Protease-Dead Separase Is Dominant Negative in the C. elegans Embryo

Separase is a protease that promotes chromosome segregation at anaphase by cleaving cohesin. Several non-proteolytic functions of separase have been identified in other organisms. We created a transgenic C. elegans line that expresses protease-dead separase in embryos to further characterize separase function. We find that expression of protease-dead separase is dominant-negative in C. elegans embryos, not previously reported in other systems. The C. elegans embryo is an ideal system to study developmental processes in a genetically tractable system. However, a major limitation is the lack of an inducible gene expression system for the embryo. We have developed two methods that allow for the propagation of lines carrying dominant-negative transgenes and have applied them to characterize expression of protease-dead separase in embryos. Using these methods, we show that protease-dead separase causes embryo lethality, and that protease-dead separase cannot rescue separase mutants. These data suggest that protease-dead separase interferes with endogenous separase function, possibly by binding substrates and protecting them from cleavage.     

Retrospective evaluation of gynecologic cytodiagnosis. II. Interlaboratory reproducibility as shown in rescreening large consecutive samples of reported cases

Two laboratories exchanged and rescreened a large sample of cases with cervicovaginal smears they had consecutively accessioned to examine the reproducibility of gynecologic cytodiagnosis under optimum conditions. At least a "working agreement" (diagnoses within +/- 1 category on a ten-category scale) was achieved in diagnoses of normal, benign reaction and squamous abnormality (from minimal dysplasia though invasive cancer) in 18,859 cases (96.8%), of endometrial abnormality in 21 cases (42%) and of "unsatisfactory" in 99 cases (20.7%). Larger differences occurred in greater than or equal to 30% of cases except in the categories of "normal" and "benign reaction," reaching a maximum of 82% for moderate dysplasia. Reexamining 382 cases decreased disagreement by category to the 20% to 65% range only in the five categories of dysplasia plus carcinoma in situ. Agreement was not predicated on the presence of endocervical cells or squamous metaplasia; the basis for "unsatisfactory" calls was not uniform. Comparison of the laboratories' diagnoses with referee diagnoses or, on 178 cases, with tissue diagnoses also demonstrated differences in diagnostic criteria.     

Regulation of intracellular pH by a neuronal homolog of the erythrocyte anion exchanger

We have isolated AE3, a novel gene expressed primarily in brain neurons and in heart. The predicted AE3 polypeptide shares a high degree of identity with the anion exchange and cytoskeletal binding domains of the erythrocyte band 3 protein. Expression of AE3 cDNA in COS cells leads to chronic cytoplasmic acidification and to chloride- and bicarbonate-dependent changes in intracellular pH, confirming that this gene product is an anion exchanger. Characterization of an AE3 mutant lacking the NH2-terminal 645 amino acids demonstrates that the COOH-terminal half of the polypeptide is both necessary and sufficient for correct insertion into the plasma membrane and for anion exchange activity. The NH2-terminal domain may play a role in regulating the activity of the exchanger and may be involved in the structural organization of the cytoskeleton in neurons.     

Octanoylation of the lipoyl domains of the pyruvate dehydrogenase complex in a lipoyl-deficient strain of Escherichia coli

The overexpression of a subgene encoding a hybrid lipoyl domain of the dihydrolipoamide acetyltransferase component of the pyruvate dehydrogenase complex of Escherichia coli has previously been shown to result in the formation of lipoylated and unlipoylated products. Overexpression of the same subgene in a lipoic acid biosynthesis mutant growing under lipoate-deficient conditions has now been shown to produce domains modified by octanoylation as well as unmodified domains. It was concluded from the mass of a lipoyl-binding-site peptide that the modification involves N6-octanoylation of the lysine residue (Lys244) that is normally lipoylated, and this was confirmed by the trypsin-insensitivity of the corresponding Lys244-Ala-245 bond, and the absence of modification in a mutant domain in which Lys244 is replaced by Gln. This novel protein modification raises interesting questions concerning the pathway of lipoic acid biosynthesis and the mechanism of enzyme lipoylation.     

Ankylosis of hock joints in group caged male B6C3F1 mice

Enlarged hock joints were observed during 1983 in B6C3F1 mice of chronic toxicity and carcinogenicity studies sponsored by the National Toxicology Program (NTP). Subsequently, approximately 9,500 B6C3F1 mice on 32 NTP chemical toxicity and carcinogenicity studies were evaluated for this condition by clinical examination. Group caged male B6C3F1 mice had thickening and reduced mobility of the hock joints at prevalences of 1.2% up to 6 months of age; 23% at 6 to 12 months of age; and 62% at 13 to 26 months of age. Group caged female B6C3F1 mice had a prevalence of 2% or less. Histologically, affected mice had periarticular exostoses on the bones of the hock joints, with formation of bony bridges around joints and deposition of new bone in joint spaces, resulting in partial or complete ankylosis. Individually caged male and female B6C3F1 mice were not affected. The cause of the ankylosis was not determined, but its occurrence in the NTP studies has been reduced by individual caging.     

The effect of grasses on the quality of transmitted radiation and its influence on the growth of white clover Trifolium repens

Summary Plants of white clover Trifolium repens were grown under the canopies of three grass species, Lolium perenne, Agrostis tenuis and Holcus lanatus, and under simulated canopies of black polythene and controls were exposed to unfiltered natural radiation. The canopies were adjusted so that they transmitted equal intensities of Photosynthetically Active Radiation (P.A.R.). The ratio of red to far red radiation () was unchanged under the black polythene canopies but was reduced under canopies of Lolium and Agrostis and even more so under Holcus. The effect of canopy filtered radiation on the growth of clover was greatly to reduced internode length, mean number of nodes, the number of branched nodes and the number of rooted nodes and greatly to increase petiole length. The effect of canopies of Holcus was greater than that of the other grass species both in its effect on and on the responses of the clover plant to its shade.     

A study on the cytoplasmic granules of the pericardial gland cells of some bivalve molluscs

The pericardial glands of three bivalve molluscs are composed of convoluted epithelium that appears as pouches on the auricles of Mytilus and as tubules in the connective tissue at the anterior-lateral sides of the pericardial cavity of Mercenaria and Anodonta. The pericardial gland cells are attached to each other by many randomly placed desmosome-like cell junctions and gap junctions. Belt-desmosomes that are characteristic of epithelial cells were not observed. The basal membrane of these cells is invaginated producing complex interdigitating cytoplasmic processes and filtration slits. The pericardial gland cells stain for the presence of iron with Prussian blue stain. Electron-dense and electron-lucent granules of various diameters are present in the cytoplasm. Many electron-dense granules contain ferritin-like particles in which the presence of iron has been demonstrated by microanalysis. It is suggested that these particles are the iron storage protein ferritin since they contain iron, and are water soluble, heat stable, and morphologically similar to mammalian ferritin. Ferritin particles are probably both synthesized and broken down by the pericardial gland cells; thus the pericardial gland cells may be involved in iron homeostasis in these molluscs.     

Syringe exchange schemes for drug users in England and Scotland

In 1987 experimental schemes for distributing injecting equipment to intravenous drug users to help prevent the spread of the human immunodeficiency virus were started by the government. After six months the schemes were found to have been reasonably successful in attracting clients but were less successful in keeping them. It has been shown that equipment can be distributed to drug users on an exchange basis. Many of the clients who were attracted to these schemes had had no treatment or other help for their drug problems.