Genetic analysis of the yeast NUD1 endo-exonuclease: a role in the repair of DNA double-strand breaks

The deoxyribonucleases (DNases) have been shown genetically to be important in the vital processes of DNA repair and recombination. The NUD1 gene, which codes for an endo-exonuclease of Saccharomyces cerevisiae, was analyzed for its role in the DNA double-strand break (DSB) repair processes. While the nud1 strain is only slightly sensitive to ionizing radiation, expression of the HO-endonuclease to introduce a DSB at the MAT locus in that strain results in cell death. Cell survival is inversely proportional to the duration of HO-endonuclease expression. Analysis of the surviving colonies from the nud1 strain indicated that many of the survivors are sterile and that the proportion of these sterile survivors increases with the time of HO-endonuclease expression. On the other hand, the surviving colonies from the isogenic NUD1 strain are mating-proficient. Interestingly, double mutants of nud1 rad52 are more resistant to ionizing irradiation than the rad52 strain and have a cell-survival fraction of 32% for rad52-1 nud1 and 9% for rad52::URA3 nud1 following prolonged HO-endonuclease expression, indicating that nud1 has a suppressor effect on the DSB-induced lethality in rad52. Polymerase chain reaction analysis showed that many of the nud1 survivors contained small alterations within the MAT locus, suggesting that the survivors arose through the process of non-homologous end-joining. These results suggest that the endo-exonuclease acts at a DSB to promote DNA repair via the homologous recombination pathway. Received: 20 July / 20 September 1998     

Newer aspects of antiarrhythmic drug development: Introduction

Although cardiac arrhythmias remain a serious clinical problem in many patients with heart disease, the exact role of antiarrhythmic drug therapy is currently under intense evaluation. Within the last several years it has become clear that there are significant risks as well as potential benefits associated with existing agents. Ongoing studies in large patient populations should help determine the benefit/risk ratio of traditional therapy. Regardless of the outcome of these trials, current electrophysiological dogma will have to be re-evaluated and newer concepts evolve for drug development to make further progress. The goal of this symposium is to exchange information among basic and clinical investigators so as to facilitate the emergence of novel electrophysiological concepts that will form the basis for future generations of antiarrhythmic drugs.     

Expression of β-catenin in normal breast tissue and breast carcinoma: a comparative study with epithelial cadherin and α-catenin

Expression of β-catenin was investigated in normal breast tissue and 66 breast carcinomas in conjunction with expression of epithelial cadherin (E-CD) and α-catenin. In normal mammary ducts and acini, intense β-catenin immunoreactivity was present at the basolateral surfaces of luminal epithelium and weak immunoreactivity was observed at the lateral borders of myoepithelial cells. No β-catenin was revealed at the myoepithelial basal surface. The intercellular expression of β-catenin, as well as of E-CD and α-catenin, was also observed in carcinoma tissues with varying staining intensity. Almost all of 10 intraductal carcinomas and approximately 70% of 41 invasive ductal carcinomas expressed the three molecules at the same level as in normal glands, whereas approximately 80% of 13 invasive lobular carcinomas showed severe deficiency of them. Two lobular carcinomas in situ showed complete absence of all of the proteins. Some of these findings were confirmed biochemically by immunoblotting analysis. In invasive ductal carcinomas, α-catenin was reduced more frequently in diffuse than in solid type tumours, whereas the level of expression of β-catenin and E-CD was unchanged between them. No correlation was present between reduced expression of the adhesion molecules and lymph node metastasis.     

The role of α2-adrenoceptors of the medullary lateral reticular nucleus in spinal antinociception in rats

We attempted to find out the role of α₂-adrenoceptors of the medullary lateral reticular nucleus (LRN) in antinociception in rats. Spinal antinociception was evaluated using the tail-flick test, and supraspinal antinociception using the hotplate test. Antinociceptive effects were determined following local electric stimulation of the LRN, and following microinjections of medetomidine (an α₂-adrenoceptor agonist; 1–10 μg), atipamezole (an α₂-adrenoceptor antagonist; 20 μg) or lidocaine (4%) into the LRN. The experiments were performed using intact and spinalized Hannover-Wistar rats with a unilateral chronic guide cannula. Electric stimulation of the LRN as well as of the periaqueductal gray produced a significant spinal antinociceptive effect in intact rats. Medetomidine (1–10 μg), when microinjected into the LRN, produced no significant antinociceptive effect in the tail-flick test in intact rats. However, following spinalization, medetomidine in the LRN (10 μg) produced a significant atipamezole-reversible antinociceptive effect in the tail-flick test in the hot-plate test, medetomidine (10 μg) in the LRN produced a significant atipamezole-reversible increase of the paw-lick latency in intact rats. Microinjection of atipamezole (20 μg) or lidocaine alone into the LRN produced no significant effects in the tail-flick test. The results are in line with the previous evidence indicating brat the LRN and the adjacent ventrolateral medulla is involved in descending inhibition of spinal nocifensive responses. However, α₂-adrenoceptors in the LRN do not mediate spinal antinociception but, on the contrary, their activation counteracts antinociception at the spinal cord level. The spinal aninociceptive effect of supraspinally administered medetomidine in spinalized rats can be explained by a spread of the drug (e.g., via circulation) which then directly activates α₂-adrenoceptors at the spinal cord level.     

Microtubule-associated protein tau is required for axonal neurite elaboration by neuroblastoma cells.

NB2a/d1 neuroblastoma cells constitutively express multiple isoforms of the microtubule-associated protein tau and incorporate this protein into the axonal neurites elaborated during serum deprivation. To examine whether or not tau played an essential role in axonal outgrowth, cells cultured in serum-free medium were treated at 24 h intervals with antisense- and sense-oriented cDNA oligonucleotides (25 or 36 mers that span or are upstream of tau initiation codon) and were simultaneously serum deprived. Oligonucleotide uptake was confirmed by determination of intracellular levels of radiolabeled oligonucleotides. Treatment for 48 h with tau antisense oligonucleotides reversibly inhibited the expression of tau and the number of neurite-bearing cells compared with treatment with sense oligonucleotides. By contrast, tubulin expression was not affected. When cells were treated with antisense oligonucleotide simultaneously with serum deprivation, the initial outgrowth of neurites was unaffected, but continued neurite elongation was prevented. By contrast, neurite outgrowth at 4 h was inhibited when cells were pretreated with tau antisense 24 h before serum deprivation. Furthermore, intracellular delivery of anti-tau antiserum prevented neurite outgrowth and, in cells that had previously been deprived of serum for 24 h, induced retraction of existing neurites. These findings indicate that both the initiation and the continued outgrowth of neurites are dependent on tau and that pre-existing cytoplasmic pools of tau can mediate initial neuritogenesis.     

Distribution of extracellular matrix components and their receptors in human lymphoid tissue and B-cell non-Hodgkin lymphomas

In this study the distribution patterns of various extracellular matrix components and their receptors (i.e. β1 integrins) in B-cell non-Hodgkin lymphomas were examined and compared to those in reactive lymphoid tissue. Neoplastic follicles within follicular lymphomas showed similar patterns to that observed in reactive follicles, which appeared to be strongly associated with the presence of follicular dendritic cells. Diffuse lymphomas of low and intermediate malignancy grade revealed features comparable to those of interfollicular areas of reactive lymphoid tissue, irrespective to which compartment the tumour cells were related. Highly malignant lymphomas, however, displayed unique extracellular matrix configurations, resulting from active matrix degradation by macrophages; this may support rapid tumour growth. Extranodal lymphomas showed virtually the same matrix patterns as their nodal counterparts, suggesting that (malignant) lymphoid cells generate (at least partly) their own specific microenvironment. In reactive lymphoid tissue β1 integrins were mainly found on resident cells and except for α4, α5 (and β1) the lymphoid cells expressed very little, if any, β1 integrins. In comparison, expression of these integrins on lymphoma cells was reduced (follicular lymphomas) or could not be detected at all (diffusely growing lymphomas); this might contribute to the growth pattern and metastatic properties of the tumours.     

Reversible low-molecular-weight proteinuria in patients with distal renal tubular acidosis

Four patients with untreated renal tubular acidosis had a urinary excretion of low-molecular-weight (LMW) proteins which was restored to normal by alkali therapy. Hypokalaemic proximal tubular damage in untreated patients with distal renal tubular acidosis is believed to be the cause of LMW proteinuria. An examination of urinary excretion of LMW proteins is useful for determining hypokalaemic proximal tubular dysfunction, as well as the efficiency of alkali therapy.     

Insulin-like growth factor I promotes leiomyoma cell growth in vitro

Objective: Our purpose was to determine whether insulin-like growth factors I and II preferentially stimulate uterine leiomyoma cells versus myometrial cells in monolayer culture.Study design: Leiomyomas and normal myometrium were obtained at hysterectomy from five premenopausal women. Specimens were enzymatically digested for use in primary monolayer cell cultures. By use of serum-free media, insulin-like growth factor I or II was added in 1, 10, and 100 ng/ml concentrations to both cell types with the patient serving as her own control. Cell number, prolactin production, and proliferative index values were measured on day 15 of cell culture.Results: Significant increases in cell number were found in the leiomyoma cultures (p < 0.05) treated with 10 and 100 ng/ml insulin-like growth factors I but not with insulin-like growth factos II. Neither factor exerted a stimulatory effect on myometrial cells.Conclusion: Insulin-like growth factor sI preferentially stimulates leiomyoma cells in monolayer culture. These results suggest an autocrine-paracine role in vivo for this factor in conjuction with gonadal steroids in promoting leiomyoma growth.     

Case-control study of the alpha-synuclein interacting protein gene and Parkinson's disease.

We conducted a case-control study of the alpha-synuclein-interacting protein gene (SNCAIP, also known as synphilin-1) and Parkinson's disease (PD). A total of 319 PD cases and 195 controls were genotyped for four SNCAIP variants, including a microsatellite repeat in intron 4 and three restriction fragment length polymorphisms (RFLP) proximal to the 5' terminal of exons 1, 4, and 6. None of the variants were found associated with PD overall. Global score statistics were not significant for four, three, and two loci haplotypes. All four loci were in linkage disequilibrium for cases, controls, or both groups combined (P < 0.0001). Recursive partitioning showed no interactions between variants of the SNCAIP gene and variants of the alpha-synuclein gene (SNCA) or the parkin (PARK2) gene.