Nucleotide sequence and evolution of the orangutan ε globin gene region and surrounding Alu repeats

Summary We have mapped and sequenced the globin gene and seven surrounding Alu repeat sequences in the orangutan globin gene cluster and have compared these and other orangutan sequences to orthologously related human sequences. Noncoding flanking and intron sequences, synonymous sites of , , and globin coding regions, and Alu sequences in human and orangutan diverge by 3.2%, 2.7%, and 3.7%, respectively. These values compare to 3.6% from DNA hybridizations and 3.4% from the globin gene region. If as suggested by fossil evidence and molecular clock calculations, human and orangutan lineages diverged about 10–15 MYA, the rate of noncoding DNA evolution in the two species is 1.0–1.5×10^–9 substitutions per site per year. We found no evidence for either the addition or deletion of Alu sequences from the globin gene cluster nor is there any evidence for recent concerted evolution among the Alu sequences examined. Both phylogenetic and phenetic distance analyses suggest that Alu sequences within the and globin gene clusters arose close to the time of simian and prosimian primate divergence (about 50–60 MYA). We conclude that Alu sequences have been evolving at the rate typical of noncoding DNA for the majority of primate history.Presented at the FEBS Symposium on Genome Organization and Evolution, held in Crete, Greece, September 1–5, 1986     

Calcium-dependent anion channel in the water mold,Blastocladiella emersonii

Summary Injection of depolarizing current into vegetative cells of the water moldBlastocladiella emersonii elicits a regenerative response that has the electrical characteristics of an action potential. Once they have been taken past a threshold of about –40 mV, cells abruptly depolarize to +20 mV or above; after an interval ranging from several hundred milliseconds to a few seconds, the cells spontaneously return to their resting potential near –100 mV. When the action potential was analyzed with voltage-clamp recording, it proved to be biphasic. The initial phase reflects an influx of calcium ions through voltage-sensitive channels that also carry Sr²⁺ ions. The delayed, and more extended, phase of inward current results from the efflux of chloride and other anions. The anion channels are broadly selective, passing chloride, nitrate, phosphate, acetate, succinate and even PIPES. The anion channels open in response to the entry of calcium ions, but do not recognize Sr²⁺. Calcium channels, anion channels and calcium-specific receptors that link the two channels appear to form an ensemble whose physiological function is not known. Action potentials rarely occur spontaneously but can be elicited by osmotic downshock, suggesting that the ion channels may be involved in the regulation of turgor.     

Haemagglutinating and Hydrophobic Properties of Corynebacterium (Eubacterium) Suis

Corynebacterium (Eubacterium) suis strains from boars and sows haemagglutinated erythrocytes of different animal species (calf, guinea pig, poultry, pig, and human). The haemaigglutination was man nose resistant (MR) and was neither inhibited by L-fucose nor D-galactose. The hydrophobicity measured by salt aggregation test (0.1–0.9 mol/1 (NH4)2SO4) and the hydrophobic interaction chromatography test (90 % retention in octyl sepharose) together with the haemagglutinating activity, indicated the presence of fimbriae on the bacteria. The haemagglutinating and hydrophobic properties were heat-sensitive (60°C for 10 min) suggestive of the presence of a protein structure. Two types of fimbria-tion were demonstrated by electron microscopy. Fetuin and glyco^ protein inhibited the haemagglutination, whereas porcine mucin was without any effect. These results indicate that branched glycoproteins might be important receptors for these fimbriae. The pathogenic aspects of C. suis are discussed, based on recent acquired knowledge of the effect of other pyelonephritogenic bacteria.     

Elevated levels of erythrocyte hypoxanthine phosphoribosyltransferase associated with allelic variation of murine Hprt

Murine stocks with wild-derived hypoxanthine phosphoribosyltransferase (HPRT) A alleles (Hprt a) have erythrocyte HPRT activity levels that are approximately 25-fold (Mus musculus castaneus) and 70-fold (Mus spretus) higher than those of laboratory strains of mice with the common Hprt b allele (Mus musculus: C3H/HeHa or C57B1/6). Since the purified HPRT A and B enzymes have substantially similar maximal specific activities (64 and 46 units/mg of protein, respectively), we infer that these HPRT activity levels closely approximate the relative levels of HPRT protein in these cells. Red blood cells of HPRT A and B mice have similar levels of adenine phosphoribosyltransferase activity (APRT; EC 2.4.2.7) and reticulocyte percentages, which suggests that the elevated levels of HPRT in erythrocytes of HPRT A mice are not secondary consequences of abnormal erythroid cell development. The HPRT activity levels in reticulocytes of HPRT B mice are approximately 35-fold higher than the levels in their erythrocytes and approach the HPRT activity levels in reticulocytes of HPRT A mice. Thus, the marked differences in the levels of HPRT protein in erythrocytes of HPRT A and B mice result from differences in the extent to which the HPRT A and B proteins are retained as reticulocytes mature to erythrocytes. The substantial and preferential loss of HPRT B activity from reticulocytes is paralleled by an equivalent loss of HPRT immunoreactive protein (i.e., CRM) from that cell, and we infer that the HPRT B protein is degraded or extruded as reticulocytes mature to erythrocytes.(ABSTRACT TRUNCATED AT 250 WORDS)     

Association of the M blood group system with bovine mastitis

Associations of the 11 bovine blood group systems with mastitis were examined in Red Danish dairy cattle. The mastitis status was followed during three lactational periods. A significant effect of the M blood group system on mastitis incidence was observed in the first and second lactation periods and a lower frequency of mastitis is found among animals lacking the M' factor as compared to those having the M' blood group factor. The significance of these results are discussed in view of the close relation between the M blood group system and the bovine lymphocyte antigens (BoLA), and the expected effect of eliminating the M' gene from the breed is estimated. Among the remaining 10 blood group systems, the T' system was the only system showing an overall effect on mastitis, and only in first and third lactation. However, the T' system was inconsistent with regard to the effect of the T' gene on the various mastitis diagnoses.     

Transposon donor plasmids, based on ColIb-P9, for use in Pseudomonas putida and a variety of other gram negative bacteria

Summary The properties of pLG221, a derivative of the ColIb plasmid carrying the transposon Tn5 are described. This plasmid can be used to introduce Tn5 by conjugation from Escherichia coli into a variety of Gram negative bacteria outside the host range for maintenance of ColIb. Plasmid pLG221, and a similar plasmid pLG223 carrying Tn10 may be of general utility as vectors for transposon-mediated mutagenesis in a variety of Gram negative bacteria.     

Immunochemical Characterization of Pyruvate Dehydrogenase Complex in Rat Brain

Pyruvate dehydrogenase complex (PDHC) in rat brain was studied immunochemically, using antibodies against the bovine kidney PDHC, by immunoblotting, immunoprecipitation, inhibition of enzyme activity, and enzyme-linked immunoabsorbent assay (ELISA). The immunoblots showed that the antibodies bound strongly to the alpha peptide of the pyruvate dehydrogenase (E1) component, and to the dihydrolipoyl transacetylase (E2) and the dihydrolipoyl dehydrogenase (E3) components of PDHC. A similar immunoblotting pattern was observed in all eight brain regions examined. On immunoblotting of the subcellular fractions, these PDHC peptides were observed in mitochondria and synaptosomes but not in the postmitochondrial supernatants. This agrees with other evidence that brain PDHC is localized in the mitochondria. These results, together with those from sodium dodecyl sulfate-polyacrylamide gel electrophoresis of the immunoprecipitin, also showed that the alpha E1, beta E1, and E3 peptides of rat brain PDHC are very similar in sizes to those of the bovine kidney PDHC, being 42, 36, and 58 kD, respectively. The size of the E2 peptide, 66 kD, is different from that of bovine kidney E2, 73 kD. The relative abundance of PDHC protein in nonsynaptic mitochondria was compared by enzyme activity titration and ELISA. Both methods demonstrated that the amount of PDHC antigen in the mitochondria from cerebral cortex is greater than that in the olfactory bulb mitochondria. This is consistent with the results of the activity measurement. The ELISA also showed that the PDHCs in both mitochondrial populations are antigenically similar.(ABSTRACT TRUNCATED AT 250 WORDS)     

Influence of pregnancy and sex steroids on concentration, motor effect and receptor binding of VIP in the rabbit female genital tract

The influence of sex steroid and pregnancy on the tissue concentration, uterine motor effect and receptor binding of VIP has been studied in the female genital tract of pregnant rabbits and oophorectomized rabbits during progesterone and/or oestrogen substitution. The concentration of immunoreactive VIP was high in the vagina and cervix, and lower in the uterine body of both pregnant and non-pregnant rabbits. A significant decrease in the VIP concentration (pmol/g wet weight) of the uterine body was observed toward term of pregnancy. The total uterine content of VIP, however, seems unchanged. Treatment of oophorectomized rabbits with ovarian steroids had no effect on the VIP concentration. The sensitivity for and potency of VIP on the relaxation of uterine muscle was significantly higher in oophorectomized rabbits treated with a combination of progesterone and oestrogen than in control rabbits. No difference was observed between non-pregnant and pregnant rabbits. The degradation and binding affinity for 125I-labelled VIP was highest in oophorectomized rabbits substituted with both oestrogen and progesterone. In the pregnant rabbits, the amount of receptors was decreased near term. In conclusion, sex steroids are able to influence the motor effect of VIP at the receptor level, but have no effect on the VIP concentration in the female genital tract.     

Differences in reactivity of confluent and nonconfluent cultures of human endothelial cells toward thrombin-stimulated platelets or heparinized salt solution

Summary This study examined whether nonconfluent endothelial cell cultures reacted differently than confluent ones toward thrombin-stimulated platelets or a heparinized salt solution. The adherence to the endothelial cell cultures of⁵¹Cr-labeled human platelets stimulated at different thrombin concentrations was studied. There was significantly higher adherence of stimulated platelets to nonconfluent cultures compared with confluent ones. This was confirmed by scanning electron microscopy, which also revealed a tendency for the platelets to adhere at the cell periphery. Electron microscopy also showed that thrombin-stimulated platelets induced endothelial cell contraction. Part of the peripheral endothelial cell surface toward the bottom of the culture dish was inverted, facing the lumen of the dish. This phenomenon was particularly seen in nonconfluent cultures. When⁵¹Cr-labeled endothelial cultures were incubated with a mildly injurious fluid as heparinized sodium acetate and 20% serum, at 20° C for 30 min, the nonconfluent cultures showed significantly more cell detachment and release of⁵¹Cr than the confluent ones. We conclude that under the conditions of the present experiments there are differences in the reactivity of confluent and nonconfluent endothelial cell cultures. These differences probably reflect biological dissimilarities. In experiments where properties of cultured endothelium are studied, care should be taken that the degree of confluency is standardized.