Number and evolutionary conservation of alpha- and beta-tubulin and cytoplasmic beta- and gamma-actin genes using specific cloned cDNA probes

A partial library of cloned human DNA was screened for sequences represented on and specific to the X chromosome. The library was constructed from Bam HI-digested human DNA from cells with X chromosome polyploidy, and was cloned in pBR322. The screening was performed by individually hybridizing ³²P-labeled cloned plasmids to Southern blots containing Bam HI-digested DNA from mouse-human hybrid cells having the human X chromosome and from derivative hybrids lacking the human X. Of 45 clones assayed, 33 contained sequences homologous to ones represented many times on the X. In situ hybridization to metaphase chromosomes demonstrated that at least four of these clones were homologous to autosomes as well. Only one of the 18 clones of this kind tested cross-hybridized with another. Two recombinant plasmids were shown to be derived from the X chromosome and to be X chromosome-specific by three criteria: they hybridized to a single band in the Southern blots of Bam HI-digested DNA from hybrid cells containing the X chromosome; they hybridized to a band of the same molecular weight as did the inserted DNA fragment; and they showed a dosage effect when hybridized to Southern blots of Bam HI-digested DNA from XY and XXX cells. One of these hybridized as a single-copy or low-order reiterated sequence in a Cot analysis using male DNA as driver. Our methods can be applied to the identification of any chromosome-specific clone. The two X-specific clones identified here should be useful in investigating the mechanism of X inactivation and in isolating a Barr body.     

The effect of pentobarbital on the antinociceptive action of morphine in morphine-tolerant and non-tolerant rats

The interaction of sodium pentobarbital with morphine sulfate in both morphine-tolerant and non-tolerant rats was investigated using the tail-compression test for analgesia. Male Sprague-Dawley rats (300–350 g) were given pentobarbital (4, 8, or 16 mg/kg) 5 min before morphine (2, 4, 6, or 8 mg/kg). Control animals received two saline injections, or pentobarbital plus saline, or saline plus morphine. All injections were subcutaneous. Prior to the first injection, a baseline nociceptive threshold was determined for each rat by applying a modified micrometer to its tail and increasing the pressure until a squeak was elicited. Test readings were taken every half-hour for 2 hr beginning 30 min after the second injection. For the chronic studies, animals were first made tolerant to morphine by the administration of the narcotic twice a day for 3 days, increasing the dose from 10 to 50 mg/kg/injection. Identical testing procedures were then followed with these rats except that the test dose of morphine given on day 4 was in the range 8–128 mg/kg. It was found that Na pentobarbital, in the subanesthetic doses used, had neither antinociceptive nor hyperalgesic properties. Furthermore, the barbiturate had no effect on the antinociceptive action of morphine in either morphine-tolerant or non-tolerant rats.     

Lymphocytes express Ia antigens of foreign haplotype following treatment with neuraminidase

Evidence is presented which indicates that neuraminidase (NA) treatment of spleen cells both destroys old Ia antigens and reveals new Ia specificities which are not normally expressed by splenocytes. It was found that NA treatment unmasked alien I-A^k-like specificities on A.TH (I ^s ) spleen cells, and I^s-like antigens on A.TL (I ^k ) spleen cells. These conclusions were based on direct testing of NA-treated targets with a range of alloantisera and on cell-absorption experiments. Furthermore, the cellular distribution of NA-exposed antigens resembled that of convential Ia antigens, the new antigens being expressed on more than 90 percent of splenic B cells and a subpopulation of splenic T cells. However, although some of the antigens exposed by NA on A.TH cells appeared to resemble the Ia. 3 and 15 specificities, additional antigens were involved which did not correlate with any previously described Ia antigens.Sugar inhibition experiments demonstrated the NA-exposed antigens to be carbohydrate in nature, D-galactose being an effective inhibitor in these studies. The proportion of- and-linked D-galactose residues associated with the new antigens depended upon the target cell used and the anti-Ia serum tested. Furthermore, glycolipid extracts from lymphoid cells were shown to contain the NA-exposed antigens.Collectively, these results support the existence of carbohydrate-defined Ia antigens. The simplest interpretation of the findings is that NA clips off terminal sialic acid residues from carbohydrate-defined Ia antigens on the cell surface and exposes subterminal sugars which resemble antigens expressed by otherI-region haplotypes.     

Survival of patients treated for end-stage renal disease by dialysis and transplantation.

The results of treatment in 213 patients with end-stage renal disease who underwent hemodialysis, peritoneal dialysis or transplantation, or a combination, between 1962 and 1975 were analysed. Comparison by censored survival analysis showed significantly better (P less than 0.01) patient survival with the integrated therapy of dialysis and transplantation than with either form of dialysis alone. There was no significant difference in survival of males and females but survival at the extremes of age was poorer. Analysis of survival by major cause of renal failure indicated best survival in patients with congenital renal disease. Graft and patient survival rates at 1 year after the first transplantation were 42% and 69%. The major cause of death in this series was vascular disease but infection was responsible for 50% of deaths after transplantation. While integration of dialysis with transplantation produces best patient survival, this course is possible only when sufficient cadaver kidneys are available.     

Stereospecificity of peptide transport by germinating barley embryos

The stereospecific requirements for peptide transport in the scutellum of germinating barley (Hordeum vulgare) embryos are described. Replacement of an L-amino acid residue in a peptide by its D-stereoisomer decreases the affinity of the peptide for the transport site, leading to a reduction in transport. Substitution of a second D-residue reduces affinity still further. The extent to which transport is inhibited depends upon the position of the D-residue in the primary sequence, with D-residues at the C-terminus of the peptide having the greatest effect. Competition between D- and L-peptides indicates that they both enter via the same transport system. Although D-amino acids can be accumulated when presented as a peptide, these same D-residues are not transported when supplied as the free amino acids. L-Leu-D-leu is accumulated intact against a concentration gradient, indicating the operation of an active transport mechanism that can function without the involvement of peptidase activity.     

The properties of immobilized caldolysin, a thermostable protease from an extreme thermophile

Caldolysin, the extracellular thermostable metal-chelator-sensitive lytic protease from Thermus T-351 was immobilized to Sepharose 4B, CM-cellulose, and controlled pore glass (CPG). Although protein binding efficiencies were high (96, 88, and 95%), some loss of enzyme activity occurred on immobilization (26, 69, and 89%). The pH optimum of both CM-cellulose and CPG-immobilized Caldolysin was decreased by about one pH unit. The K(m) for Sepharose-Caldolysin was unchanged with respect to the free protease, while those for CM-cellulose-Caldolysin and CPG-Caldolysin were lower by approximately one order of magnitude. Immobilization to both Sepharose and CM-cellulose increased the thermostability of Caldolysin at high temperatures, while CPG-Caldolysin was less thermostable than the free protease.