An Autosomal Gene That Affects X Chromosome Expression and Sex Determination in CAENORHABDITIS ELEGANS

Recessive mutant alleles at the autosomal dpy-21 locus of C. elegans cause a dumpy phenotype in XX animals but not in XO animals. This dumpy phenotype is characteristic of X chromosome aneuploids with higher than normal X to autosome ratios and is proposed to result from overexpression of X-linked genes. We have isolated a new dpy-21 allele that also causes partial hermaphroditization of XO males, without causing the dumpy phenotype. All dpy-21 alleles show hermaphroditization effects in XO males that carry a duplication of part of the X chromosome and also partially suppress a transformer (tra-1) mutation that converts XX animals into males. Experiments with a set of X chromosome duplications show that the defects of dpy-21 mutants can result from interaction with several different regions of the X chromosome. We propose that dpy-21 regulates X chromosome expression and may be involved in interpreting X chromosome dose for the developmental decisions of both sex determination and dosage compensation.     

Cyclic AMP levels during induction and repression of cellulase biosynthesis in Thermomonospora curvata.

Specific cellulase production rates (SCPR) were compared with intracellular cyclic AMP (cAMP) levels in the thermophilic actinomycete, Thermomonospora curvata, during growth on several carbon sources in a chemically defined medium. SCPR and cAMP levels were 0.03 U (endoglucanase [EG] units) and 2 pmol per mg of dry cells, respectively, during exponential growth on glucose. These values increased to about 6 and 25, respectively, during growth on cellulose. Detectable EG production ceased when cAMP levels dropped below 10. Cellobiose (usually considered to be a cellulase inducer) caused a sharp decrease in cAMP levels and repressed EG production when added to cellulose-grown cultures. 2-deoxy-D-glucose, although nonmetabolizable in T. curvata, depressed cAMP to levels observed with glucose, but unlike glucose, the 2DG effect persisted until cells were washed and transferred to fresh medium. SCPR values and cAMP levels in cells grown in continuous culture under conditions of cellobiose limitation were markedly influenced by dilution rate (D). The maxima for both occurred at D = 0.085 (culture generation time of 11.8 h). When D was held constant and cellobiose concentration was increased over a 14-fold range to support higher steady state population levels, SCPR values decreased about fivefold, indicating that extracellular catabolite accumulation may be a factor in EG repression. The role of cAMP in the mechanism of this repression appears to be neither simple nor direct, since large changes (up to 200-fold) in SCPR accompany relatively small changes (10-fold) in cellular cAMP levels.     

Proline excretion by Escherichia coli K12

Proline excretion from proline overproducing strains of E. coli K12 has been studied as a model chemical production system. We have isolated proline overproducing mutants of E. coli and have shown that uncontrolled synthesis is not sufficient to cause excretion of this amino acid. An episomal mutation causing proline over production has been introduced into a series of otherwise isogenic strains that bear well defined, chromosomal lesions affecting the active uptake and catabolism of L-proline. A syntropism test reveals that L-proline is excreted by overproducing strains only if transport and/or catabolism are impaired. Dansyl derivatization and chromatographic analysis of culture supernatants shows that proline is the only amino acid excreted. Batch cultures of an excreting strain in an amino acid production medium yield culture supernatants containing 1 g proline/L, whereas no proline is detectable in supernatants derived from cultures of an overproducing strain with normal transport and catabolic activities. These data reveal that genetic lesions eliminating active uptake can be used to specifically enhance metabolite excretion.     

Production of auto-anti-idiotypic antibody during the normal immune response: IX. Characteristics of the auto-anti-idiotype antibody and its production

Using hapten-reversible inhibition of plaque formation as an assay for auto-anti-idiotype antibody (anti-Id) and as a means for following idiotype (Id) expression, we have obtained evidence that following immunization with trinitrophenyl (TNP) conjugates (a) there are differences in Id expression in the anti-TNP antibody response to different TNP conjugates although there is some overlap; (b) different strains, although showing some differences in Id expression, tend to produce cross-reactive Ids, thus no obvious allotype linked inheritance of Id expression is observed in this heterogeneous immune response; (c) the auto-anti-Id produced following immunization with TNP-Brucella abortus or TNP-Ficoll tends to be of the IgG_2a and IgG_2b isotypes.     

Non-targeted mutagenesis of unirradiated lambda phage in Escherichia coli host cells irradiated with ultraviolet light

Non-targeted mutagenesis of lambda phage by ultraviolet light is the increase over background mutagenesis when non-irradiated phage are grown in irradiated Escherichia coli host cells. Such mutagenesis is caused by different processes from targeted mutagenesis, in which mutations in irradiated phage are correlated with photoproducts in the phage DNA. Non-irradiated phage grown in heavily irradiated uvr+ host cells showed non-targeted mutations, which were 3/4 frameshifts, whereas targeted mutations were 2/3 transitions. For non-targeted mutagenesis in heavily irradiated host cells, there were one to two mutant phage per mutant burst. From this and the pathways of lambda DNA synthesis, it can be argued that non-targeted mutagenesis involves a loss of fidelity in semiconservative DNA replication. A series of experiments with various mutant host cells showed a major pathway of non-targeted mutagenesis by ultraviolet light, which acts in addition to "SOS induction" (where cleavage of the LexA repressor by RecA protease leads to din gene induction): (1) the induction of mutants has the same dependence on irradiation for wild-type and for umuC host cells; (2) a strain in which the SOS pathway is constitutively induced requires irradiation to the same level as wild-type cells in order to fully activate non-targeted mutagenesis; (3) non-targeted mutagenesis occurs to some extent in irradiated recA recB cells. In cells with very low levels of PolI, the induction of non-targeted mutagenesis by ultraviolet light is enhanced. We propose that the major pathway for non-targeted mutagenesis in irradiated host cells involves binding of the enzyme DNA polymerase I to damaged genomic DNA, and that the low polymerase activity leads to frameshift mutations during semiconservative DNA replication. The data suggest that this process will play a much smaller role in ultraviolet mutagenesis of the bacterial genome than it does in the mutagenesis of lambda phage.     

A low-cost antifoam additive for agar-based culture media

A highly effective, low-cost silicone-based antifoam emulsion is described which, at a final concentration of 100 ppm, prevents bubble formation during the preparation and dispensing of agar media. The compound is heat-stable, has a long shelf-life and offers considerable savings in cost by reduction in wastage of time and materials. It has no demonstrable deleterious effect on the growth of a wide range of pathogenic and commensal bacteria, or on antibiotic disc susceptibility testing, when examined using a range of different media.     

The role of H-2 and non-H-2 antigens and genes in the rejection of murine cardiac allografts

Genes of the major histocompatibility complex (MHC) in the mouse (H-2 complex) have been shown to be an important factor in determining the immune responsiveness of various strains of mice to isolated antigens (e. g., lysozyme). The role of MHC genes in controlling the responsiveness of mice to multiple alloantigens is less well-defined, and although non-MHC genes have been shown to be important in determining responsiveness in some systems (e. g., haptens), they have not been demonstrated as yet to influence the rejection of vascularized organ allografts. In this study, the responsiveness of mice to vascularized cardiac allografts transplanted across well-defined major (H-2) and minor (non-H-2) histocompatibility barriers was investigated using congenic mice in 32 different donor/recipient combinations. The results show that both H-2 and non-H-2 gene products can act as target alloantigens for rejection. At the responder level, they may interact to effect responsiveness of a recipient strain to multiple alloantigens. In no case in this study has any one gene or group of genes been found to confer universal high or low responder status.     

Binding of inositol trisphosphate by a liver microsomal fraction.

To test the hypothesis that inositol trisphosphate (InsP3) mediates adaptation and excitation in invertebrate photoreceptors, we measured its formation on a rapid time scale in squid retinas. For squid, excitation and adaption occurs within 0.1 and 1-2 s respectively. We could detect an elevation in InsP3 within 200 ms of a bright flash. This increase is about 240% over dark basal levels and is maintained for at least 2 min after a flash. The increase probably occurs in the photoreceptors, which are the only neurons in squid retinas. Analysis by h.p.l.c. indicates that the light-regulated isomer is Ins(1,4,5)P3, which is formed by the hydrolysis of phosphatidylinositol bisphosphate (PtdInsP2).     

Coypu insulin. Primary structure, conformation and biological properties of a hystricomorph rodent insulin.

Insulin from a hystricomorph rodent, coypu (Myocaster coypus), was isolated and purified to near homogeneity. Like the other insulins that have been characterized in this Suborder of Rodentia, coypu insulin also exhibits a very low (3%) biological potency, relative to pig insulin, on lipogenesis in isolated rat fat-cells. The receptor-binding affinity is significantly higher (5-8%) in rat fat-cells, in rat liver plasma membranes and in pig liver cells, indicating that the efficacy of coypu insulin on receptors is about 2-fold lower than that of pig insulin. The primary structures of the oxidized A- and B-chains were determined, and our sequence analysis confirms a previous report [Smith (1972) Diabetes 21, Suppl. 2, 457-460] that the C-terminus of the A-chain is extended by a single residue (i.e. aspartate-A22), in contrast with most other insulin sequences, which terminate at residue A21. In spite of a large number of amino acid substitutions (relative to mammalian insulins), computer-graphics model-building studies suggest a similar spatial arrangement for coypu insulin to that for pig insulin. The substitution of the zinc-co-ordinating site (B10-His----Gln) along with various substitutions on the intermolecular surfaces involved in the formation of higher aggregates are consistent with the observation that this insulin is predominantly 'monomeric' in nature. The c.d. spectrum of coypu insulin is relatively similar to those of casiragua insulin and of bovine insulin at low concentration.