Munc-18–1 and cysteine string protein (csp) in pinealocytes of the gerbil pineal gland

Mammalian pinealocytes contain several synaptic membrane proteins which probably play a role in the targeting and exocytosis of secretory vesicles, in particular of synaptic-like microvesicles (SLMVs). The latter are considered as the endocrine equivalent of neuronal synaptic vesicles. By means of immunocytochemical techniques and immunoblot analyses, we now show that two further key components of the molecular apparatus regulating neurotransmitter release are present in the gerbil pineal gland, i.e., munc-18–1 and cysteine string protein (csp). In addition to varicosities of nerve fibres, munc-18–1 and csp could be localized to pinealocytes where both proteins were markedly enriched in process swellings. When using antibodies against csp for an immunogold electron-microscopic study of pinealocytes, gold particles consistently decorated profiles of pleomorphic SLMVs. Interestingly, we found that also the cytosolic protein munc-18, which is partially recruited to the plasmalemma in neurons, was associated to a significant extent with SLMVs of pinealocytes and synaptic vesicles of neurons, respectively. This localization implies that munc-18 at least partially exerts its regulatory functions while being bound to secretory vesicle membranes. Our results indicate that in endocrine cells such as pinealocytes the synaptic proteins munc-18–1 and csp play essential roles during the life cycle of SLMVs.     

Cation sensitivity and kinetics of guard-cell potassium channels differ among species

in ward rectifying g uard c ell K ⁺ c hannel, GCKC1_in, from three major crop plants Solanum tuberosum L., Nicotiana tabacum L., and Vicia faba L. Selecting guard cells for our analyses we aimed to test whether K⁺ channels of the same cell type differ among species. The channels shared basic features including voltage-dependence, selectivity and single-channel conductance. They activated at hyperpolarization (V _1/2 ≈ −164 mV) with single channels of 7 pS underlying the whole-cell current. The channel density in S. tuberosum was higher than in V. faba and N. tabacum while the activation and deactivation kinetics were faster in the latter two species. Among different monovalent cations the K⁺ channels discriminated strongly against Na⁺, Li⁺, and Cs⁺. The sensitivity to Cs⁺ was similar for the three species. Extracellular Ca²⁺ blocked the V.␣faba K⁺ channel at concentrations ≥1 mM but only affected its functional homologs in S. tuberosum and N.␣tabacum at higher concentrations and more-negative membrane potentials. Like the differences in Ca²⁺-sensitivity, protoplasts from the three species differed remarkably in their response towards extracellular pH changes. Whereas protons neither altered the open probability nor the kinetic parameters of the V. faba GCKC1_in, in S. tuberosum and N. tabacum this cation affected the voltage-dependent properties strongly. An increase in proton concentration from pH 8.5 to 4.5 shifted the potential of half-maximal open probability to less-negative values with a maximum effect around pH 6.2. The pH modulation of the K⁺ channels could be described assuming a two-state model where the open and closed channel can be protonated. The observed differences in cation-sensitivity and voltage-dependent kinetics between K⁺ channels reflect the diversification of guard-cell channels that may contribute to species-specific variations in the control of stomatal aperture. Received: 19 July 1997 / Accepted: 2 October 1997     

Toxicity screening of materials from buildings with fungal indoor air quality problems (Stachybotrys chartarum)

Samples of building materials visibly contaminated with moisture-related fungi (drywall, fiberglass, wallpaper, wood) were tested with indirect (FFL) and direct (MTT) cytotoxicity screening tests that are particularly sensitive toStachybotrys chartarum toxins. In addition, microscopic, chemical, immunochemical (Roridin A enzyme immunoassay) and mycological culture analyses were performed. In all cases in which building occupants had reported verifiable skin, mucous membrane, respiratory, central nervous system or neuropsychological abnormalities, cytotoxicity was identified. Results of a cytotoxicity screening test of field samples, such as the direct MTT test method, will give investigators of health problems related to indoor air quality problems important toxicity information.     

Involvement of the PS03 gene of Saccharomyces cerevisiae in intrachromosomal mitotic recombination and gene amplification

Using a genetic system of haploid strains of Saccharomyces cerevisiae carrying a duplication of the his4 region on chromosome III, the pso3-1 mutation was shown to decrease the rate of spontaneous mitotic intrachromosomal recombination 2- to 13-fold. As previously found for the rad52-1 mutant, the pso3-1 mutant is specifically affected in mitotic gene conversion. Moreover, both mutations reduce the frequency of spontaneous recombination. However, the two mutations differ in the extent to which they affect recombination between either proximally or distally located markers on the two his4 heteroalleles. In addition, amplifications of the his4 region were detected in the pso3-1 mutant. We suggest that the appearance of these amplifications is a consequence of the inability of the pso3-1 mutant to perform mitotic gene conversion.     

Cellular prion protein in the bovine mammary gland is selectively expressed in active lactocytes.

The cellular prion protein (PrP(c)) is a highly conserved glycoprotein with a still enigmatic physiological function. It is mainly expressed in the central nervous system but accumulating data suggest that PrP(c) is also found in a broad spectrum of non-neuronal tissue. Here we investigated the cell-type-related PrP(c) expression in the bovine mammary gland by using immunohistochemistry (IHC), ELISA, Western blot, and real-time RT-PCR. Specific immunostaining of serial sections revealed that PrP(c) is selectively localized in mammary gland epithelial cells. Particularly strong expression was found at the basolateral surface of those cells showing active secretion. Results obtained by RT-PCR and ELISA complemented IHC findings. No correlation was found between the level of PrP(c) expression and other parameters such as age of the animals under study or stage of lactation.     

Genetic adaptation in emergence time ofClunio populations to different tidal conditions

Summary 1. The emergence times of intertidalClunio-species (Diptera, Chironomidae) are correlated with special tidal conditions in such a way that the immediately following reproduction of the short-lived imagos can take place on the exposed habitat.2. If the habitat of aClunio-species is situated in the middle tidal region and exposed twice a day by the tidal cycle (T = 12.4 h), a tidal rhythm of emergence with an average period of 12.4 hours may result (example:Clunio takahashii).3. If the habitat is located in the lower tidal zone, exposed only at about the time of the spring tides, a semilunar rhythm of emergence is expected (examples:Clunio marinus andClunio mediterraneus). These semilunar rhythms are correlated with certain conditions of low tide which occur at the coastal locations every 15 days at about the same time of day. The semilunar rhythm is therefore exactly characterized by two dates: the lunar emergence time (a few successive days around full and new moon) and the diurnal emergence time.4. According to experimental investigations on the control of the emergence rhythm, the midges are able to determine both dates in advance.5. Coastal populations differ in their lunar and diurnal emergence times. These differences correspond to the time of low tide which exists at each location during the emergence days of the semilunar rhythm.6. Crossbreeding between stocks of different populations showed that the differences in diurnal emergence time are gene-controlled.

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Genetische Adaptation der Schlüpfzeiten vonClunio-Populationen an verschiedene Gezeitenbedingungen

Kurzfassung Die Schlüpfzeiten der in der Gezeitenzone lebendenClunio-Arten (Diptera, Chironomidae) sind mit bestimmten Wasserstandsbedingungen synchronisiert, und zwar derart, daß die unmittelbar anschließende Fortpflanzung der kurzlebigen Imagines auf dem trockengefallenen Habitat stattfinden kann. Wenn das Habitat einerClunio-Art in der mittleren Gezeitenzone liegt und parallel zu dem halbtägigen Gezeitenzyklus (T = 12,4 h) zweimal täglich auftaucht, dann kann sich eine 12,4stündige Schlüpfperiodik einstellen (Beispiel:Clunio takahashii). Wenn das Habitat in der unteren Gezeitenzone liegt und nur um die Zeit der Springtiden auftaucht, dann ist eine 15tägige (semilunare) Schlüpfperiodik zu erwarten (Beispiele:Clunio marinus undC. mediterraneus). Diese 15tägige Schlüpfperiodik ist synchronisiert mit bestimmten Niedrigwasserbedingungen, die an einem Küstenort alle 15 Tage jeweils um die gleiche Tageszeit auftreten. Sie wird daher durch zwei Daten eindeutig gekennzeichnet: (1) die lunaren Schlüpftage (wenige aufeinanderfolgende Tage um Voll- und Neumond) und (2) die tägliche Schlüpfzeit. Wie experimentelle Untersuchungen über die Steuerung der Schlüpfperiodik zeigten, können die Tiere beide Daten richtig vorausbestimmen. Die einzelnen Küstenpopulationen unterscheiden sich allerdings in Anpassung an die örtlichen Gezeiten- und Standortbedingungen recht auffällig in ihren lunaren und täglichen Schlüpfzeiten. Kreuzungsversuche zwischen Laboratoriumsstämmen verschiedener Populationen belegen, daß die Unterschiede in der täglichen Schlüpfzeit genkontrolliert sind.

     

Mapping of gal minus-mutants by transducing lambda dg carrying deletions of the gal-region

Summary To map numerousgal ^--mutants ofE. coli, advantage is taken of the fact that transducing dg's can carry different amounts of bacterial DNA of the host from which they originated (Adler andTempleton, 1963).A method is described with which a large number of transducing dg can be easily isolated, differing from each other with respect to the amount of bacterial DNA of thegal-region. By observing whethergal ⁺-colonies can arise as the result of recombination betweengal ^--mutants and dg's carrying deletions in thegal-region, so far 104 kinaseless mutants and 96 transferaseless mutants could be ordered into 26 groups. The mapping-tests were done by spotting the mutants with 52 HFT-lysates of dg's lacking more or less of the kinase- or the kinase- and transferase gene.