Topoisomerase I has a strong binding preference for a conserved hexadecameric sequence in the promoter region of the rRNA gene from Tetrahymena pyriformis.

Topoisomerase I is in situ associated with DNaseI hypersensitive sites located in the promotor and terminator regions of the extrachromosomal rDNA in Tetrahymena thermophila at sites with sequences fitting the motif (sequence in text) Reconstitution experiments with purified topoisomerase I and cloned fragments of rDNA demonstrate that the enzyme exhibits the same binding and cleavage properties on naked DNA. These observations are striking as topoisomerase I previously has been found to exhibit low sequence specificity. The specific binding of the enzyme has an absolute requirement for divalent cations with a preference for Ca2+. The strong binding to the hexadecamer has been characterized by competition experiments, and it has been used to determine the molecular weight of the enzyme.     

Further segregation analysis of the fragile X syndrome with special reference to transmitting males

Summary A new series of 96 pedigrees with the fra(X) syndrome was analysed using complex segregation analysis with pointers, defining affection as any degree of mental impairment. These families were found to exhibit the same segregation pattern as the first series of 110 pedigrees (Sherman et al. 1984). The best estimate for penetrance of mental impairment in males was 79% and in females was 35% for the combined data. Again, there was little evidence for sporadic cases among affected males.Many more intellectually normal transmitting males have been observed since the existence of such males and the concomitant need to investigate the paternal side of pedigrees was recognized. On further investigation of all 206 pedigrees from the old and new data sets, the sibships of nonexpressing males appeared to be different from those of expressing males. Our analysis, using mental impairment as the phenotype, suggested that obligate carrier mothers and daughters of intellectually normal transmitting males are rarely, if ever, mentally impaired and that the sibs of transmitting males are much less likely to be retarded than the sibs of mentally impaired males. Though mothers and daughters of transmitting males are similar in phenotype, the expression of the gene in their offspring appears to be different: the penetrance of mental impairment is higher in offspring of intellectually normal daughters of transmitting males than in offspring of intellectually normal mothers of transmitting males. The implications of these observations for genetic counseling and for genetic models of the fra(X) syndrome are discussed.     

immunocytochemical localization of urokinase-type plasminogen activator in lewis lung carcinoma

The invasively growing and metasizing Lewis lung carcinoma consistently contained urokinase-type plasminogen activator (u-PA) enzyme activity. When investigated immunocytochemically with antibodies against u-PA, different parts of individual tumors showed a pronounced heterogeneity in staining intensity. Strong staining was found in areas with invasive growth and degradation of surrounding normal tissue, while other areas were completely devoid of staining. Immunoreactivity occurred both with a perinuclear cytoplasmic localization in tumor cells and associated with apparently extracellular material. SDS PAGE of tumor extracts, under both reducing and nonreducing conditions, followed by immunoblotting, showed only one immunocytochemically stainable band with an electrophoretic mobility corresponding to that of purified proenzyme to u-PA, while no two-chain u-PA was detected. This indicates that the major part of the activator in Lewis lung carcinoma is present as one-chain pro-u-PA.     

Effect of vasoactive intestinal polypeptide (VIP) on steroidogenesis in women

The VIPergic nervous system appears to be the major peptide-containing neuronal component in the female genital tract. Evidence has been put forward that exogenous VIP is able to stimulate progesterone secretion. In the present study the effect of human VIP (900 pmol/kg body weight per h i.v. during 30 min) on steroidogenesis in six female volunteers was investigated. The experiments were performed between the 6th and 14th day of their menstrual cycle, and peripheral venous blood was collected before, during and after infusion of VIP. The concentrations of VIP, oestradiol, progesterone, testosterone, androstenedione (AD), dihydrotestosterone (DHT), dehydroepiandrosterone sulphate (DHAS), sex hormone binding globulin (SHBG) and cortisol were measured. The infusion of VIP was accompanied by a 15% increase (P less than 0.05) in serum oestradiol concentrations, from a mean basal concentration of 0.58 nmol/l. The concentrations of testosterone and DHT also increased significantly. No effect of VIP on progesterone, AD, DHAS, SHBG or cortisol was observed. In the light of the presence of VIP in nerve fibres of the steroid producing tissue, this stimulatory effect of VIP might reflect a direct action on the ovary or the adrenal gland.     

An intron in a ribosomal protein gene from Tetrahymena

We have cloned and sequenced a single copy gene encoding a ribosomal protein from the ciliate Tetrahymena thermophila. The gene product was identified as ribosomal protein S25 by comparison of the migration in two-dimensional polyacrylamide gels of the protein synthesized by translation in vitro of hybrid-selected mRNA and authentic ribosomal proteins. The proteins show strong homology to ribosomal protein S12 from Escherichia coli. The coding region of the gene is interrupted by a 979-bp intron 68 bp downstream of the translation start. This is the first intron in a protein encoding gene of a ciliate to be described at the nucleotide sequence level. The intron obeys the GT/AG rule for splice junctions of nuclear mRNA introns from higher eukaryotes but lacks the pyrimidine stretch usually found in the immediate vicinity of the 3' splice junction. The structure of the intron and the fact that it is found together with the well described self-splicing rRNA intron is discussed in relation to the evolution of RNA splicing.     

Cat pancreatic eicosapeptide and its biosynthetic intermediate. Conservation of a monobasic processing site.

Pancreatic eicosapeptide is synthesized together with the hormone pancreatic polypeptide in a common precursor in the major endocrine cell type of the duodenal pancreas. This processing has been previously demonstrated in man and in the dog. In the present study the cat pancreatic eicosapeptide and a C-terminally extended form of this were isolated and characterized from acid/ethanol extracts of pancreas by gel filtration and reverse-phase h.p.l.c. The sequence homology in the C-terminal part of the eicosapeptides from different species was shown to continue to the other side of the monobasic cleavage site in the extended intermediate form, whereas the end of the extension differed both in chain length and amino acid sequence. It is concluded that the processing site in the intermediate form of the pancreatic eicosapeptide is an example of a proline-directed monobasic cleavage site that has been conserved during evolution.     

Fluid balance in exercise dehydration and rehydration with different glucose-electrolyte drinks

After exercise dehydration (3% of body weight) the restoration of water and electrolyte balance was followed in 6 male subjects. During a 2 h rest period after exercise, a drink of one of four solutions was given as 9 X 300 ml portions at 15 min intervals: control (C-drink), high potassium (K-drink), high sodium (Na-drink) or high sugar (S-drink). An exercise test (submaximal and supramaximal work) was performed before dehydration and after rehydration. Dehydration reduced plasma volume by 16%, a process reversed on resting even before fluid ingestion began, due to release of water accumulated in the muscles during exercise. After 2 h rehydration, plasma volume was above the initial resting value with all 4 drinks. The final plasma volumes after the Na-drink (+14%) and C-drink (+9%) were significantly higher than after the K- and S-drinks. The Na-drink favoured filling of the extracellular compartment, whereas the K- and S-drinks favoured intracellular rehydration. In spite of the higher than normal plasma volume after rehydration, mean heart rate during the submaximal test was 10 bpm higher after rest and rehydration than in the initial test, and was not different between the drinks. The amount of work which could be performed in the supramaximal test (105% VO2max) was 20% less after exercise dehydration and subsequent rest and rehydration than before. This reduction was similar for all drinks, and may be due to a decreased muscle glycogen content (70% of initial) at the time of the second test.     

Effect of captopril on kidney function in insulin-dependent diabetic patients with nephropathy.

The influence of angiotensin II on kidney function in diabetic nephropathy was assessed by studying the effect of 12 weeks'' monotherapy with captopril (25-50 mg twice a day) in 16 hypertensive insulin dependent diabetic patients with persistent albuminuria. In an initial one week randomised single blind trial of captopril versus placebo, captopril (for nine patients) reduced arterial blood pressure from 148/94 (SD11/6) to 135/88 (8/7) mm Hg (p less than 0.05) and albuminuria from 1549 (range 352-2238) to 1170 (297-2198) micrograms/min (p less than 0.05), while glomerular filtration rate remained stable. No significant changes occurred in seven patients treated with placebo. During the 12 weeks of captopril treatment arterial blood pressure in all patients fell from 147/94 (11/6) to 135/86 (13/7) mm Hg (p less than 0.01), albuminuria fell from 1589 (range 168-2588) to 1075 (35-2647) micrograms/min (p less than 0.01), and glomerular filtration rate fell from 99 (SD19) to 93 (25) ml/min/1.73 m2 (p less than 0.01). The renin-angiotensin system showed suppressed plasma concentrations of angiotensin II and increased concentrations of angiotensin I and renin. The study showed that glomerular filtration rate is not dependent on angiotensin II, that captopril reduces albuminuria, probably by lowering glomerular hypertension, and that captopril represents a valuable new drug for treating hypertension in diabetics dependent on insulin with nephropathy.