Determining the order of genes,centromeres, and rearrangement breakpoints inNeurospora by tests of duplication coverage

Nontandem segmental duplications provide a useful alternative to conventional recombination mapping for determining gene order in a haploid organism such asNeurospora. When an insertional or terminal rearrangement is crossed by Normal sequence, a class of progeny is produced that have a precisely delimited chromosome segment duplicated. In such Duplication progeny, a recessive gene in the Normal-sequence donor chromosome may or may not be masked (“covered”) by its dominant wild-type allele in the translocation-sequence recipient chromosome. Coverage depends upon whether the gene in question is left or right of the rearrangement breakpoint. The recessive gene will be heterozygous and covered (not expressed) if its locus is within the duplicated segment, but it will be haploid and expressed if the locus is outside the segment. Not only genes but also centromeres can be mapped by means of duplications, because genes included in. the same viable duplication must reside in the same chromosome arm. - Numerous sequences in the current genetic maps ofN. crassa have been determined using duplications. Gene order in the albino region and in the centromere region of linkage group I provide examples. Over 50 insertional or terminal rearrangements are available from which nontandem duplications of defined content can be obtained at will; collectively these cover about 75% of the genome. - Intercrosses between partially overlapping chromosome rearrangements also produce Duplication progeny containing two copies of regions between the breakpoints. The 180 mapped reciprocal translocations and inversions include numerous overlapping combinations that can be used for duplication mapping.     

Desorption chemical ionization mass spectrometry of 1-O-alkyl-2-O-acetyl-sn-glycero-3-phosphocholines and analogues

Ammonia desorption chemical ionization of ether-linked phospholipids of the type 1-O-alkyl-2-O-acetyl-sn-glycero-3-phosphocholine (platelet-activating factors) and a series of analogues revealed a systematic fragmentation pattern that is characteristic for these compounds. The predominant ions included the protonated molecular ion and a series of fragments derived from the molecular ion having the following nominal mass losses: MH-14, MH-42, MH-59, and MH-183. Deuterated ammonia was used to elucidate the nature of several fragments. In addition, desorption chemical ionization was used to quantitate 1-O-hexadecyl-2-O-acetyl-sn-glycero-3-phosphocholine at the nanogram/sample level.     

Platelet-activating factor induces the expression of early pregnancy factor activity in female mice

When synthetic platelet-activating factor (PAF, 1-0-alkyl-2-acetyl-sn-glycero-3-phosphocholine) was injected into mature female mice during dioestrus, pro-oestrus or oestrus, it induced the expression of early pregnancy factor (EPF) activity in the sera of these animals within 1 h of injection. The sera of similarly injected males, metoestrous or immature females did not display any EPF activity. The results suggest that embryo-derived PAF may be the ovum factor responsible for triggering the generation of serum EPF activity during the preimplantation stages of pregnancy.     

Variants of BALB/c 3T3 cells lacking complex gangliosides retain a fibronectin matrix and spread normally on fibronectin-coated substrates

Evidence has accumulated that di- and trisialogangliosides are involved in the interaction of cells with fibronectin. We have therefore tested the ability of variants of BALB/c 3T3 deficient in such gangliosides to organize a fibronectin matrix and to spread on fibronectin-coated substrates. Whereas BALB/c 3T3 cells contained gangliosides GM3, GM1, and GD1a, direct chemical analysis showed that five out of six variants isolated contained no detectable GD1a. By the overlaying of thin layer chromatograms of cellular gangliosides with 125I-cholera toxin, these variants were also found to lack ganglioside GM1. In contrast, the sialogalactoprotein profile of these cells, analyzed using an 125I-ricin/SDS polyacrylamide gel overlay technique, was similar to that of the parent cell line. All variants organized an extensive fibronectin matrix comparable to that of BALB/c 3T3, as shown using either immunofluorescence or lactoperoxidase-catalyzed iodination. The variants could also spread on fibronectin-coated substrates and adopt a morphology similar to that of BALB/c 3T3 cells, with little or no difference in the concentration of fibronectin required for 50% cell spreading. Cell spreading of the variants was accompanied by the formation of focal contacts and microfilament bundles, in a manner closely resembling that seen with BALB/c 3T3 cells. Treatment of BALB/c 3T3 cells with neuraminidase, which converts much of the cellular GD1a to GM1, did not affect cell spreading on fibronectin. The results clearly demonstrate that complex gangliosides are not essential for retention of a fibronectin matrix or for spreading on fibronectin-coated substrates.     

Mechanism of interferon action. Expression of vesicular stomatitis virus G gene in transfected COS cells is inhibited by interferon at the level of protein synthesis

The effect of interferon on the expression of the vesicular stomatitis virus glycoprotein G gene was examined in simian COS cells transfected with the expression vector pSVGL containing the G gene under the control of the SV40 late promoter. When COS cells were treated with interferon 24 h after transfection, the synthesis of vesicular stomatitis virus G protein was inhibited by about 80% as compared to that in untreated controls. By contrast, under the same conditions, neither the plasmid copy number nor the G gene mRNA levels were detectably affected by interferon treatment. Likewise, the synthesis of simian virus 40 large T-antigen was not inhibited by interferon treatment of transfected COS cells even though the synthesis of vesicular stomatitis virus G protein was markedly inhibited. The residual G protein synthesized in transfected, interferon-treated COS cells appeared to be normally glycosylated.     

Release of Endogenous Amino Acids from Striatal Neurons in Primary Culture

Following partial purification, the characteristics of a cytosol protein kinase were investigated. The protein kinase was purified by ammonium sulfate precipitation and diethylaminoethyl-cellulose, ATP-agarose, and hydroxyapatite chromatography. Analysis of the purified protein kinase preparation by polyacrylamide gel electrophoresis revealed three major protein bands. The cytosol protein kinase was purified approximately 442-fold, as calculated from the cyclic nucleotide independent protein kinase activity in the 40,000 g supernatant. The activity of the kinase was found to be independent of either cyclic AMP or cyclic GMP. Moreover, the kinase activity was unaffected by the addition of the endogenous protein kinase inhibitor, or the regulatory subunit from the type II cyclic AMP-dependent protein kinase from bovine heart. The molecular weight of the enzyme was determined to be 95,000 by Sephadex G-200 gel filtration. The activity of the kinase was increased approximately twofold in the presence of 10 microM Ca+2 and calmodulin. This increase was reversed by the addition of EGTA. The subcellular distribution of the protein kinase was also examined. The soluble fraction from nerve terminal was found to have the highest concentration of the kinase activity.     

Wright (a + b-) human erythrocytes and Plasmodium falciparum malaria

We find Wr(a + b-) erythrocytes of donor M. Fr., which appear to carry a rare glycophorin A variant, to be fully susceptible to invasion by nine isolates of Plasmodium falciparum. Thus we fail to confirm the previous publication on the refractoriness of these erythrocytes. In addition the serum of donor M. Fr., which is known to contain anti-Wrb directed against an epitope located on glycophorin A in close proximity to the erythrocyte membrane, was not found to inhibit P. falciparum invasion of blood group O Rh- red blood cells. Despite this, different lines of evidence still indicate that glycophorin A is one of the receptors for erythrocyte invasion by P. falciparum. The Wrb epitope, however, does not appear to represent a distinct receptor site, which is in contrast to previous suggestions.     

DNA sequence patterns in human,mouse, and rabbit immunoglobulin kappa-genes

Summary DNA sequences of the human, mouse, and rabbit immunoglobulin kappa-gene (J-C regions) are compared with respect to various DNA patterns, including dyad symmetry pairings, runs of nucleotides, repeat clusters, and repeats that occur with unusually high frequency. The significant dyad symmetry pairings within each of the sequences emphasize the two control-enhancer elements of the J₅-C intron. Dyad symmetry pairs between the J-C region and a number of kappa variable (V)-gene domains suggest differences in the affinities between the V and J segments. It is the consensus heptamer rather than the consensus nonamer that embodies the longest V-J dyad symmetry combinations. In the rabbit there are long runs and repeat clusters of the sequences that identify regions of high duplication; these regions are absent in the human and mouse sequences. High-frequency oligonucleotides feature the consensus nonamer 5 to the J segments, especially in the mouse sequence.     

Temporal fluctuations of silver,copper and zinc in the bivalve Macoma balthica at five stations in South San Francisco Bay

Concentrations of Cu, Ag and Zn were measured in the soft tissues of the estuarine bivalve Macoma balthica in South San Francisco Bay at near-monthly intervals for periods of two to three years at four stations, and eight years at a metal-enriched station. The amplitude and frequency of fluctuations differed among stations and among metals. Fluctuations were greatest at stations with the greatest metal enrichment and with the least dilution and flushing of wastes. A consistent seasonal pattern of fluctuation in Cu and Ag concentrations was evident in M. balthica at the metal-enriched station. These seasonal changes in tissue metal concentrations appeared to be affected by metal inputs, hydrologic processes that may affect both metal concentrations and bioavailability, and seasonal changes in the weight of the bivalve. The contributions of each of these interacting factors could not be determined quantitatively. At the metal-enriched station significant variation in the amplitude of seasonal fluctuations was also evident from year to year. Interpretation of metal concentrations in bivalves from estuaries will require careful consideration of the processes which affect metal dynamics in these complex environments.