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The effects of T-2 toxin, HT-2 toxin, T-2 triol, T-2 tetraol, diacetoxyscirpenol, verrucarol, roridin A, and verrucarin A on the growth of yeast were studied. The influence of these trichothecenes on the growth of the yeasts varied markedly. Kluyveromyces fragilis was most sensitive to verrucarin A. The minimal inhibitory concentration and effective concentrations50 values for verrucarin A were 0.24 and 0.05 nmol/ml, respectively. T-2 triol, T-2 tetraol, and verrucarol did not inhibit growth at the highest concentration tested. These eight trichothecenes inhibited mitochondrial function in Saccharomyces cerevisiae. Verrucarin A had the greatest toxicity toward the mitochondria of this yeast. In general, the order of toxicity was verrucarin A > roridin A > T-2 toxin > diacetoxyscirpenol > HT-2 toxin > verrucarol ≥ T-2 triol ≥ T-2 tetraol. These results show that these trichothecenes have different reactivities toward intracellular target(s). [ABSTRACT FROM PUBLISHER]

For sorption studies, the saturation vapor pressure p 0 above an adsorbate is of great significance. For example, it is needed for the determination of the pore size distribution, the Laplace pressure, and the chemical potential. Above the bulk triple point, T 3 bulk , this pressure is identical with the saturation vapor pressure above the bulk liquid. However, below T 3 bulk , the correct value of p 0 ( T ) is controversial. Nanoconfined fluids exhibit a shift of the freezing and melting temperatures in comparison to the bulk state. Thus, the adsorbed fluid is supercooled in a certain temperature range below T 3 bulk . Here, we show that it is possible to determine the appropriate saturation vapor pressure above the nanoconfined supercooled liquid experimentally. For this purpose, we have performed sorption measurements with liquid argon in nanoporous Vycor glass in the temperature range of the supercooled liquid and at temperatures above the bulk triple point. In order to determine the unknown and temperature-dependent saturation vapor pressure of the supercooled confined adsorbate, p 0 ( T ), we use the Kelvin equation relating this quantity to the pore radius, r P ( p 0 ), that is independent of temperature. The knowledge of the absolute values for the liquid-vapor surface tension of the supercooled adsorbate, γ lv ( T ), is not required. However, we presuppose that its dependence on the unknown vapor pressure, γ lv ( p 0 ), is bulk-like. Our results indicate that the saturation vapor pressure above the supercooled nanoconfined liquid corresponds to that above supercooled bulk argon (i.e., to the pressure obtained by an extension of the usual vaporization curve to T < T 3 bulk ). We expect that this method can be used for the determination of p 0 above other supercooled adsorbates., Competing Interests: The authors declare no competing financial interest., (Copyright © 2020 American Chemical Society.)

Liquids in nanoporous media are exposed to an adsorption-induced pressure, a consequence of the interaction with the pore surface. The smaller the pore diameter, d P , the higher the pressure at saturation and thus the bulk modulus of the confined liquid. Therefore, it has been proposed to use ultrasonic measurements on saturated nanoporous media for the determination of the average pore size. Here, we discuss the requirements for such an analysis. Although predictions for the size-dependent pore pressure and the liquid's modulus, K iso ( d P ), are based on isothermal simulations, an experimentalist studying the propagation of ultrasonic waves determines adiabatic moduli, K ad ( d P ). We show that the quantity relating adiabatic and isothermal moduli, the heat capacity ratio γ = c p / c v = K ad / K iso , exhibits a strong pressure dependence for many bulk liquids. In nanopores, this translates into a size-dependent γ( d P ), provided the confinement does not alter the heat capacity ratio. Disregarding this effect in the analysis of ultrasonic data would yield an underestimate of the isothermal modulus and thus an overestimate of the average pore size. For a correct analysis, an experimentalist thus needs to know the size dependence of three quantities: the isothermal modulus, adsorption-induced pressure, and heat capacity ratio., Competing Interests: The authors declare no competing financial interest.

The change of surface stress is an important quantity characterising the behaviour of nanoporous systems, however, it is difficult to assess experimentally. In this letter we develop and demonstrate an experimental method for the determination of adsorption-induced changes of the surface stress in nanoporous materials. With the aid of ultrasonic measurements we determine the dependence of the adsorbate's longitudinal modulus [Formula: see text] on the adsorption-induced normal pressure, [Formula: see text], which is exerted by the adsorbate on the porous matrix. From this dependence we deduce the normal pressure at saturation, [Formula: see text], and thereby changes of the surface stress [Formula: see text] at the interface between the solid matrix and the liquid adsorbate. For the model system of argon in nanoporous glass (pore radius [Formula: see text] nm) the ultrasonic method reveals a value for [Formula: see text] that is in very good agreement with the theoretical value known for the argon-silica interface. The disclosure of this experimental method and its application on other systems will enable a better understanding of the behaviour of adsorbates in nanoporous materials.

In this article we study the dependence of the sorption-induced deformation of nanoporous glass on the liquid-solid phase transition of adsorbed argon. During cooling we observe a continuous reduction of the expansion of the porous glass matrix caused by the adsorbate. The contraction is attended by a likewise continuous change of the adsorbed argon's phase state from liquid to solid. This simultaneous behavior evidences that the liquid-solid phase transition leads to a reduction of the pressure the adsorbate exerts on the pore walls. Furthermore, the study shows that small temperature changes can temporarily cause strong deformations of the porous material that decay in long time intervals of up to 1 week. We expect that our observations for the model system of argon and porous glass can be generalized to other systems. Consequently, this study will have implications when considering porous materials for applications, e.g., as a medium for storage.

Sorption of substances in pores generally results in a deformation of the porous matrix. The clarification of this effect is of particular importance for the recovery of methane and the geological storage of CO2. As a model system, we study the macroscopic deformation of nanoporous Vycor glass during the sorption of argon using capacitative measurements of the length change of the sample. Upon desorption we observe an unpredicted sharp contraction and re-expansion peak, which contains information on the draining mechanism of the porous sample. We have modified the theoretical model by Gor and Neimark1 to predict the sorption-induced deformation of (partly) filled porous samples. In this analysis, the contraction is attributed to a metastable or nonequilibrium configuration where a thin surface layer on the pore walls coexists with capillary bridges. Alternatively, pore blocking and cavitation during the draining of the polydisperse pore network can be at the origin of the deformation peak. The results are a substantial step toward a correlation between the spatial configuration of adsorbate, its interaction with the host material, and the resulting deformation.

We have measured sorption isotherms and determined the intrinsic longitudinal elastic modulus β(Ar,ads) of nanoconfined material via ultrasonic measurements combined with a special effective medium analysis. In the liquid regime the adsorbate only contributes to the measured effective properties when the pores are completely filled and the modulus is bulklike. At partial fillings its contribution is cancelled out by the high compressibility of the vapour phase. In contrast, at lower temperatures frozen argon as well as underlying liquid surface layers cause a linear increase of the effective longitudinal modulus upon filling. During sorption the contribution of the liquid surface layers near the pore wall β(Ar,surf) increases with the thickness of the solid layers reaching the bulk value β(Ar,liquid) only in the limit of complete pore filling. We interpret this effect as due to the gradual stiffening of the solid argon membrane. The measurements and their analysis show that longitudinal ultrasonic waves are well suited to the study of the elastic properties and liquid-solid phase transitions in porous systems. This method should also help to detect the influence of nanoconfinement on elastic properties in further research.

We have studied the phase transition of argon in completely filled mesopores. Our effective medium analysis of ultrasonic measurements clearly indicates a continuous phase transition of argon in completely filled pores over a broad temperature range of about 45 K. With decreasing temperature, the amount of frozen argon increases and below about 30 K all adsorbed argon (including the first few layers near the pore wall) is frozen with a shear modulus about equal to the bulk shear modulus. It is remarkable, that in a system showing such a pronounced confinement effect--a continuous phase transition over 45 K--the bulk properties are preserved. A comparison with temperature cycles with one and two adsorbed layers shows, that due to the presence of solid argon in the center of the pores the first few layers are already frozen at a higher temperature (30 K) compared to single layers (20 K). The transfer of our technique from the simple model system that we present in this Letter to other more complex adsorbates and different porous samples should help to enlighten the phase behavior under confinement in further studies.

Recent reports on a potential association between the K-variant of the gene for butyrylcholinesterase (BCHE-K) and Alzheimer's disease (AD) are discordant. An initial finding of association through a synergistic enhancement of risk of APOE epsilon4 with late-onset AD has not been confirmed by others. We have conducted a case-control study of histopathologically confirmed AD (n=135) and non-AD (n=70) cases (age of death > or =60 years), in which we have genotyped for APOE epsilon4, BCHE-K, and BCHE-A1914G, a silent polymorphism 299 bp downstream of the BCHE-K mutation. The allelic frequency of BCHE-K was 0.13 in the controls and 0.23 in the AD cases, giving a carrier odds ratio (OR(c)) of 2.1 (95% C.I. 1.1-4.1) for BCHE-K in confirmed AD. The allelic frequency for the BCHE-1914G variant was 0.19 and 0.33 in controls and AD cases, respectively (OR(c)=2.4; 95% C.I. 1.3-4.5). In an older sub-sample of 27/70 controls and 89/135 AD patients with ages of death > or =75 years, the OR(c) was increased to 4.5 (95% C.I. 1.4-15) for BCHE-K and 2.7 (95% C.I. 1.0-7.2) for BCHE-1914G carriers. The BCHE-K association with AD became even stronger in carriers of at least one APOE epsilon4 allele. Only three out of 19 controls compared with 39/81 AD cases carried BCHE-K in addition to APOE epsilon4, giving an odds ratio of confirmed AD of 5.0 (95% C.I. 1.3-19) for BCHE-K carriers within APOE epsilon4 carriers. Five out of 19 controls and 52/81 AD cases carried BCHE-1914G, giving the same odds ratio of confirmed AD of 5.0 (95% C.I. 1.6-16) for BCHE-1914G carriers within APOE epsilon4 carriers. In addition, our results suggest strong linkage disequilibrium between BCHE-K and BCHE-1914G but no major association of the sole BCHE-1914G chromosome with AD. We conclude that BCHE through its K-variant, rather than a nearby marker, is a susceptibility factor for AD and enhances the AD risk defined by APOE epsilon4 alone in an age-dependent manner.

We discovered the missense mutation, A226V, in the ornithine-delta-aminotransferase (OAT) genes of two unrelated patients with gyrate atrophy of the choroid and retina (GA). One patient, who was a compound for A226V and for the premature termination allele R398ter, showed a significant (P < .01) decrease in mean plasma ornithine levels, following pyridoxine supplementation with a constant protein intake: 826 +/- 128 microM (n = 5; no pyridoxine supplementation) versus 504 +/- 112 microM (n = 6; 500 mg pyridoxine/d) and 546 +/- 19 microM (n = 6; 1,000 mg pyridoxine/d). In extracts of fibroblasts from a second GA patient homozygous for A226V and from Chinese hamster ovary cells expressing an OAT-cDNA-containing A226V, we found that OAT activity increased from undetectable levels to approximately 10% of normal when the concentration of pyridoxal phosphate was increased from 50 to 600 microM. A226V is the fourth disease-causing pyridoxine-responsive human mutation to be reported.

Six new mutations have been identified in the CFTR gene. These mutations, representing three different categories--missense (R31L, W1098R), nonsense (E1104X), and frameshift (441delA, 681delC, 1461ins4)--are located in exons 2, 4, 5, 9, and 17b of the gene and presumed to cause cystic fibrosis (CF) in patients. All these mutations are probably rare in the population, as no additional examples were found for any of them in a cohort of 545 CF patients. Our study also revealed a benign sequence variation (3499 + 45T-->C) in intron 17b.

Mitochondrial 3-hydroxy-3-methylglutaryl CoA synthase (mHS) is the first enzyme of ketogenesis, whereas the cytoplasmic HS isozyme (cHS) mediates an early step in cholesterol synthesis. We here report the sequence of human and mouse liver mHS cDNAs, the sequence of a HS-like cDNA from Caenorhabditis elegans, the structure of a partial human mHS genomic clone, and the mapping of the human mHS gene to chromosome 1p12-p13. The nucleotide sequence of the human mHS cDNA encodes a mature mHS peptide of 471 residues, with a mean amino acid identity of 66.5% with cHS from mammals and chicken. Comparative analysis of all known mHS and cHS protein and DNA sequences shows a high degree of conservation near the N-terminus that decreases progressively toward the C-terminus and suggests that the two isozymes arose from a common ancestor gene 400-900 million years ago. Comparison of the gene structure of mHS and cHS is also consistent with a recent duplication event. We hypothesize that the physiologic result of the HS gene duplication was the appearance of HS within the mitochondria around the time of emergence of early vertebrates, which linked preexisting pathways of beta oxidation and leucine catabolism and created the HMG CoA pathway of ketogenesis, thus providing a lipid-derived energy source for the vertebrate brain.

Long range physical mapping within the p21 region of the X chromosome identified a CpG rich island approximately 180 kb centromeric to the chronic granulomatous disease (CGD) locus. The segments adjacent to the CpG island hybridized to discrete bands in DNAs of several species and when used to screen retinal cDNA libraries led to the identification of cDNAs that detected a mRNA of 2.1 kb in many tissues. Molecular characterization of corresponding genomic clones of this novel human gene confirmed the origin of the cDNA clones and indicated a genomic structure with five exons spanning a total of 9 kb. The complete cDNA sequence revealed that this gene contained a putative open reading frame of 116 amino acids with a 3' untranslated region of 1.74 kb. The amino acid sequence shows a high degree of similarity to the predicted product of the tctex-1 gene of the mouse t complex. As linkage studies and patients with deletions have implicated the Xp21 region as containing the retinitis pigmentosa defect (RP3), the gene was assessed as a candidate disease gene in RP3 families. A single base pair polymorphism was identified within the coding region but no disease associated changes were found by single strand conformational polymorphism and sequencing analysis of amplified exons of 20 RP patients. Analysis of a dinucleotide repeat polymorphism within this gene in families affected with RP3 suggested refinement of the RP3 region.

The 51-kDa flavoprotein subunit of mitochondrial NADH:ubiquinone oxidoreductase (Complex I) [NADH dehydrogenase (ubiquinone), flavoprotein 1 (51 kDa); EC 1.6.5.3] plays an important role in the formation of the NADH-binding site and is believed to be the principal site of entry for electrons donated by NADH into the respiratory chain. Human cDNA fragments of the 51-kDa protein were generated by polymerase chain reaction and used to localize the gene (NDUFV1) for this subunit to 11q13 by two separate techniques. This region of the human genome is strongly implicated in a number of different forms of cancer.

Central core disease (CCD) is a morphologically distinct, autosomal dominant myopathy with variable clinical features. A close association with malignant hyperthermia (MH) has been identified. Since MH and CCD genes have been linked to the skeletal muscle ryanodine receptor (RYR1) gene, cDNA sequence analysis was used to search for a causal RYR1 mutation in a CCD individual. The only amino acid substitution found was an Arg2434His mutation, resulting from the substitution of A for G7301. This mutation was linked to CCD with a lod score of 4.8 at a recombinant fraction of 0.0 in 16 informative meioses in a 130 member family, suggesting a causal relationship to CCD.

Endothelial nitric oxide (NO) synthase is a unique NO synthase isoform that is expressed constitutively by vascular endothelium both in vivo and in vitro and is believed essential to local vascular homeostasis. This calcium/calmodulin-dependent isoform is distinct from neuronal NO synthase. Genomic clones encoding the human endothelial NO synthase were isolated and the structural organization of the gene was determined. The gene contains 26 exons spanning approximately 21 kilobases of genomic DNA, encodes a messenger RNA of 4052 nucleotides, and is present as a single copy in the haploid human genome. Characterization of 5'-flanking genomic regions indicates that the endothelial NO synthase promoter is "TATA-less" and exhibits proximal promoter elements consistent with a constitutively expressed gene that is found in endothelial cells, namely Sp1 and GATA motifs. The 5'-flanking region contains putative AP-1, AP-2, NF-1, heavy metal, acute-phase response shear stress, and sterol-regulatory cis-elements. The human endothelial NO synthase gene was assigned to the 7q35-->7q36 region of chromosome 7 by Southern blot hybridization of human-rodent somatic cell hybrid lines and fluorescence in situ hybridization, whereas human neuronal NO synthase localized to the 12q24.2 region of chromosome 12.

A family of proinflammatory cytokines sharing several structural features has been described and includes, for example, interleukin-8, monocyte chemoattractant protein-1, and melanocyte growth stimulatory activity. Recently, the receptors for interleukin-8 have been isolated and found to belong to the seven-transmembrane domain class of G protein-coupled receptors. As other members of this cytokine family likely interact with similar receptors, the polymerase chain reaction was employed to isolate related receptors from human peripheral blood adherent cells. Degenerate oligonucleotide primers based on the rabbit interleukin-8 receptor sequence were used. The corresponding full-length cDNA was isolated from a human spleen cDNA library. The predicted protein sequence of this clone, designated pBE1.3, was 93% identical to that of a cDNA isolated from bovine locus coeruleus, which apparently encodes a neuropeptide Y receptor, and also shows similarity with the interleukin-8 receptor and the human cytomegalovirus US28 sequences. The gene, designated D2S201E was localized to human chromosome 2q21. By Northern blotting, transcripts hybridizing to this cDNA were present in a variety of tissues and cells, including those of hemopoietic origin.

Rom-1 and peripherin are related membrane proteins of the photoreceptor outer segments. Both proteins are located at the rims of the photoreceptor disks, where they may act jointly in disk biogenesis. Mutations in the gene (RDS) encoding peripherin cause autosomal dominant retinitis pigmentosa, autosomal dominant punctata albescens and butterfly macular degeneration in man, and retinal degeneration slow in mice. To facilitate ROM1 mutation and linkage analysis in inherited retinal diseases, we cloned and characterized the human and murine ROM1 genes. In both species, the ROM1 coding region is contained within approximately 1.8 kb of genomic DNA and is interrupted by only two introns. The structures of the ROM1 and RDS genes are similar, with perfect conservation of the intron splice sites. Putative transcription regulatory regions of the ROM1 locus, 5' to an apparent transcription start site, were identified by cloning the mouse Rom-1 gene and comparing the sequence to the human homologue. Alignment of the human and murine rom-1 predicted protein sequences with the peripherin polypeptides of four species reveals a high degree of conservation (47% overall identity between the six proteins) in the central hydrophilic domain of the two family members. Despite this conservation of sequence, the predicted pI's of only this region of rom-1 and peripherin differ substantially, being 5.2 and 8.2, respectively. The charge difference in this region may mediate the non-covalent association of these two proteins in vivo. The conserved genomic structure and sequence of ROM1 and RDS indicates that these genes evolved from a common ancestor by duplication event.

3-Hydroxy-3-methylglutaryl coenzyme A lyase (HL) is a homodimeric mitochondrial matrix enzyme that catalyzes the last step of ketogenesis. Using a human HL cDNA as a probe, we isolated a 1.4-kb mouse HL cDNA (HLM) from a mouse liver library and extended the sequence in the 5' direction, using RACE PCR to include the complete coding sequence. The nucleotide sequence of the mouse HL coding region is 85.7% identical to human HL, and 52.6% to Ps. mevalonii HL. Peptide identities of 87.4% and 54.3% respectively were observed. Southern analysis of 29 strains of laboratory mice and of Mus spretus revealed a total of about 25 kb of hybridizing fragments and three polymorphic fragments in both EcoRI and Hin-dIII digestions. The mouse HL locus (Hmgcl) was localized on Chromosome (Chr) 4: Pmv-19-12.6 +/- 3.6 cM-Hmgcl-7.3 +/- 2.3 cM-Xmv-8-1.5 +/- 1.0 cM-Gpd-1. The human HL locus (HMGCL) was mapped to distal Chr 1p by analysis of a human-hamster hybrid cell panel and by in situ hybridization.

Myosin participates in a varying repertoire of cellular functions ranging from cytokinesis, receptor capping and secretion to sarcomere contraction. In vertebrates this functional complexity is achieved through the regulated expression of gene families encoding isoproteins for each of the myosin subunits. We report here the identification and characterization of a gene (MYL5) that encodes a novel regulatory myosin light chain isoprotein and maps 700 kb from the human chromosome 4p telomere. Identical cDNAs have been isolated from human adult retina and fetal muscle cDNA libraries. A full length 519 bp open reading frame was identified in the cDNA sequence encoding a predicted protein of 173 residues. Sequence analysis of a 5.6 kb genomic region that encodes these cDNAs revealed the presence of 7 exons which span 4 kb. Expression of this gene has been detected in human adult retina, cerebellum, basal ganglia and fetal skeletal muscle. Whereas Northern analysis fails to detect transcription of this gene in human adult skeletal muscle it reveals an abundant transcript in monkey skeletal muscle. Phylogenetic comparison of the predicted proteins primary structure to those of related myosin light chains from Drosophila, rat and human reveal evolutionarily conserved structural motifs important for both calcium binding and phosphorylation.

The genetic defect underlying Huntington's disease (HD) has been mapped to 4p16.3. Refined localization using recombinant HD chromosome analysis and allelic association analyses have identified two distinct candidate regions. Using a cDNA hybrid selection procedure we have cloned the gene for alpha-adducin, a subunit of a cytoskeletal protein crucial for spectrin-actin membrane plasticity. This gene maps to the proximal 2.2 Mb candidate region within 20 kb of D4S95. Alleles of markers at this locus have been shown to exhibit significant linkage disequilibrium with HD. A 4 kb alpha-adducin transcript was identified which is abundantly expressed in the caudate nucleus, the site of major neuronal loss in HD. Sequencing of the brain alpha-adducin cDNA from two HD patients and an age-matched control did not detect any sequence alterations specific to HD. However, we identified in brain cDNA of both patients and control samples, two alternately spliced brain exons, not previously described in the erythrocyte cDNA. A 93 bp exon is inserted in frame between codon 471 and 472 while a 34 bp exon inserted within codon 621 disrupts the frame and introduces a stop codon after 11 novel amino acids. The mapping of the adducin gene adjacent to D4S95 and its pattern of expression, as well as its potential for distinct alternately spliced variants, reinforces the necessity to accurately assess the role of the expression of this gene in the pathogenesis of HD.

The constitutive calcium/calmodulin-dependent nitric oxide (NO) synthase expressed in vascular endothelium shares common biochemical and pharmacologic properties with neuronal NO synthase. However, recent cloning and molecular characterization of NO synthase from bovine endothelial cells indicated the existence of a family of constitutive NO synthases. Accordingly, we undertook molecular cloning and sequence analysis of human endothelial NO synthase. Complementary DNA clones predict a protein of 1,203 amino acids sharing 94% identity with the bovine endothelial protein, but only 60% identity with the rat brain NO synthase isoform. Northern blot analysis with an endothelial-derived cDNA identified a 4.6-4.8 kb mRNA transcript in HUVEC and in situ hybridization localized transcripts to vascular endothelium but not neuronal tissue.

The biosynthesis of protein-bound complex N-glycans in mammals requires a series of covalent modifications governed by a large number of specific glycosyltransferases and glycosidases. The addition of oligosaccharide to an asparagine residue on a nascent polypeptide chain begins in the endoplasmic reticulum. Oligosaccharide processing continues in the Golgi apparatus to produce a diversity of glycan structures. UDP-N-acetylglucosamine:alpha-3-D-mannoside beta-1,2-N-acetylglucosaminyltransferase I (EC 2.4.1.101; GlcNAc-TI) is a key enzyme in the process because it is essential for the conversion of high-mannose N-glycans to complex and hybrid N-glycans. We have isolated the mouse gene encoding GlcNAc-TI (Mgat-1) from a genomic DNA library. The mouse sequence is highly conserved with respect to the human and rabbit homologs and exists as a single protein-encoding exon. Mgat-1 was mapped to mouse Chromosome 11, closely linked to the gene encoding interleukin-3 by the analysis of multilocus interspecies backcrosses. RNA analyses of Mgat-1 expression levels revealed significant variation among normal tissues and cells.

TFIIIA is an RNA polymerase III transcription factor that binds to the internal control region of the 5 S RNA gene as the first step in the assembly of a transcription complex. We have identified the gene encoding TFIIIA from Saccharomyces cerevisiae. Protein synthesized in vitro from the cloned gene has the same size, DNA-binding properties, and transcription factor activity as does purified yeast TFIIIA. Examination of the deduced sequence of the 50-kDa yeast transcription factor revealed the presence of nine zinc-finger motifs, a characteristic of Xenopus TFIIIA. Although the conservation of these nine putative DNA-binding domains is striking, the amino acid sequence throughout the corresponding fingers of the yeast and amphibian TFIIIAs has diverged extensively and in many instances the spacing between the residues that coordinate the zinc ions differs between the two proteins. A unique feature of the yeast protein is an 81-amino acid domain interrupting the repeated zinc-finger motifs between fingers 8 and 9. Additionally, the yeast and amphibian proteins differ in both the size and sequence of the amino- and carboxyl-terminal domains flanking the zinc fingers. The gene encoding yeast TFIIIA is present in single copy in the S. cerevisiae genome and is essential for cell viability. A carboxyl-terminal truncated form of the protein containing 4.5 zinc-finger motifs retains the ability to bind to DNA but is no longer active in promoting transcription in vitro.

As part of the search for the Huntington disease (HD) gene we have cloned and sequenced 34 kb of genomic DNA containing the full-length gene for the beta-subunit of the human cGMP phosphodiesterase (beta-cGMP PDE). This gene is localized to 4p16.3 about 700 kb proximal to the 4p telomere and represents the most telomeric gene characterized on 4p to date. We show that this gene is comprised of 22 exons spanning approximately 43 kb of genomic DNA. We also provide 400 bp immediately 5' to the putative initiator methionine and 700 bp of 3' flanking sequences. Northern blot analysis of several human tissues revealed a highly abundant 3.5 kb transcript and a minor signal of 4.5 kb in retinal tissue. Alignment of the deduced amino acid sequence to the previously identified beta-subunits of the cGMP PDEs of mouse and cow demonstrates highly significant similarities and, therefore, confirms the identity of the cloned gene. A defect in the beta-subunit of the cGMP PDE gene has been shown recently to be the cause for the retinal degeneration in the rd mouse. The cloning of the human homolog and the knowledge of its genomic organization with exon/intron boundaries will allow rapid assessment of the role of this gene in the causation of human retinopathies.

PRP11 is a gene that encodes an essential function for pre-messenger RNA (mRNA) processing in Saccharomyces cerevisiae. We have carried out a mutational study to locate essential and non-essential regions of the PRP11 protein. The existing temperature-sensitive (ts) mutation (prp11-1) was isolated from the chromosome of the original mutant and its position in the gene was determined. When the prp11-1 gene was transcribed from the GAL1 promoter, the overproduced protein was able to reverse the ts prp11-1 phenotype; this is compatible with the possibility that the defect in the prp11-1 gene product affects its binding to the spliceosome. Thirteen linker-insertion mutations were constructed. Only five (prp11-4, 11-6, 11-10, -13 and -14) resulted in a null phenotype. One of these became temperature-sensitive when the insertion was reduced in size from four (prp11-10) to two (prp11-15) amino acids. A sequence of ten amino acids of which also occurs in the human U1 small nuclear ribonucleoprotein particle (snRNP) A protein and the U2 snRNP B" protein, when deleted from PRP11, had no phenotype and thus appears to be nonessential for PRP11 function. However, a linker-insertion mutation (prp11-10) immediately adjacent to this region resulted in a null phenotype.

The influence of the trichothecene mycotoxin T-2 on the mitochondria of Saccharomyces cerevisiae was studied. T-2 is a cytotoxic molecule inhibiting growth and macromolecular synthesis in S. cerevisiae. At low concentrations, T-2 toxin arrested yeast growth on glycerol medium and at higher concentrations, it arrested growth on glucose medium. The toxin was not capable itself of inducing petite mutations. Its inhibitory effect on the growth of petite strains, of both chromosomally isogenic and non-isogenic strains was less than that of grande strains. One exception to this was equally low susceptibility of psi+ SUP4-3 strain in both rho+ and rho- state. T-2 toxin was also capable of retarding the petite inducing activity of the mutagen, ethidium bromide. T-2 toxin inhibited the polymerization of P-ribo-sylaminoimidazole in an ade2 strain of S. cerevisiae. These results show that T-2 toxin is capable of interfering with the activity of the mitochondria in addition to its well studied effects on cytoplasmic protein synthesis.

A Fusarium metabolite, T-2 toxin, inhibits the growth of Saccharomyces carlsbergensis and Saccharomyces cerevisiae. The growth inhibitory concentrations of T-2 toxin were 40 and 100 mug/ml, respectively, for exponentially growing cultures of the two yeasts. S. carlsbergensis was more sensitive to the toxin and exhibited a biphasic dose-response curve. Addition of the toxin at 10 mug/ml of S. carlsbergensis culture resulted in a retardation of growth as measured turbidimetrically, after only 30 to 40 min. This action was reversible upon washing the cells free of the toxin. The sensitivity of the yeasts to the toxin was dependent upon the types and concentrations of carbohydrates used in the growth media. The sensitivity of the cells to the toxin decreased in glucose-repressed cultures. These results suggest that T-2 toxin interferes with mitochondrial functions of these yeasts.

An outbreak of irritant contact dermatitis associated with residual percholorethylene (PCE) in dry-cleaned garments was studied at a large semiconductor manufacturing facility. A new method was developed to measure PCE levels which detected concentrations of 0.83 to 32.01 ppm in the garment.