Maniatis
Keyword(s):  
Binding Site ◽  
Fat Body ◽  
Regulatory Element ◽  
Negative Control ◽  
Nuclear Extracts ◽  
Eukaryotic Proteins ◽  
Adh Gene ◽  
A Site

Expression of the Drosophila melanogaster Adh gene in adults requires a fat body-specific enhancer called the Adh adult enhancer (AAE). We have identified a protein in Drosophila nuclear extracts that binds specifically to a site within the AAE (adult enhancer factor 1 [AEF-1]). In addition, we have shown that AEF-1 binds specifically to two other Drosophila fat body enhancers. Base substitutions in the AEF-1 binding site that disrupt AEF-1 binding in vitro result in a significant increase in the level of Adh expression in vivo. Thus, the AEF-1 binding site is a negative regulatory element within the AAE. A cDNA encoding the AEF-1 protein was isolated and shown to act as a repressor of the AAE in cotransfection studies. The AEF-1 protein contains four zinc fingers and an alanine-rich sequence. The latter motif is found in other eukaryotic proteins known to be transcriptional repressors.

scholarly journals Novel model for the in vivo study of central nervous system infection due to Acanthamoeba spp. (T4 genotype)

2009 ◽  
Vol 58 (4) ◽  
pp. 503-508 ◽  
Author(s):  
Parisa Nakhostin Mortazavi ◽  
Graham Goldsworthy ◽  
Ruth Kirk ◽  
Naveed Ahmed Khan
Keyword(s):  
Culture Medium ◽  
Fat Body ◽  
Central Nervous System Infection ◽  
Nutrient Agar ◽  
Migratory Locust ◽  
Mature Adult ◽  
Faecal Pellets ◽  
Sterile Culture ◽  
First Time

In this study it was shown for what is believed to be the first time that the African migratory locust can be used as a model for the study of Acanthamoeba pathogenesis. Mature adult locusts were injected intra-abdominally with 10 μl suspension of 106 Acanthamoeba (a clinical isolate of the T4 genotype) in culture medium, or with the same volume of sterile culture medium. Locusts injected with Acanthamoeba showed significant weight loss and reduced production of faeces compared with control locusts. Furthermore, injection of amoebae killed all of the locusts within 17 days at room temperature, although the speed of kill was temperature and dose dependent. When samples of faecal pellets and various tissues of infected locusts were cultured on non-nutrient agar plates containing bacterial lawns, live amoebae were recovered from haemolymph, flight muscle and fat body samples, but not from faeces. When brains dissected from locusts were incubated with an anti-amoebic drug (100 μM chlorhexidine) to kill extracellular amoebae, and then washed, homogenized and cultured on bacteria-seeded non-nutrient agar plates, only lysates from amoebae-infected locusts were positive for Acanthamoeba. This strongly suggests that amoebae invade the locust brain and, indeed, trophozoites of Acanthamoeba could be identified within the brain in histological sections of brains from infected locusts, but not from uninfected locusts. These findings support the view that locusts can be used as a model for the study of Acanthamoeba pathogenesis in vivo.

scholarly journals Vitellogenesis in the stick insect Carausius morosus I. Specific protein synthesis during ovarian development

1980 ◽  
Vol 46 (1) ◽  
pp. 1-16
Author(s):  
F. Giorgi ◽  
F. Macchi
Keyword(s):  
Amino Acids ◽  
Molecular Weight ◽  
Protein Synthesis ◽  
Gel Electrophoresis ◽  
Fat Body ◽  
Polyacrylamide Gel Electrophoresis ◽  
Stick Insect ◽  
Polyacrylamide Gel ◽  
Carausius Morosus

Vitellogenesis in the stick insect Carausius morosus (Br.) has been studied with the goal of identifying vitellogenin in various tissues. Following exposure to in vivo to radioactive amino acids, oocytes in the medium size range are labelled with a minimum delay of 6 h after the time of injection. Incorporation of radioactivity under these conditions is shown to depend upon accumulation of proteins rather than on a differential rate of protein synthesis in succeeding stages of oogenesis. By immunochemical analyses, it is shown that at least two antigens are common to both haemolymph and ovary and that one of these is also present in the fat body. Both antigens are labelled during exposure to radioactive amino acids. When analysed by the SDS polyacrylamide gel electrophoresis, extracts from both haemolymph and ovary appear to share a number of protein fractions which range in molecular weight from 40 000 to 200 000 Daltons. The labelling pattern exhibited by these fractions is clearly indicative of a protein transfer from the fat body to the oocyte. Fat body cultured in vivo for up to 4 h releases a major macromolecular complex in the external medium. The latter has been identified as vitellogenin by both immuno-precipitation assay and SDS polyacrylamide gel electrophoresis. The protein which is synthesized and secreted under these conditions results from the processing of a protein complex of higher molecular weight.

scholarly journals Measurement of average decoding rates of the 61 sense codons in vivo

eLife ◽  
2014 ◽  
Vol 3 ◽  
Author(s):  
Justin Gardin ◽  
Rukhsana Yeasmin ◽  
Alisa Yurovsky ◽  
Ying Cai ◽  
Steve Skiena ◽  
...  
Keyword(s):  
Amino Acids ◽  
Ribosome Profiling ◽  
High Coverage ◽  
Peptide Bonds ◽  
Synonymous Codons ◽  
Specific Effects ◽  
Exit Tunnel ◽  
A Site ◽  
Novel Algorithm

Most amino acids can be encoded by several synonymous codons, which are used at unequal frequencies. The significance of unequal codon usage remains unclear. One hypothesis is that frequent codons are translated relatively rapidly. However, there is little direct, in vivo, evidence regarding codon-specific translation rates. In this study, we generate high-coverage data using ribosome profiling in yeast, analyze using a novel algorithm, and deduce events at the A- and P-sites of the ribosome. Different codons are decoded at different rates in the A-site. In general, frequent codons are decoded more quickly than rare codons, and AT-rich codons are decoded more quickly than GC-rich codons. At the P-site, proline is slow in forming peptide bonds. We also apply our algorithm to short footprints from a different conformation of the ribosome and find strong amino acid-specific (not codon-specific) effects that may reflect interactions with the exit tunnel of the ribosome.

Aufnahme von 22Na in die Zellkerne der Speicheldrüsen von Chironomus thummi

1966 ◽  
Vol 21 (3) ◽  
pp. 274-277 ◽  
Author(s):  
Markus Lezzi ◽  
Heinrich Kroeger
Keyword(s):  
Salivary Glands ◽  
Polytene Chromosomes ◽  
Inorganic Ions ◽  
Saturation Curve ◽  
Cell Nuclei ◽  
Chironomus Thummi ◽  
In Vitro Uptake

The in vivo and in vitro uptake of 22Na from the hemolymph into cell nuclei of larval salivary glands was measured and compared. The uptake of 22Na in vivo follows approximately a saturation curve. The respective in vitro curve has a much steeper slope during the first 8 min and this phase is followed by an interval during which the 22Na content of the nuclei decreases (8 to 16 min). After the 16th min it increases again to reach a nuclear 22Na content approximately twice as high as that of nuclei in vivo at 60 minutes. The uptake curves are discussed in relation to recent findings on the induction of puffs in polytene chromosomes by inorganic ions.

scholarly journals Drosophila transcriptional repressor protein that binds specifically to negative control elements in fat body enhancers.

1992 ◽  
Vol 12 (9) ◽  
pp. 4093-4103 ◽  
Author(s):  
D Falb ◽  
T Maniatis
Keyword(s):  
Binding Site ◽  
Fat Body ◽  
Regulatory Element ◽  
Negative Control ◽  
Nuclear Extracts ◽  
Eukaryotic Proteins ◽  
Adh Gene ◽  
A Site

Expression of the Drosophila melanogaster Adh gene in adults requires a fat body-specific enhancer called the Adh adult enhancer (AAE). We have identified a protein in Drosophila nuclear extracts that binds specifically to a site within the AAE (adult enhancer factor 1 [AEF-1]). In addition, we have shown that AEF-1 binds specifically to two other Drosophila fat body enhancers. Base substitutions in the AEF-1 binding site that disrupt AEF-1 binding in vitro result in a significant increase in the level of Adh expression in vivo. Thus, the AEF-1 binding site is a negative regulatory element within the AAE. A cDNA encoding the AEF-1 protein was isolated and shown to act as a repressor of the AAE in cotransfection studies. The AEF-1 protein contains four zinc fingers and an alanine-rich sequence. The latter motif is found in other eukaryotic proteins known to be transcriptional repressors.

Selective stimulation of puffing activity in vitro with a juvenile hormone analogue by polytene chromosomes in the salivary glands of Chironomus thummi (Diptera)

1989 ◽  
Vol 67 (10) ◽  
pp. 2528-2532
Author(s):  
X. Vafopoulou ◽  
C. G. H. Steel ◽  
H. Laufer
Keyword(s):  
Juvenile Hormone ◽  
Salivary Glands ◽  
Polytene Chromosomes ◽  
Hormone Analogue ◽  
Juvenile Hormone Analogue ◽  
Balbiani Rings ◽  
Chironomus Thummi ◽  
Stimulation Of

Salivary glands of Chironomus thummi prepupae were treated in vitro with various concentrations of the juvenile hormone analogue (JHA) methoprene and the puffing activities of Balbiani rings (BR) b and c were scored in cytological preparations of polytene chromosomes. In control cultures, both BRb and BRc regress rapidly. JHA treatment in vitro prevented regression at both these sites. In addition, BRb was found to expand within 60 min to a size two to three times larger than in control contralateral glands. The most effective concentration for stimulation of BRb and BRc in vitro was 10−7 M. In vivo treatment of prepupae with 10−7 M JHA also induced puffing activity in BRb and BRc to a degree similar to that observed following in vitro treatment. It is concluded that BRb, and perhaps also BRc, are juvenile hormone (JH) inducible chromosomal sites in Chironomus thummi salivary glands. This is the first report of chromosomal sites that are stimulated by JH in vitro in the absence of exogenous ecdysteroids. These findings support the view that JH may act at the gene level at separate loci from ecdysone.

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