The purified yeast pre-mRNA splicing factor PRP2 is an RNA-dependent NTPase.

Unlike autocatalyzed self-splicing reactions, nuclear pre-mRNA splicing requires transacting macromolecules and ATP. A protein encoded by the PRP2 gene of Saccharomyces cerevisiae is required, in conjunction with ATP, for the first cleavage-ligation reaction of pre-mRNA splicing. In this study, we have purified two forms of the PRP2 gene product with apparent molecular weights of 100 kDa and 92 kDa, from a yeast strain overproducing the protein. Both proteins were indistinguishable in their ability to complement extracts derived from a heat-sensitive prp2 mutant. Furthermore, we show that the PRP2 protein is capable of hydrolyzing nucleoside triphosphates in the presence of single-stranded RNAs such as poly(U). However, purified PRP2 by itself did not unwind double-stranded RNA substrates. The fact that an RNA-dependent NTPase activity is intrinsic to PRP2 may account for the ATP requirement in the first catalytic reaction of pre-mRNA splicing.     

Alternate use of divergent forms of an ancient exon in the fructose-1,6-bisphosphate aldolase gene of Drosophila melanogaster.

The fructose-1,6-bisphosphate aldolase gene of Drosophila melanogaster contains three divergent copies of an evolutionarily conserved 3' exon. Two mRNAs encoding aldolase contain three exons and differ only in the poly(A) site. The first exon is small and noncoding. The second encodes the first 332 amino acids, which form the catalytic domain, and is homologous to exons 2 through 8 of vertebrates. The third exon encodes the last 29 amino acids, thought to control substrate specificity, and is homologous to vertebrate exon 9. A third mRNA substitutes a different 3' exon (4a) for exon 3 and encodes a protein very similar to aldolase. A fourth mRNA begins at a different promoter and shares the second exon with the aldolase messages. However, two exons, 3a and 4a, together substitute for exon 3. Like exon 4a, exon 3a is homologous to terminal aldolase exons. The exon 3a-4a junction is such that exon 4a would be translated in a frame different from that which would produce a protein with similarity to aldolase. The putative proteins encoded by the third and fourth mRNAs are likely to be aldolases with altered substrate specificities, illustrating alternate use of duplicated and diverged exons as an evolutionary mechanism for adaptation of enzymatic activities.     

Videomicroscopic recording of Pneumocystis carinii motion.

Videomicroscopy in combination with differential-contrast optics was used to study fresh preparations of Pneumocystis carinii from immunosuppressed rats. Certain spherical intracystic bodies appeared to move freely within the cyst wall. Flexing type movement was observed in intracystic ellipsoidal forms attached at a common point in the inner margin of the cyst wall. Greater movement was seen in non-attached thinner elongated forms. Possible extracellular trophic forms and movement were also identified. The movement of the morphological forms of P. carinii has been recorded in real time onto videotape. These initial observations suggest P. carinii is capable of movement and additional studies are under way to substantiate this possibility.     

Immunologic comparisons of Pneumocystis carinii strains obtained from rats, ferrets, and mice using convalescent sera from the same sources.

Pneumocystis carinii (Pc) infections were developed in animals immunosuppressed by dexamethasone treatment either from activation of latent infection (ferret) or trans-tracheal inoculation of Pc obtained from infected lungs of the homologous species (rat, mouse). Convalescent antisera were obtained by stopping dexamethasone treatment after 2-4 wk and allowing 5-8 wk for recovery. Parasites from infected lungs were purified by differential filtration, solubilized in loading buffer, subjected to sodium dodecyl sulfate- polyacrylamide gel electrophoresis, and blotted to polyvinylidene fluoride sheets for Western analysis. Antisera from each animal species were reacted on Western blots of antigens from rat, ferret, and mouse. Each combination of antigen and antibody from the same species of animal showed reaction with 5 or more bands of Pc antigen. Convalescent mouse antibody did not react with rat or ferret antigens. Convalescent rat antibody reacted with a mouse antigen at about 66 kDa but not with ferret antigen, and convalescent ferret antibody showed minimal, probably non-specific reactions with both rat and mouse antigens. Variations in reactions indicate antigenic differences in Pc strains infecting these animals.     

A case of anisakiasis causing intestinal obstruction

A 31-year old salesman living in Seoul developed suddenly abdominal pain due to intestinal obstruction. Exploratory laparotomy exhibited segmental jejunal cellulitis caused by penetrating Anisakis larva. The patient had eaten raw fish. The typical history of intestinal anisakiasis was presented with a short review of Korean patients of anisakiasis.     

A single-stage two-flap method of total ear reconstruction

A single-stage two-flap method of total ear reconstruction in congenital microtia is reported. This method was derived from the one-stage reconstruction described by Song and Song. Two flaps defined by vascular basis were elevated on the mastoid area: the superficial skin flap supplied mainly by subcutaneous pedicled arteriole perforators from the posterior auricular artery and the deeper axial-pattern fascial flap including the posterior auricular artery itself. The ear framework, exaggeratedly carved using autologous rib cartilage, could be inserted easily between the two flaps, simultaneously producing the auriculocephalic angle and the conchal wall. Intraoperative expansion of the skin flap and postoperative external ear molding also were performed to create aesthetically pleasing ears.     

Anaerobic degradation of toluene by a denitrifying bacterium

A denitrifying bacterium, designated strain T1, that grew with toluene as the sole source of carbon under anaerobic conditions was isolated. The type of agar used in solid media and the toxicity of toluene were determinative factors in the successful isolation of strain T1. Greater than 50% of the toluene carbon was oxidized to CO2, and 29% was assimilated into biomass. The oxidation of toluene to CO2 was stoichiometrically coupled to nitrate reduction and denitrification. Strain T1 was tolerant of and grew on 3 mM toluene after a lag phase. The rate of toluene degradation was 1.8 mumol min-1 liter-1 (56 nmol min-1 mg of protein-1) in a cell suspension. Strain T1 was distinct from other bacteria that oxidize toluene anaerobically, but it may utilize a similar biochemical pathway of oxidation. In addition, o-xylene was transformed to a metabolite in the presence of toluene but did not serve as the sole source of carbon for growth of strain T1. This transformation was dependent on the degradation of toluene.