DNA ligase IV mutations identified in patients exhibiting developmental delay and immunodeficiency.

DNA ligase IV functions in DNA nonhomologous end-joining and V(D)J recombination. Four patients with features including immunodeficiency and developmental and growth delay were found to have mutations in the gene encoding DNA ligase IV (LIG4). Their clinical phenotype closely resembles the DNA damage response disorder, Nijmegen breakage syndrome (NBS). Some of the mutations identified in the patients directly disrupt the ligase domain while others impair the interaction between DNA ligase IV and Xrcc-4. Cell lines from the patients show pronounced radiosensitivity. Unlike NBS cell lines, they show normal cell cycle checkpoint responses but impaired DNA double-strand break rejoining. An unexpected V(D)J recombination phenotype is observed involving a small decrease in rejoining frequency coupled with elevated imprecision at signal junctions.     

G(alpha)(i) controls the gating of the G protein-activated K(+) channel, GIRK.

GIRK (Kir3) channels are activated by neurotransmitters coupled to G proteins, via a direct binding of G(beta)(gamma). The role of G(alpha) subunits in GIRK gating is elusive. Here we demonstrate that G(alpha)(i) is not only a donor of G(beta)(gamma) but also regulates GIRK gating. When overexpressed in Xenopus oocytes, GIRK channels show excessive basal activity and poor activation by agonist or G(beta)(gamma). Coexpression of G(alpha)(i3) or G(alpha)(i1) restores the correct gating parameters. G(alpha)(i) acts neither as a pure G(beta)(gamma) scavenger nor as an allosteric cofactor for G(beta)(gamma). It inhibits only the basal activity without interfering with G(beta)(gamma)-induced response. Thus, GIRK is regulated, in distinct ways, by both arms of the G protein. G(alpha)(i) probably acts in its GDP bound form, alone or as a part of G(alpha)(beta)(gamma) heterotrimer.     

Kinetic study of an enzyme-catalysed reaction in the presence of novel irreversible-type inhibitors that react with the product of enzymatic catalysis

In the present paper a kinetic study is made of the behaviour of a Michaelis-Menten enzyme-catalysed reaction in the presence of irreversible inhibitors rendered unstable in the medium by their reaction with the product of enzymatic catalysis. A general mechanism involving competitive, non-competitive, uncompetitive and mixed irreversible inhibition with one or two steps has been analysed. The differential equation that describes the kinetics of the reaction is non-linear and computer simulations of its dynamic behaviour are presented. The results obtained show that the systems studied here present kinetic co-operativity for a target enzyme that follows the simple Michaelis-Menten mechanism in its action on the substrate, except in the case of an uncompetitive-type inhibitor.     

Isolation of Host-Dependent and Nonparasitic Mutants of the Facultative Parasitic Bdellovibrio UKi2

Obligate host-dependent and nonparasitic mutants were isolated from a facultative parasitic Bdellovibrio strain. Thus it is possible to separate host-dependency from the ability to parasitize in bdellovibrios.     

ANABOLIC RESPONSES OF EMBRYONIC DORSAL ROOT GANGLIA TO NERVE GROWTH FACTOR, INSULIN, CONCANAVALIN A OR SERUM IN VITRO

—Intact and dissociated dorsal root ganglia from 8-day chick embryos were examined for their ability to incorporate radio-precursors into RNA and protein in unsupplemented medium or in medium supplemented with Nerve Growth Factor, insulin, Concanavalin A, fetal calf serum, or several combinations of such agents. In the absence of any agent, incorporation into RNA and protein declined with time. All four agents maintained or improved the initial incorporation rates, and optimal doses were determined in each case. Different combinations of two agents led to potentiated, full or partially additive, or inhibited effects; in particular, NGF promoted incorporation even in conjunction with insulin (additive) or serum (potentiating). Several differences were noted between the responses of intact and of dissociated ganglia.     

Elongation and cell division in Bdellovibrio bacteriovorus

Elongation and division of Bdellovibrio bacteriovorus were studied in axenic synchronous cultures. The cells elongate unidirectionally from one end attaining a length of several unit cells, and then divide into the corresponding number of cells. The length of the filament and, consequently, the progeny number, vary within the range of two to several dozen cells, according to the conditions used. A protein and a low molecular weight component are required for normal division.     

Evaluation and selection of tandem repeat loci for <Emphasis Type="Italic">Streptococcus pneumoniae</Emphasis> MLVA strain typing

Background  

Precise identification of bacterial pathogens at the strain level is essential for epidemiological purposes. InStreptococcus pneumoniae, the existence of 90 different serotypes makes the typing particularly difficult and requires the use of highly informative tools. Available methods are relatively expensive and cannot be used for large-scale or routine typing of any new isolate. We explore here the potential of MLVA (Multiple Loci VNTR Analysis; VNTR, Variable Number of Tandem Repeats), a method of growing importance in the field of molecular epidemiology, for genotyping ofStreptococcus pneumoniae.     

Possible association of the allele status of the CS.7/HhaI polymorphism 5′ of the CFTR gene with postnatal female survival

Cystic fibrosis (CF) patients show a high degree of linkage disequilibrium between the CF transmembrane conductance regulator (CFTR) gene and polymorphisms 5′ of that gene. To determine whether the region 5′ of CFTR contains biologically important sequences, the allele frequencies of six CFTR-linked polymorphisms (metH/MspI, XV-2c/TaqI, CS.7/HhaI, KM19/PstI, MP6-d9/MspI, J44/XbaI) were determined in 417 randomly selected elderly individuals (over 75 years of age) from the Czech population. The elderly individuals were considered “escapees” of strong selective pressures that had operated during their lifetime, prior to the introduction of modern health care since 1950. The pooled allele frequencies of the analyzed marker polymorphisms in the elderly did not significantly differ from published data. However, when analyzed by sex, the allele frequencies of markers CS.7/HhaI and KM19/PstI differed significantly (P < 0.05) between elderly females and males. The allele frequencies of the six polymorphisms were then determined in 646 newborns and 345 young adults of reproductive age; these individuals were selected in a similar manner and drawn from the same population. In these control groups, the studied marker polymorphisms exhibited no statistically significant differences between sexes and/or between individuals of the same sex, only between different age groups. A gradual relative increase in the frequency of allele “2” of marker CS.7/HhaI was observed from newborn females to elderly women, the overall difference in allele frequencies of this marker polymorphism between newborn females and elderly women reaching statistical significance (P < 0.05). Interestingly, allele “2” is the major constituent of the extended “B-haplotype”, which is in strong linkage disequilibrium with common CF alleles. Taken together, our data suggest that the region spanning markers CS.7 and KM19 is associated with a genetic factor that influences postnatal female survival, providing a possible mechanism for increasing the frequency of particular mutations in the adjacent CFTR gene. Received: 30 January 1996 / Revised: 16 December 1996