Isolated deficiency of immunocytochemically detected somatostatin in Snell dwarf, but not in “little,” mice

Immunocytochemical staining of the neuropeptide somatostatin was evaluated in the brains of two growth hormone-deficient mouse mutants, Snell dwarf (dw/dw), and “little” (lit/lit). In Snell dwarf mice, in which GH is undetectable, an isolated somatostatin deficiency was observed in hypothalamic median eminence and in anterior periventricular hypothalamic neurons which are afferent to median eminence, compared to somatostatin staining in normal mice of the same strain (DW/?). Somatostatin staining in all other CNS areas in dwarfs was identical to that in normal mice. In contrast, in little mice, which exhibit 5–10% of normal GH levels, somatostatin expression was comparable between mutants and normal mice (LIT/?) in all CNS areas, including median eminence-afferent neurons in anterior periventricular hypothalamus. The results suggest that expression of somatostatin in hypophysiotropic CNS is dependent upon minimal pituitary GH secretion.     

Genesis and structure of the so-called “balbiani body” or “yolk nucleus” in the oocyte of dugesia dorotocephala (turbellaria,tricladida)

Oogenesis of the fresh-water triclad Dugesia dorotocephala has been studied by electron microscopical methods, with particular regard to the genesis and composition of the so-called “Balbiani body.” Its origin is clearly recognizable in young oocytes where the few mitochondria present seem to gather at the level of the perinuclear ooplasm. Here they surround dense masses of finely granular, fibrillar material probably coming from the nucleus. During the previtelloge ic period, mitochondria rapidly increase in number while the dense masses progressively dissolve. In the vitellogenic oocytes the Balbiani body shows its final configuration: it appears as a large area (up to 15-20 pm in diameter) consisting of innumerable densely packed mitochondria, some smooth vesicles and free ribosomes. This aggregate of cytoplasmic organelles remains unmodified in the mature oocytes. The function of the “Balbiani body” of D. dorotocephala is as yet unclear; it can only be asserted that it is not correlated with yolk production in which the endoplasmic reticulum and the Golgi complex are involved.     

Testosterone increases acetylcholine receptor number in the “levator ani” muscle of the rat

The “levator ani” muscle of male rats provides a neuromuscular system in which both the muscle and its motoneurons have high levels of androgen receptors. Two weeks of castration caused a 48% loss of acetylcholine receptors in this muscle. One week of testosterone propionate injections initiated one week after castration increased receptor number by 27% over untreated castrate levels. These changes paralleled changes in muscle protein content. In contrast, castration and testosterone treatments of castrates had no effect on total, Triton X-100-extractable acetylcholinesterase activity. This system may provide a useful model of synaptic plasticity.     

Manganese and copper promote the binding of dopamine to “serotonin binding proteins” in bovine frontal cortex

The authors previously reported that Fe²⁺ is capable of increasing the binding of dopamine and of serotonin to “serotonin binding proteins” which are present in soluble extracts from calf brain. In this study, it is shown that Mn²⁺ and Cu²⁺ are also capable of increasing the binding, but for dopamine only. As for Fe²⁺, Mn²⁺ and Cu²⁺ are likely to promote the binding by virtue of their ability to enhance the oxidation of dopamine into dopamine-O-quinone, a derivative which is known to undergo covalent association with sulfhydryl groups of proteins. Data such as the irreversible nature of the majority of the binding, the inhibitory action of reducing agents (sodium ascorbate) and of reagents which contain, or modify sulfhydryl groups (reduced glutathione) are compatible with such a mechanism. The three metal ions are also capable of inactivating part of the binding sites on SBP directly; this effect is more pronounced for Cu²⁺ than for Fe²⁺ and it is only weak for Mn²⁺. The Fe²⁺-mediated binding of dopamine is inhibited by the superoxide dismutase enzyme, and it was therefore suggested that Fe²⁺ enhances the oxidation of dopamine by virtue of its ability to produce superoxide radicals out of dissolved molecular oxygen. Such a mechanism does not appear to take place in the case of Mn²⁺ and Cu²⁺. Instead, it is likely that Cu²⁺ and dopamine form a complex which is highly susceptible towards oxidation by dissolved molecular oxygen. Mn²⁺, on the other hand, can easily be oxidized into Mn³⁺, which is capable to oxidize dopamine by itself. Chronic manganese intoxication (from exposure to manganese) and Wilson's disease (related to inadequate elimination of copper) go along with neurological symptoms which are very similar to those encountered in Parkinson's disease. Our data indicate that manganese and copper ions accelerate the oxidation of catecholamines to produce toxic quinones. These quinones could, at least in part, account for the degeneration of dopamininergic neurons in such pathologies.     

Suicide is not the inevitable outcome of “perpetual” selfing in tetrahymenines collected from natural habitats

A significant fraction of the Tetrahymena clones isolated from natural habitats self (mating occurs within a clone). Early attempts to study such clones failed because stable subclones were rarely, if ever, observed, and isolated pairs all died. Isozyme analysis revealed that these wild selfers were a diverse group; some were very similar to T. australis, a species with synclonal mating type determination and to T. elliotti, shown recently to have a karyonidal mating type system. One originally stable clone of T. australis included some selfing clones after a few years in our laboratory. Other clones manifested unique zymograms. Subclones isolated from 18 selfer strains were heterogeneous. All subclones of several selfers mated massively at each transfer through 100 fissions. Selfing among subclones of other selfers was highly variable or not observed. Although 77% of the pairs isolated died, and 9% of the pair cultures selfed, 15 selfers yielded some viable nonselfing “immature” progeny. Additional immature progeny were obtained by isolating pairs from macronuclear retention synclones. Although some “immature” progeny eventually selfed, most remained stable. Giemsa staining revealed macronuclear anlagen in nearly all mating pairs and some anomalies. Crosses among the F1 progeny clones of the T. elliotti selfers yield viability data comparable to those from crosses among normal strains. Perhaps perpetual selfing is a mechanism of getting rid of deleterious combinations of genes and uncovering better combinations in homozygous state by playing genetic roulette. © 1992 Wiley-Liss, Inc.     

The last of the Nomads. the last of the Ethnographies or “All Anthropologists are Liars”

Pintupi Country, Pintupi Self; Sentiment, Place, and Politics Among Western Desert Aborigines. By Fred R. Myers. Smithsonian Institute Press, Washington and Australian Institute of Aboriginal Studies, Canberra, 1986. Pp. 7+334; black and white diagrams. $32.95 (hard cover). ISBN 0–85575–171–1 hard cover — AIAS); ISBN 0–87474–690–0 (hard cover —Smithsonian).     

DNA “fingerprinting” and the genetic management of a captive chimpanzee population (Pan troglodytes)

DNA fingerprinting probes are cloned sequences which simultaneously detect a large number of similar hypervariable loci in the target DNA. The resulting highly polymorphic pattern visualized on an autoradiograph allows resolution of questions concerning individual identification and parentage. M13 bacteriophage has been used as a DNA fingerprinting probe for paternity ascertainment among captive chimpanzees housed in multi-male groups as part of the National Chimpanzee Breeding and Research Program. In 31 cases of unknown paternity where DNA samples for mother, offspring, and all potential sires were available, DNA fingerprinting with M13 resulted in the unambiguous assignment of paternity for all 31 infants. Knowledge of pedigrees among the captive-born animals is used to address several issues important in the genetic management of captive breeding colonies, including estimation of effective population size and of the rate of decline in genetic variability, variance in male and female reproduction, and the effect of social dominance on male reproductive success. Our analysis demonstrates the beneficial effects of genetic management by comparing the managed dedicated cohort to the Bastrop colony as a whole.     

Progesterone receptor associated with the “melanosome” fraction of Pleurodeles waltlii oocytes (urodela amphibian)

A proteinaceous receptor associated with the melanosome fraction of Pleurodeles waltlii oocytes binds progesterone with high affinity and limited capacity (K_D ≈ 5 · 10⁻⁸ M).     

Evidence for “Deleted” or “Silent” gene homozygous at the locus coding for the constant region of the γ3 chain

Three uncommon stable Gm haplotypes, Gm3;23;--, Gm1,2,17;..;-- and Gm1,17;..;-- have been transmitted through 3 generations of two related Lebanese and Syrian families. No pathological consequence was noted in seven individuals, aged 14--65, whose sera were deficient for all the allotypes carried by the IgG3 chains. Among the different genetic events which could have produced these haplotypes (alteration of a regulatory gene, point mutation, gene hybridization, gene deletion), it appears that a structural deletion is the most probable explanation. The observed data can be explained by either a partial or a total deletion of the constant portion of the IgG3 heavy chain.     

Synthesis and evaluation of two novel “mixed” chiral stationary phases deriving from tyrosine: Csps designed for the resolution of either π-acid or π-basic racemates

The synthesis and chromatographic evaluation of two novel chiral stationary phases (CSPs) deriving from (S)-tyrosine are reported. The chiral graft has been designed in order to bear both π-acid and π-basic sites, each one being connected to a distinct asymmetric centre. An intramolecular π-π interaction may take place within these CSPs, leading to an energetically favoured conformation of the chiral selector (CS). The enantiorecognition ability of these CSPs was investigated for various classes of either π-acid or π-basic racemates. It is shown that these CSPs are able to separate simultaneously π-acid and π-basic racemates. Finally, chiral recognition mechanisms and mobile phase optimization are discussed.     

Mutations in the ubiquitin binding UBZ motif of DNA polymerase eta do not impair its function in translesion synthesis during replication

Treatment of Saccharomyces cerevisiae cells with DNA-damaging agents elicits lysine 164-linked PCNA monoubiquitination by Rad6-Rad18. Recently, a number of ubiquitin (Ub) binding domains (UBDs) have been identified in translesion synthesis (TLS) DNA polymerases and it has been proposed that the UBD in a TLS polymerase affects its binding to Ub on PCNA and that this binding mode is indispensable for a TLS polymerase to access PCNA at the site of a stalled replication fork. To evaluate the contribution of the binding of UBDs to the Ub moiety on PCNA in TLS, we have examined the effects of mutations in the C2H2 zinc binding motif and in the conserved D570 residue that lies in the alpha-helix portion of the UBZ domain of yeast Poleta. We find that mutations in the C2H2 motif have no perceptible effect on UV sensitivity or UV mutagenesis, whereas a mutation of the D570 residue adversely affects Poleta function. The stimulation of DNA synthesis by Poleta with PCNA or Ub-PCNA was not affected by mutations in the C2H2 motif or the D570 residue. These observations lead us to suggest that the binding of Ub on PCNA via its UBZ domain is not a necessary requirement for the ability of polymerase eta to function in TLS during replication.     

On the application of polyelectrolyte “limiting laws” to the helix-coil transition of DNA. I. Excess univalent cations

A general theory of polyelectrolyte solutions is here used to calculate the differences in Gibbs free energy, enthalpy, and entropy between the coil and helix forms of DNA at any temperature and salt concentration. The salt has univalent cations and is assumed present in excess over the base concentration. The results are restricted to sufficiently dilute solutions. It is shown that the salt concentrations effect is entirely entropic in origin. When applied to the melting temperature, the calculations yield a relation between the enthalpy difference at the melting temperature and the slope of the plot of melting temperature vs. the logarithm of the salt concentration. In accord with observation, both the Gibbs free energy difference at any fixed temperature and the melting temperature are predicted to be linear functions of the log of the salt concentration. However, the theory is not in quantitative agreement with enthalpy data. Data on various colligative and transport properties of both helix and coil forms are reviewed in the text and in Appendix B, and good agreement is found with theory for both forms. No attempt is made to explain why the theory is quantitative for these properties but not for heat measurements. Finally, in Appendix A, an approximate calculation is made of the free energy contributions due to ionic effects not associated with the salt concentration.     

DNA binding and synapsis by the large C-terminal domain of ϕC31 integrase

The integrase (Int) from phage ϕC31 acts on the phage and host-attachment sites, attP and attB, to form an integrated prophage flanked by attL and attR. Excision (attL × attR recombination) is prevented, in the absence of accessory factors, by a putative coiled-coil motif in the C-terminal domain (CTD). Int has a serine recombinase N-terminal domain, required for synapsis of recombination substrates and catalysis. We show here that the coiled-coil motif mediates protein–protein interactions between CTDs, but only when bound to DNA. Although the histidine-tagged CTD (hCTD) was monomeric in solution, hCTD bound cooperatively to three of the recombination substrates (attB, attL and attR). Furthermore, when provided with attP and attB, hCTD brought these substrates together in a synaptic complex. Substitutions in the coiled-coil motif that greatly reduce Int integration activity, L460P and Y475H, prevented CTD–CTD interactions and led to defective DNA binding and no detectable DNA synapsis. A substitution, E449K, in full length Int confers the ability to perform excision in addition to integration as it has gained the ability to synapse attL × attR. hCTD^E449K was similar to hCTD in DNA binding but unable to form the CTD synapse suggesting that the CTD synapse is not essential but could be part of the mechanism that controls directionality.     

The thyrotropin receptor hinge region is not simply a scaffold for the leucine-rich domain but contributes to ligand binding and signal transduction

The glycoprotein hormone receptor hinge region connects the leucine-rich and transmembrane domains. The prevalent concept is that the hinge does not play a significant role in ligand binding and signal transduction. Portions of the hinge are redundant and can be deleted by mutagenesis or are absent in certain species. A minimal hinge will be more amenable to future investigation of its structure and function. We, therefore, combined and progressively extended previous deletions (Delta) in the TSH receptor (TSHR) hinge region (residues 277-418). TSHRDelta287-366, Delta287-371, Delta287-376, and Delta287-384 progressively lost their response to TSH stimulation of cAMP generation in intact cells, consistent with a progressive loss of TSH binding. The longest deletion (TSHRDelta287-384), reducing the hinge region from 141 to 43 amino acids, totally lost both functions. Surprisingly, however, with deletions extending from residues 371-384, constitutive (ligand-independent) activity increased severalfold, reversing the suppressive (inverse agonist) effect of the TSHR extracellular domain. TSHR-activating point mutations I486F and I568T in the first and second extracellular loops (especially the former) had reduced activity on a background of TSHRDelta287-371. In summary, our data support the concept that the TSHR hinge contributes significantly to ligand binding affinity and signal transduction. Residues within the hinge, particularly between positions 371-384, appear involved in ectodomain inverse agonist activity. In addition, the hinge is necessary for functionality of activating mutations in the first and second extracellular loops. Rather than being an inert linker between the leucine-rich and transmembrane domains, the TSHR hinge is a signaling-specificity domain.