Expression of the Arabidopsis G-protein GPα1: purification and characterisation of the recombinant protein

The Arabidopsis G subunit, GP1, was expressedwithin Escherichia coli by co-transformation with the expressionvector and the dnaY gene which encodes tRNA^Arg _AGA/AGG. Isolation of the recombinant GP1 in a highly pureform could be achieved by a combination of anion exchange and dyeaffinity chromatography or by a single step affinity procedure viachromatography on 4-amino-anilido-GTP agarose. The recombinant proteinyielded by both procedures was highly active and bound GTPS withan apparent K_d in the nM range. GTPS binding wasstimulated two-fold in the presence of Zn²⁺ compared with that inthe presence of Mg²⁺, Mn²⁺ or Ca²⁺.Abbreviations: 4aaGTP, 4-amino-anilido-GTP; GTPS,guanosine- 5-(3-O-thiotriphosphate), PMSF,phenylmethylsulphonyl fluoride; PVDF, polvinylidene fluoride;rGP1, recombinant GP1     

Selection of Penicillium funiculosum Strains with High Glucose Oxidase Activity

Metabolic inhibitors, riboflavin, and end products of glucose oxidation were shown to be effective for the selection ofPenicillium funiculosum mutant strains with a high glucose oxidase activity. The incidence of positive mutations was highest in clones resistant to sodium azide, riboflavin, and -D-glucono--lactone. Enzyme activity in Penicillium funiculosum mutants was studied in submerged culture. The level of glucose oxidase synthesis in seven cultures was 24–56% higher than that in the parent strain of Penicillium funiculosum NMM95.132.     

Race-specific interactions between wheat genotypes and Indian cultures of stem rust

Summary Near isogenic/substitution lines of stem rust resistance genes in different backgrounds of Marquis, Chinese Spring and W 2691 and certain varieties with known genes for stem rust resistance were tested against each of 19 Indian cultures of stem rust races/biotypes (14, 15, 17, 21, 21A-1, 24, 34, 40, 40A, 42, 42B, 117, 117A, 117A-1, 122, 184, 194, 222 and 295). Sr 24 (Sear's 3D/Ag), Sr 24 (TR 380-27 4/3 Ag 14-White seeded recombinant with Agent type resistance), Sr 25 (Sear's 7D/Ag), Sr 26 (Eagle), Sr 26 (Knott's 6A/Ag translocation), Sr 27 (WRT 238-5), Combination line (Sr Tt1 + Sr 9b) were observed to be completely effective against all the 19 cultures tested. In addition, a number of lines, such as TAF2d (Sr Agi), Line W(Sr Tt2) and Combination III (Sr Tt1 + Sr 9e), were found to be effective against at least three of the most prevalent races (21, 40A and 117A-1) and a virulent race 122 in Indian natural population. Lines carrying genes other than Sr 2, Sr 9a, Sr 9f (Chinese Spring) and Sr 15 (Norka), and Line E were found to be resistant to one or more cultures of stem rust.The background effect upon the expression of a gene was observed by comparing the range of infection on single gene host lines in either different backgrounds and/or in cultivars with known genes for stem rust resistance against the 12 cultures of stem rust races found in India.     

Cloning,characterization and functional expression of a new beta-D-fructofuranosidase (Os beta fruct2) cDNA from Oryza sativa

A new cDNA (Osfruct2) encoding an acid –d-fructofuranosidase from rice has been cloned, sequenced and expressed in Pichia pastoris. The full-length cDNA is 2453 base pairs long and encodes a pre-pro-protein of 682 amino acids. The cDNA fragment coding for mature enzyme was sub-cloned into vector pPICZA for extracellular expression in the methylotrophic yeast. The recombinant product was purified by Ni²⁺-nitrilotriacetic acid agarose column and biochemically characterized. The enzyme could hydrolyze sucrose and raffinose. Molecular mass is 66 kDa. The activity optimum was at pH 4.8 and 40 °C.     

Physiological relationship between the temperate phages lambda and phi80

Summary This work deals with the ability of phage 80 to provide defective mutants of with their missing functions. Functions Involved in Recombination. As shown by others, the Int mechanism of 80 cannot excise prophage . However, 80 efficiently excises recombinants from tandem dilysogens, using its Ter mechanism. Likewise, the nonspecific mechanism Red is interchangeable between 80 and . Maturation of DNA by 80. The Ter recombinants excised by 80 from tandem dilysogens are packaged into a 80 protein coat. This contrasts with the fact, already mentionned by Dove, that 80 is extremely inefficient for packaging phage superinfecting a -lysogen. The latter result is also found when the helper phage is a hybrid with the left arm of (80hy4 or 80hy41 — see Fig. 1). However, the maturation of the superinfecting is much more efficient if the 80hy used as a helper has the att-N region of (like 80hy1). Conversely a with the att-N region of 80 (hy6 — see Fig. 1) is packaged more efficiently by 80 or 80hy4 than by 80hy1. It is suggested that the maturation of chromosome superinfecting an immune cell requires a recombination with the helper phage. Vegetative Functions. Among the replicative functoons O and P, the latter only can be supplied by 80. That N mutants are efficiently helped by 80 does not tell that 80 provides the defective with an active N product; the chromosomes are simply packaged into a 80 coat. This shows that 80 is unable to switch on the late genes of . That neither 80 nor any of the 80hy tested can provide an active N product is shown in a more direct way by their complete failure to help N ^-r₁₄; this phage carries a polar mutation which makes the expression of genes O and P entirely N-dependant. The maturation of a N ^- by 80 contrasts with the fact that mutants affected in late genes (A, F or H) are not efficiently helped by 80. This suggests that the products coded by these genes are not interchangeable between 80 and , and that packaging of DNA into 80 coats is possible but inhibited when late proteins are present in the cell. Activation of the Late Genes. Among the im ₈₀ h ⁺ hybrids tested, only 80hy41 is able to switch on the late genes of a N defective mutant. This hybrid differs from the other hybrids studied here, by the fact that it has the Q-S-R region of (see Fig. 1). The results are consistant with the view that the product of Q gene is sufficient for activating the late genes of a DNA. N would thus control the expression of late genes only indirectly by controlling the expression of gene Q (Couturier & Dambly have independantly reached the same conclusion, 1970). Furthermore the failure of 80 and of the 80hy1 and 80hy4 to activate the late genes of would imply that these phages are unable to provide an Q product active on the chromosome Reciprocally, switches on the late genes of prophage 80hy41, but not of prophages 80hy1 and 80hy4. This suggests that the initiation of late genes expression takes place at a main specific site located in the Q-S-R region of the chromosome. The expression of the late genes would thus be sequential, and proceed through the left arm only when steaky ends cohere. Similar conclusions were reached independantly by Toussaint (1969) and by Herskowitz and Signer (1970).

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Ce travail a été réalisé dans le cadre du contrat d'association Euratom-U. L. B. 007-61-10 ABIB et avec l'aide du Fonds de la Recherche Fondamentale Collective.     

Ultrastructure of spermatogenesis and sperm ofMulticotyle purvisi (Plathelminthes,Aspidogastrea)

Summary The ultrastructure of spermatogenesis is described for the first time in an aspidogastrean. The zone of differentiation which is usually formed during digenean spermiogenesis was not observed inMulticotyle purvisi. Instead, spermatid components are assembled within the common cytoplasmic mass before the outgrowth of spermatids. Microtubules, mitochondrion, nucleus and axonemes including their basal body regions, migrate from the cytoplasm into the spermatid which is pinched off at the level of the arching membrane. An unusual, complex structure of the basal body region is described. Intercentriolar bodies and striated rootlets are left behind and quickly disappear from the residual cytoplasm. Despite these atypical aspects, spermiogenesis results in the formation of mature sperm with the classical structure common to Digenea and Monogenea Polyopisthocotylea with the addition of some extra, non-cortical microtubules and a dense rod along part of the length of the sperm.Abbreviations used in the figures A cell type A, primary spermatogonium - AM arching membrane - AX axoneme - AZ attachment zone - B cell type B — spermatogonium - BB basal body region - C cell type C — spermatogonium - CEL central element - CI cisternae - CY cytophore - D cell type D — primary spermatocyte - DO doublet of microtubules - DR dark rod - E cell type E — multinucleate condensed cytophore - ER endoplasmic reticulum - G glycogen - GO Golgi body - I intercentriolar body - LB lamellate body - M mitochondrion - ME remnants of arching membranes - MT microtubules - N nucleus - R rootlet - S spermatid     

A simple method for maintaining high multiplication of Narcissus shoot cultures in vitro

A simple method for stimulating and maintaining high in vitro multiplication of Narcissus shoot clump cultures was developed. Shoot clumps were subjected either to normal cutting where leaves were trimmed to 20 mm in length at the beginning of each culture passage or to severe cutting where shoot clumps were cut down to the basal plate region removing all green tissue. Severe cutting at the beginning of each culture passage initially doubled the leaf multiplication, compared to normal cutting, but the difference between cutting treatments declined in successive passages. The improvement in leaf multiplication was maintained when shoot clumps were subjected to severe cutting only at every other culture passage, with no cutting in the alternate recovery passages. In vitro multiplication was increased by severe cutting in all seven Narcissus cultivars which were tested.Abbreviations NAA-1 naphthylacetic acid - BAP benzylaminopurine     

Average phase difference theory and 1∶1 phase entrainment in interlimb coordination

The dynamics of coupled biological oscillators can be modeled by averaging the effects of coupling over each oscillatory cycle so that the coupling depends on the phase difference between the two oscillators and not on their specific states. Average phase difference theory claims that mode locking phenomena can be predicted by the average effects of the coupling influences. As a starting point for both empirical and theoretical investigations, Rand et al. (1988) have proposed d/dt= — K sin ), with phase-locked solutions =arcsin( /K), where is the difference between the uncoupled frequencies and K is the coupling strength. Phase-locking was evaluated in three experiments using an interlimb coordination paradigm in which a person oscillates hand-held pendulums. was controlled through length differences in the left and right pendulums. The coupled frequency _c was varied by a metronome, and scaled to the eigenfrequency _v of the coupled system K was assumed to vary inversely with _c. The results indicate that: (1) and K contribute multiplicatively to (2) =0 or = regardless of K when =0; (3) 0 or regardless of when K is large (relative to ); (4) results (1) to (3) hold identically for both in phase and antiphase coordination. The results also indicate that the relevant frequency is _c/_v rather than _c. Discussion high-lighted the significance of confirming =arcsin(/K) for more general treatments of phase-locking, such as circle map dynamics, and for the 11 phase-entrainment which characterizes biological movement systems.     

Receptors for phorbol esters are primarily localized in neurons: Comparison of neuronal and glial cultures

Binding of [³H]PDB has been measured in the present study to determine the levels of protein kinase C in the neuronal and astrocytic glial cells in culture from rat brain. Binding of [³H]PDB to homogenates of cultured neuronal cells from the brains of normotensive and hypertensive rats was time-dependent and specific. The relative potency for competition by various phorbol esters to [³H]PDB binding was TPA > -PDD > POE > -PDD 4phorbol. Scatchard analysis showed that neuronal cultures from normotensive rat brains contained 2–3 fold more phorbol ester receptors compared with the glial cultures from the same brains. No differences in theK _d andB _max were observed between neuronal cultures from normotensive and spontaneously hypertensive rat brains. These studies suggest that the phorbol ester receptors are primarily localized in neuronal cells.     

Effects of growth environment on recombinant plasmid stability in Saccharomyces cerevisiae grown in continuous culture

A recombinant strain of Saccharomyces cerevisiae, containing a 2-m-fragment-based plasmid (pYEa4) was grown under non-selective conditions in continuous culture. The decrease in the population carrying the plasmid-encoded auxotrophic marker, LEU2, was examined under different physiological conditions. The difference in growth rate (µ) between plasmid-free and plasmid-containing cells and the rate of plasmid segregation (R) were determined using a non-linear regression technique. Loss rates were greater in defined glucose-limited cultures than in complex glucose-limited cultures. Plasmid loss was µ-dominated in cultures grown on defined media whereas µ and R were co-dominant in cultures grown on complex medium. Loss rates increased with increasing dilution rate in complex glucose-limited cultures. The reverse was found in defined glucose-limited cultures. Plasmid retention and loss kinetics determined from defined magnesium-limited cultures were not significantly different from those observed in defined glucose-limited cultures. Although plasmid retention in defined phosphate-limited culture was not significantly different from that in defined glucose-limited culture, reduced R and increased µ indicated an alternative physiological effect of phosphate limitation on plasmid stability.     

Problems in estimating growth rates of marine phytoplankton from short-term14C assays

Growth rate estimates () of phytoplankton populations that were sampled from nitrogen-limited continuous cultures and then incubated for short durations in batch culture with added¹⁴C-HCO₃ ^– were significantly different than steady-state growth rates () for 3 of 5 marine phytoplankton species. Two diatoms,Thalassiosira weissflogii andChaetoceros simplex, displayed virtually identical growth rates (=) over a wide range of, whereas for a third diatom,Phaeodactylum tricornutum, was overestimated by an average of 40% compared to. In contrast, was underestimated by the¹⁴C technique for the two remaining species: up to 40% at a steady-state of 1.0 day^–1 for the chlorophyteDunaliella tertiolecta and up to 100% at of 1.4 day^–1 for the haptophytePavlova lutheri. For the latter two species the divergence between and appeared to increase with increasing steady-state. A simple model of labeled and total carbon flow between the aqueous phase and cellular biomass was constructed to demonstrate that respiration was negligible when=, but was significant when>. In the cases in which<, a rapid physiological alteration presumably took place once the steady state was disturbed and cells were placed in the incubation chambers, which perhaps was related to the nutritional state of the cultures at the time of sampling. Questions thus are raised regarding our ability to measure accurately primary productivity from shipboard experiments with confined samples of phytoplankton from nutrient-impoverished waters that probably are less hardy than the laboratory cultures used in these studies.     

Microspore cultures as donor tissue for the initiation of embryogenic cell suspensions in barley

We have initiated embryogenic cell suspension cultures of barley (Hordeum vulgare L.) Igri from isolated microspore cultures. Data were obtained on the time required for establishment, frequency of establishment, i.e. number of calluses out of the total number of initiations giving rise to suspensions, and embryogenic capacity of the suspension cultures. For comparison, establishment of embryogenic cell suspensions from callus derived from immature zygotic embryos of Igri, Dissa and Golden Promise was also carried out. The results revealed that embryogenic suspension cultures were established in half the time and with a seven-fold higher frequency from microspore cultures than from zygotic embryo-derived calluses. The suspension cultures were still capable of embryo formation after two years. However, only albino plantlets were regenerated. For comparison, long term callus cultures derived from microspores, anthers and zygotic embryos were established. From the anther and zygotic embryo-derived callus cultures green plants were continuously regenerated, whereas the microspore-derived callus cultures lost this ability after the second subculture.     

Synthesis of Cationic Amphiphiles on the Basis of Deoxycholic Acid

Cationic derivatives of deoxycholic acid with N,N-dimethylenediamine, -aminocaproic acid, and pyridine as polar heads were synthesized. The cationic groups were linked to 3- and 12-hydroxy groups of the steroid moiety through ester or urethane bonds. Liposomal formulations of the compounds synthesized may be used for gene delivery in cells.     

Structural Peculiarities of Na+,K+-ATPase Isozymes from the Calf Brain

Functionally active preparations of Na⁺,K⁺-ATPase isozymes from calf brain that contain catalytic subunits of three types (1, 2, and 3) were obtained using two approaches: a selective removal of contaminating proteins by the Jorgensen method and a selective solubilization of the enzyme with subsequent reconstitution of their membrane structure by the Esmann method. The ouabain inhibition constants were determined for the isozymes. The real isozyme composition of the Na⁺ pump from the grey matter containing glial cells and the brain stem containing neurons was determined. The plasma membranes of glial cells were shown to contain mainly Na⁺,K⁺-ATPase of the 11 type and minor amounts of isozymes of the 22(1) and the 31(2) type. The axolemma contains 21 and 31 isozymes. A carbohydrate analysis indicated that 11 enzyme preparations from the brain grey matter substantially differ from the renal enzymes of the same composition in the glycosylation of the 1 isoform. An enhanced sensitivity of the 3 catalytic subunit of Na⁺,K⁺-ATPase from neurons to endogenous proteolysis was found. A point of specific proteolysis in the amino acid sequence PNDNR⁴⁹² Y⁴⁹³ was localized (residue numbering is that of the human 3 subunit). This sequence corresponds to one of the regions of the greatest variability in 1-, 2-, 3-, and 4-subunits, but at the same time, it is characteristic of the 3 isoforms of various species. The presence of the 3 isoform of tubulin (cytoskeletal protein) was found for the first time in the high-molecular-mass Na⁺,K⁺-ATPase 31 isozyme complex isolated from the axolemma of brain stem neurons, and its binding to the 3 catalytic subunit was shown.     

Effects of the potential allelochemical α-asarone on growth,physiology and ultrastructure of two unicellular green algae

The effects of the natural phenylpropanoid -asarone on growth pattern, photosynthesis, respiration and cell structure of two microalgae have been investigated. In cultures ofAnkistrodesmus braunii -asarone decreases in the medium and induces a lag in growth. Both phenomena were dependent on the number of cells inoculated. By contrast, in cultures ofSelenastrum capricornutum a constant decrease of the growth rate at all inocula was observed and only a slight decrease of -asarone in the medium occurred. In both algae -asarone caused an initial inhibition of photosynthesis, followed by a resumption of control values. The respiratory rate ofA. braunii was not significantly affected by -asarone, whereas inS. capricornutum respiration lowered to 60% of the control in the first 48 h and subsequently rose to values exceeding the controls by 20%. Ultrastructural observations carried out 24 and 72 h after the addition of -asarone showed modifications of cell wall inA. braunii, an increase in the number of mithocondrial profiles per cell section inS. capricornutum, and an accumulation of electron-dense deposits in the vacuoles of both algae.Author for correspondence     

Microbial transformation of 1,6-dichloro-1,6-dideoxy-β,D-fructofuranosyl-4-chloro-4-deoxy-α,D-galactopyranoside (TGS) by soil populations

Summary Soil microorganisms from one site were shown to be consistently capable of the transformation of 1,6-dichloro-1,6-dideoxy-,d-fructofuranosyl-4-chloro-4-deoxy-,d-galactopyranoside (TGS) in laboratory batch cultures. With fresh soils, all of the available chloride ions were released from the molecule. Subcultures of a TGS-dehalogenating bacterial community produced a progressive decline in the dehalogenating capabilities towards the substrate. The soil organisms did not utilise TGS as a carbon source. The transformation was achieved by co-metabolism and was probably supported by an unknown component in the soil. Four bacterial species were isolated from the TGS-dehalogenating soil community: twoBacillus species, anAcinetobacter group isolate and aMicrococcus group isolate. Thin-layer chromatography confirmed the disappearance of the chlorosugar and high-performance liquid chromatography demonstrated that neither of the constituent monosaccharides—1,6-dichlorofructose nor 4-chlorogalactosucrose was accumulated as an intermediate.

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Resumen Microorganismos de suelo de cierto lugar demostraron consistemente ser capaces de realizar la transformación de 1,6-dicloro-1,6 dideoxi--D-fructofuranosil-4-cloro-4-deoxi-,D-alactopiranosa (TGS) in culturas de laboratorio de tipo discontinuo. Con muestras frescas de suelo, todos los iones cloruro fueron liberados de la molecula. Subculturas de una comunidad bacterial capaz de dehalogenizar TGS produjeron una declinación progresiva de la capacidad de dehalogenizar el substrato. Los microorganismos no utilizaron el TGS como fuente de carbono. La transformación se realiza por co-metabolismo y probablemente se base en un componente del suelo, no determinado. Cuatro especies bacteriales fueron aisladas de la comunidad de bacterias de suelo con capacidad de dehalogenar el TGS: dos especies deBacilo, unaAcinelobacteria y unMicrococo. Estudios de cromatografía de capa delgada confirmaron la desaparición del clorosacárido, y estadios de cromatografía liquida demostraron que ninguno de los componentes monoscáridos — 1,6-diclorofructuosa y 4-clorogalactosucrosa — eran acumulados como productors intermedios.

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Résumé Les microorganismes du sol d'un certain endroit ont été démontrés être capable, sans exception, de la transformation de 1,6-dichloro-1,6-dideoxy-,D-fructofuranosyl-4-chloro-4-deoxy-,D-galactopyranoside (TGS) en cultures de laboratoire du type discontinu. Avec des prélèvements frais du sol, tous les ions disponibles de chlorure ont été libérés de la molécule. Des souscultures d'une communauté bactérienne capable de déhalogeniser le TGS ont produit un déclin progressif de la capacité de déhalogeniser le substrat. Les microorganismes du sol n'ont pas utilisé le TGS comme source de carbone. La transformation s'est accomplie par cometabolisme et, probablement, s'est basée sur un component indéterminé du sol. Quatre espèces bactériennes ont été isolées de la communauté de bactéries du sol capable de déhalogeniser le TGS: deux espèces deBacillus, unAcinetobacter et unMicrococcus. Des études de chromatographie de couches fines ont confirmées la disparition du chlorosaccharide, tandis que des études de chromatographie liquide de haut rendement ont démontrées que, des monosaccharides constituants, ni 1,6-dichlorofructose ni 4-chlorogalactosucrose, n'ont été accumulés comme produits intermédiaires.

     

Testosterone and progesterone metabolism in the central nervous system: Cellular localization and mechanism of control of the enzymes involved

Summary This paper summarizes the most recent data obtained in the authors' laboratory on the metabolism of testosterone and progesterone in neurons and in the glia.1. The activities of 5-reductase (the enzyme that converts testosterone into dihydrotestosterone; DHT) and of 3-hydroxy steroid dehydrogenase (the enzyme that converts DHT into 5-androstane-3,17-diol; 3-diol) were first evaluated in primary cultures of neurons, oligodendrocytes, and type-1 and type-2 astrocytes, obtained from the fetal or neonatal rat brain. The formation of DHT and 3-diol was evaluated incubating the different cultures with labeled testosterone or labeled DHT as substrates. The results obtained indicate that the formation of DHT takes place preferentially in neurons; however, also type-2 astrocytes and oligodendrocytes possess considerable 5-reductase activity. A completely different localization was observed for 3-hydroxysteroid dehydrogenase; the formation of 3-diol appears to be prevalently, if not exclusively, present in type-1 astrocytes; 3-diol is formed in very low yields by neurons, type-2 astrocytes, and oligodendrocytes. Moreover, the results indicate that, in type 1 astrocytes, both 5-reductase and 3-HSD are stimulated by coculture with neurons and by the addition of neuron-conditioned medium, suggesting that secretory products released by neurons might intervene in the control of glial cell function.2. Subsequently it was shown that, similarly to what happens when testosterone is used as the substrate, 5-reductase, which metabolizes progesterone into 5-pregnane-3,20-dione, (DHP), shows a significantly higher activity in neurons than in glial cells; however, also type-1 and type-2 astrocytes as well as oligodendrocytes possess some ability to 5-reduce progesterone. On the contrary, 3-hydroxysteroid dehydrogenase, the enzyme which converts DHP into 5-pregnane-3-ol-20-one (THP), appears to be present mainly in type-1 astrocytes; much lower levels of this enzyme are present in neurons and in type-2 astrocytes. At variance with the previous results obtained using androgens as precursors, oligodendrocytes show considerable 3-hydroxysteroid dehydrogenase activity, even if this is statistically lower than that present in type-1 astrocytes. The existence of isoenzymatic forms of the enzymes involved in androgen and progesterone metabolism is discussed.     

Ongoing ultradian activity rhythms in the common vole,Microtus arvalis,during deprivations of food,water and rest

Summary The timing mechanism underlying ultradian (2–3 h) activity patterns in the common vole, Microtus arvalis, was studied using behavioural deprivation experiments. These were aimed at distinguishing between a homeostatic control mechanism, in which the rhythmic behaviour itself is part of the causal loop, and a clock mechanism, independent of the behaviour.In 175 experiments, deprivation of food during 3 ultradian cycles in (subjective) daytime did not result in significant changes in the ultradian periodicity of attempts to obtain the food, compared with ad lib. access to food and water. A minor, but significant increase in ultradian activity time () occurred in the course of the deprivation, but this was compensated by a shorter ultradian rest (). These results were obtained both in intact animals (n = 24), which showed ultradian and circadian rhythmicity in behaviour, and in animals (n = 21) with electrolytic lesions aimed at the suprachiasmatic nuclei (SCN), which lacked the circadian modulation of behaviour. Simultaneous deprivation of water and food in 8 voles without circadian rhythmicity during 40 experiments also did not lead to any change in the ultradian periodicity of feeding attempts.Rest deprivation was studied in 5 SCN lesioned voles, by forcing running wheel activity to continue following spontaneous running. Thus, the experimental activity bout was artificially lengthened to 2–9 h in 67 experiments. The onset of the subsequent rest episodes occurred independent of the duration of the preceding . The duration of was dependent on the preceding, experimental in a periodic fashion. The interval experimental (=lengthened +following ) was equal to one, two or three times the control (obtained on nonexperimental days). This result fits the prediction of a clock model and is in conflict with a monotonicincrease of with , as expected in a homeostatic, restorative process.It is concluded that the ultradian timing of activity in the common vole can be explained neither by homeostatic hunger or thirst mechanisms nor by homeostatic rest/activity regulation. The results strongly suggest an independent clock system generating ultradian feeding rhythms in the common vole.Abbreviations DD continuous darkness - LD light-dark regime - LL continuous light - RCA retrochiasmatic area - ARC arcuate nucleus - SCN suprachiasmatic nuclei - ultradian period - ultradian activity time - ultradian rest time     

Cellular localization of glycoside hydrolases inBacteroides fragilis

The localizations of six glycosidases produced byBacteroides fragilis—-glucosidase, -glucosidase, -galactosidase, -galactosidase, -N-acetylglucosaminidase, and -l-fucosidase—were studied. Cell fractions and cell extracts were obtained by Triton X-100 release, by disruption by freeze-pressing and sonication, and by osmotic release. Isoelectric focusing of a cytoplasmic and of a Triton X-100 extract of the cell wall fraction was performed and revealed differences in the relative distribution of differently charged forms of -N-acetylglucosaminidase. -Galactosidase and alkaline phosphatase were used as cytoplasmic and periplasmic markers, respectively. It is concluded that inB. fragilis -glucosidase is periplasmic, -l-fucosidase and -galactosidase are cytoplasmic, and -n-acetylglucosaminidase is cell associated and bound to the cell envelope by hydrophobic interactions. -Glucosidase and -galactosidase are localized cytoplasmically and/or located in the cell envelope.