Sterile mutants of Schizosaccharomyces pombe: Analysis by somatic hybridization

Summary Sixteen sterile mutants of Schizosaccharomyces pombe, isolated from homothallic h ⁹⁰strains, were examined. It was found that they are blocked either in copulation and meiosis or in copulation alone.Protoplasts of the sterile strains were fused with protoplasts of h^+N, h^–S or h ⁹⁰strains. In these somatic crosses, eight of the sterile strains yielded hybrids which were able to form azygotic asci. Tetrad analyses revealed that seven of these sterile strains contain a single mutation; the mutations represent at least five sterility genes (ste2, ste3, ste4, ste5, ste6). One sterile strain contains two unlinked mutations which do not represent sterility genes, but genes the interaction of which results in a sterile phenotype.     

Dominant-negative effect of hetero-oligomerization on the function of the human immunodeficiency virus type 1 envelope glycoprotein complex

The human immunodeficiency virus type 1 (HIV-1) envelope (Env) glycoprotein forms trimers that mediate interactions with the CD4 receptor and a co-receptor on the target cell surface, thereby triggering viral fusion with the cell membrane. Cleavage of Env into its surface, gp120, and transmembrane, gp41, moieties is necessary for activation of its fusogenicity. Here, we produced pseudoviruses with phenotypically mixed wild-type (Wt) and mutant, cleavage-incompetent Env in order to quantify the effects of incorporating uncleaved Env on virion infectivity, antigenicity and neutralization sensitivity. We modeled the relative infectivity of three such phenotypically mixed viral strains, JR-FL, HXBc2 and a derivative of the latter, 3.2P, as a function of the relative amount of Wt Env. The data were fit very closely (R(2) > 0.99) by models which assumed that only Wt homotrimers were functional, with different approximate thresholds of critical numbers of functional trimers per virion for the three strains. We also produced 3.2P pseudoviruses containing both a cleavage-competent Env that is defective for binding the neutralizing monoclonal antibody (NAb) 2G12, and a cleavage-incompetent Env that binds 2G12. The 2G12 NAb was not able to reduce the infectivity of these pseudoviruses detectably. Their neutralization by the CD4-binding site-directed agents CD4-IgG2 and NAb b12 was also unaffected by 2G12 binding to uncleaved Env. These results further strengthen the conclusion that only homotrimers consisting of cleaved Env are functional. They also imply that the function of a trimer is unaffected sterically by the binding of an antibody to an adjacent trimer.     

Characteristics of hepatitis B viral strains in chronic carrier patients from North-East Algeria

Objective

The goal of the study was to determine the characteristics of chronic hepatitis B virus (HBV) infections in the north-east part of Algeria. Chronic HBV infection remains a global public health issue and Algeria is considered as an intermediate prevalence area. Improving our knowledge on the epidemiology of this infection is a prerequisite to adopt the best preventive and curative strategy.

Material and methods

We have studied 75 chronic hepatitis B patients from north-east Algeria. The characteristics of HBV strains were determined by use of serological and molecular testing. Genes encoding part of the precore, the surface and the polymerase were sequenced and phylogenetically analyzed.

Results

Median age of the patients was 35 years and 80% of them had normal transaminase level. Liver histological lesions were identified in 63% of the patients who benefited from a liver biopsy and 21% of them had cirrhosis. Median viral replication was 3.9 Log IU/ml and 87% of patients had a “precore” mutant serological profile without HBe Ag. Genotype D was predominant (93%) followed by genotype A (5%) and E for one patient. Algerian strains clustered independently from other genotype D reference sequences, suggesting a possible new D subtype. Within the “precore” region, only 16% of the strains did not show any mutation at positions 1762/1764 and 1896.

Conclusion

In this original set of patients from north Algeria, the virologic characteristics of HBV are comparable to what has been described in other Mediterranean countries. Our study raises several important aspects with regard to the prevention and treatment of chronic hepatitis B in Algeria.     

The impact of envelope glycoprotein cleavage on the antigenicity, infectivity, and neutralization sensitivity of Env-pseudotyped human immunodeficiency virus type 1 particles

Endoproteolytic processing of the human immunodeficiency virus type 1 (HIV-1) envelope (Env) glycoproteins is an obligate part of the biosynthetic pathway that generates functional, fusion-competent Env complexes, which are then incorporated into infectious virions. We have examined the influence of cleavage on Env-specific antibody reactivity, Env incorporation into pseudovirions, and the infectivity and neutralization sensitivity of Env-pseudotyped viruses. To do so, we have used both incompletely processed wild-type (Wt) Env and engineered, cleavage-defective Env mutants. We find that there is no simple association between antibody reactivity to cell surface-expressed Env, and the ability of the same antibody to neutralize virus pseudotyped with the same Env proteins. One explanation for the absence of such an association is the diverse array of Env species present on the surface of transiently transfected cells. We also confirm that cleavage-defective mutants are antigenically different from Wt Env. These findings have implications for the use of Env binding assays as predictors of neutralizing activity, and for the development of cleavage-defective Env trimers for use as subunit immunogens.     

Neutralization sensitivity of HIV-1 Env-pseudotyped virus clones is determined by co-operativity between mutations which modulate the CD4-binding site and those that affect gp120-gp41 stability

Adaptation of antibody neutralization-resistant human immunodeficiency virus type I (HIV-1) to growth in vitro generally results in the acquisition of a neutralization-sensitive phenotype, an alteration of viral growth kinetics, and an array of amino acid substitutions associated with these changes. Here we examine a panel of Env chimeras and mutants derived from these neutralization-resistant and -sensitive parental Envs. A range of neutralization and infectivity phenotypes was observed. These included a modulation of the CD4 binding site (CD4bs) towards recognition by neutralizing and non-neutralizing CD4bs-directed antibodies, resulting in a globally neutralization-sensitive Env; alterations which affected Env complex stability; and interactions which resulted in differential infectivity and CCR5/CXCR4 usage. This range of properties resulted from the complex interactions of no more than three amino acids found in key Env locations. These data add to a growing body of evidence that dramatic functional alterations of the native oligomeric Env protein complex can result from relatively minor amino acid substitutions.     

Isolation and properties of yeast mutants affected in farnesyl diphosphate synthetase

Summary Two yeast mutant strains auxotrophic for ergosterol and blocked in farnesyl diphosphate synthetase (EC 2.5.1.1) were isolated. Genetic analysis has shown that these mutant strains carry additional mutations in the ergosterol pathway besides erg20-1 and erg20-2 which affect FPP synthetase. The novel feature of these mutants is their ability to excrete prenyl alcohols (farnesol and geraniol). As geraniol is toxic for yeast cells, the above leaky mutations in FPP synthetase have to be associated with others in the sterol pathway, in order to slow down geraniol synthesis.Abbreviations FPP farnesyl diphosphate - GPP geranyl diphosphate - IPP isopentenyl diphosphate - DMAPP dimethylallyl diphosphate - MVAP mevalonate phosphate - MVAPP mevalonate diphosphate - HMG CoA 3-hydroxy-3-methylglutaryl-CoA - MIC minimal inhibitory concentration     

庆大霉素产生菌23-18诱变育种研究

用青霉素、超声波对庆大霉素产生菌棘孢小单抱菌23-18的孢子进行预处理，以增加其通透性，再用钻60进行辐射诱变，结合耐自身代谢产物的筛选，获得数株高产菌株，其中79，167菌株摇瓶发酵数价比亲株23－18提高20％，发酵周期缩短10h。萌种连续传五代，生产能力稳定，其发酵液用高效液相色谱分析，C组分比例符合中国药典要求。     

Relationships between trehalose metabolism and maltose utilization in Saccharomyces cerevisiae

Summary A specific deficiency in UDPG-linked trehalose-6-phosphate synthase in the yeast, Saccharomyces cerevisiae has been associated with a single nuclear gene, sst1. Strains bearing this abnormal allele lacked the capacity to accumulate trehalose during growth on glucose or galactose medium or when incubated with glucose in nonproliferating conditions. However, sst1 strains still exhibited trehalose accumulation during growth on maltose medium, provided they contained a gene for maltose fermentation (MAL gene). Introduction of a constitutive MAL ^c gene into an sst1 strain rendered the strain capable of accumulating trehalose during growth on glucose medium, but did not restore the normal capacity to convert glucose to trehalose in nonproliferating conditions. Different systems, I and II, of trehalose accumulation are proposed. System I would require the UPDG-linked synthase, whereas system II, which is normally specific for maltose, would utilize a different enzyme. It is unlikely that system II produces trehalose by trans-glucosylation, since it converted glucose to trehalose in MAL ^c sst1 strains. The results indicate that maltose specifically induces the production of the MAL gene-product, which, in turn, would stimulate the formation (or activation) of system II.     

Characterization of susceptibility variants of poliovirus grown in the presence of favipiravir

Background

T-705 (favipiravir) is a potent inhibitor of RNA-dependent RNA polymerases of influenza viruses and no favipiravir-resistant virus has been isolated. Poliovirus RNA polymerase has been well characterized and isolation of resistant virus was examined in poliovirus.