The induction of autophagy in isolated insect fat body by β-ecdysone

The fat body in Calpodes undergoes sequential organelle specific autophagy as a first step in the cell remodeling process necessary for metamorphosis to the pupa. This autophagy begins at about 36 hr before pupation and coincides with a critical period after which an isolated abdomen will pupate without further influence from the prothoracic glands. This suggested that autophagy might be induced by ecdysone. Fat body taken before the critical period and cultured in a medium containing β-ecdysone undergoes autophagy. Fat body from the same animal maintained in hormone-free medium retains the pre-critical period morphology with no autophagy. Autophagy is therefore directly induced by β-ecdysone. Fat body taken soon after the critical period continues with the autophagic sequence in hormone-free medium. Therefore the entire autophagic sequence is induced and does not require the continuing presence of hormone. Protein storage granule formation and cell dissociation, which occur in fat body at metamorphosis, are also induced by β-ecdysone.     

Incidence of anencephaly and other major malformations when oestriol excretion is very low.

A study of 533 women with very low urinary oestriol excretion during the third trimester of pregnancy showed an incidence of major fetal malformations among their infants of 7-1% and a perinatal mortality rate of 14-6%. Thirteen of the malformations were cases of anencephaly, and 26 of the 78 perinatal deaths were due to or associated with major fetal malformations. The incidence of these complications was higher when maternal oestriol excretion was lower. Routine screening by urinary oestriol assay, with fetal radiography when values below 20-8 mumol/24 hours (6 mg/24 h) are detected is the most reliable method of detecting anencephaly before birth.     

The Degradation of Romanowsky-Type Blood Stains in Methanol

The oxidative demethylation of Romanowsky-type stains in methanol has been examined quantitatively with respect to its effect upon the staining of blood smears. Spectral changes in hound dye, observed through two color filters, have been measured for the nuclei and cytoplasm of segmented neutrophils and monocytes utilizing the LARC™ automated differential analyzer. Stain decomposition in methanol results in a large loss in staining intensity with little change in color. The loss in intensity has been correlated with the observed spectral changes in the degraded stain. High-performance liquid chromatographic analysis of degraded stain samples has shown the products of methanolic degradation to be different from those obtained in aqueous polychroming reactions. To maintain a stain of defined thiazine dye composition and thus defined staining properties, refrigeration is recommended.     

Secretion of trans-zeatin by Agrobacterium tumefaciens: A function determined by the nopaline Ti plasmid

Production of the plant cytokinin, trans-zeatin, by a number of strains of Agrobacterium tumefaciens was measured by a combination of traceenrichment, HPLC and radioimmunoassay and confirmed by mass spectrometry. Secretion of trans-zeatin into a culture medium is a constitutive function of those strains which harbor a nopaline Ti plasmid. Strains cured of the nopaline Ti plasmid and those which harbor octopine or agropine plasmids are non-producers. Reacquisition of nopaline plasmids by cured strains restores production.     

Receptor-mediated phagocytosis of zymosan is unaffected by some conditions which reduce macrophage lysosomal enzyme secretion

We have previously shown that several agents which interfere with binding of ligands to the mannose-glycoprotein receptor on macrophages can inhibit zymosan-induced lysosomal enzyme secretion. Here we show that mannose only reduces the association of zymosan with macrophages during the first hour of exposure; after longer periods of uptake no effect is detectable. We have previously shown that mannose reduces surface binding of zymosan, probably by interfering selectively with binding to the mannose receptor. The present inhibition of association of zymosan with macrophages during short exposures can be entirely explained by this reduction of binding. Macrophages must therefore internalize zymosan at sites in addition to the mannose receptor. In contrast to macrophages the murine macrophage-like cell line P388D1 is lacking the mannose-glycoprotein receptor. Accordingly we find that binding of zymosan to P388D1 is much slighter than to macrophages and is unaffected by mannose or mannose-6-phosphate. The spontaneous lysosomal enzyme secretion of P388D1 is also unaffected by mannose. The data on macrophages confirm our previous suggestion that agents interfering with the mannose receptor inhibit the induction of lysosomal enzyme secretion by acting directly on the receptor. The data on P388D1 cells support this assertion by excluding effects at later steps in the secretory pathway.     

Comparative study of the peptide composition of Complex III (quinol-cytochromec reductase)

A comparative study has been made on the subunits of Complex III from beef heart, rat liver, Neurospora, and baker's yeast mitochondria. All of the subunits of the beef heart enzyme were similar to the counterpart subunit in rat liver Complex III, both with respect to their apparent molecular weights on SDS-polyacrylamide gels and their proteolytic digestion maps obtained in the presence of S. subtilus V8 protease. In contrast, the subunits of Neurospora and yeast Complex III varied considerably from the mammalian enzyme, as well as between themselves, the only exception being cytochrome b (subunit III). Less variation was observed in the electron transport peptides (IV-V) of higher and lower eukaryotes than in those subunits (I, II, VI-VIII) for which no functions are known. However, the data imply that subunits I, II, and VI-VIII are bona fide members of the complex, and that their functions within the complex, although unknown, are also somewhat conserved. Finally, the low-molecular-weight subunits of rat liver cytochrome oxidase and Complex III were compared. They appear to contain no subunits in common, implying different roles for these peptides in the two complexes.     

Simian virus 40 large T antigen untwists DNA at the origin of DNA replication.

Simian virus 40 large tumor antigen (SV40 T antigen) untwists DNA at the SV40 replication origin. In the presence of ATP, T antigen shifted the average linking number of an SV40 origin-containing plasmid topoisomer distribution. The loss of up to two helical turns was detected. The reaction required the presence of the 64-base pair core origin of replication containing T antigen DNA binding site II; binding site I had no effect on the untwisting reaction. The presence of human single-stranded DNA binding protein (SSB) slightly reduced the degree of untwisting in the presence of ATP. ATP hydrolysis was not required since untwisting occurred in the presence of nonhydrolyzable analogs of ATP. However, in the presence of a nonhydrolyzable analog of ATP, the requirement for the SV40 origin sequence was lost. The origin requirement for DNA untwisting was also lost in the absence of dithiothreitol. The origin-specific untwisting activity of T antigen is distinct from its DNA helicase activity, since helicase activity does not require the SV40 origin but does require ATP hydrolysis. The lack of a requirement for SSB or ATP hydrolysis and the reduction in the pitch of the DNA helix by just a few turns at the replication origin distinguishes this reaction from the T antigen-mediated DNA unwinding reaction, which results in the formation of a highly underwound DNA molecule. Untwisting occurred without a lag after the start of the reaction, whereas unwound DNA was first detected after a lag of 10 min. It is proposed that the formation of a multimeric T antigen complex containing untwisted DNA at the SV40 origin is a prerequisite for the initiation of DNA unwinding and replication.