PHOTOSYNTHETIC NITRITE REDUCTASE II. FURTHER PURIFICATION AND BIOCHEMICAL PROPERTIES OF THE ENZYME

1. Further purification of photosynthetic nitrite reductase (PNiR),which catalyzes the transfer of hydrogen (or electron) fromthe photolytic system to nitrite, is reported in this paper.Chromatography on DEAE- cellulose and Sephadex gel-filtrationwere effective for the purification of PNiR.
2. PNiR could befractionated into two components. It was inferredfrom the dataobtained that one of these components is identicalwith PPNR,and the other one may probably be a hitherto unreportedflavinenzyme containing FMN as prosthetic group.
3. The propertiesof these two components of PNiR were described,and the interrelationshipbetween these catalysts and possibleintermediary carriers ofthis electron transfer system was discussed.

¹Dedicated to Prof. H. TAMIYA on the occasion of his 60th birthday.This investigation was supported by a Grant in Aid for FundamentalScientific Research from the Ministry of Education (No. 407130-1961)and a Grant in Aid for Organized Scientific Research from theMinistry of Education (No. 95037-1960), which are gratefullyacknowledged here. The authors also wish to acknowledge thatthe progress of this study was facilitated by a Grant from theKAISEI-KAI. A preliminary report on this work was read beforethe 25th Annual Meeting of the Botanical Society of Japan (1960,Osaka).     

Mutation in the Structural Gene for Diphtheria Toxin carried by Temperate Phage β

Corynebacterium diphtheriae strains lyso-genic for phage β are able to produce diphtheria toxin. This article describes evidence suggesting that the toxin structural gene is part of the phage genome.     

L-Tyrosine carboxy-lyase of barley roots

_L-Tyrosine carboxy-lyase (E.C. 4. 1. 1. 25) was isolated fromroots of germinating barley (Hordeum vulgare). The enzyme requirespyridoxal phosphate for maximum activity. The optimum pH foractivity is about 7.0. The enzyme is inhibited by p-chloromercuribenzoateand hydroxylamine at 10^–3 M. Enzyme activity is foundin extracts from young roots, especially from those in earlystages of development, but not in extracts from shoots of thesame plant. Localization and changes in the amounts of _L-tyrosinecarboxy-lyase and aromatic amines in developing barley seedlingswere measured. Participation of carboxy-lyase in the formationof aromatic amines in barley roots is suggested. (Received July 17, 1970; )     

THE DISTRIBUTION OF FORMYLTETRAHYDROFOLATE SYNTHETASE IN PLANTS, AND THE PURIFICATION AND PROPERTIES OF THE ENZYME FROM PEA SEEDLINGS

1. Formyltetrahydrofolate synthetase (E. C. 6. 3. 4. 3) was foundto be widely distributed in higher plants and the high enzymeactivity was observed in green leaves of Brassica and Alliumspecies, spinach, and in pea seedlings. In pea seedlings, theenzyme activity changed during the course of germination, andmost of the enzyme activity was located in a soluble fractionof the cytoplasm.
2. The enzyme was labile and lost the activityrapidly, even whenstored at 5 in the presence of 0.1 M mercaptoethanol.It was,however, found that ammonium sulfate was very effectivein stabilizingthe enzyme activity.
3. The enzyme has been purifiedapproximately 500-fold from extractsof pea seedlings by treatmentswith ammonium sulfate, protaminesulfate, hydroxylapatite, calciumphosphate gel, and DEAE-cellulosecolumn chromatography.
4. Thepurified enzyme was specific for formate, ATP and FAH₄,andthe Michaelis constants for these reactants were 2.1 10^–2M, 5.1 10^–4 M, and 5.6 10^–3 M, respectively.
5. The optimum pH was found to be 8.0, and the optimal temperaturewas observed at 37. Both NH₄^$ and a divalent cation (Mg^SS orMn^SS) were required for the optimal activity.

¹ Studies on the Enzymatic Synthesis and Metabolism of FolateCoenzymes in Plants. II. (For the previous paper see reference(8)) A part of this paper was presented at the Meeting of theKansai Division of the Agricultural Chemical Society of Japan,Kyoto, January 29, 1966.     

Structure of forest canopies as related to their primary productivity

Some structural features of forest canopy were analysed in relationto their role in photosynthetic production by forest communitieswhich are thought to produce more organic matter than herbaceouscommunities under the same environment. 1) Leaf area density was found to be much smaller in forestthan in herbaceous canopies. 2) Light extinction in forest canopy followed the BEER-LAMBERT'Slaw as was found for herbaceous canopy, though the coefficientof light extinction (K) was relatively small in the former. 3) A geometrical model was proposed to account for the smallvalue of K and the resultant large leaf area index, based onthe characteristically clustered distribution of tree leavesin forest canopy. 4) Stratification in forest community was interpreted as theuneven vertical distribution of leaf area density. 5) Deterioration of leaf functions, such as photosynthesis andrespiration, toward the bottom of forest canopy was noticed. 6) An attempt to estimate total canopy photosynthesis is presentedtaking this into consideration. 7) A new method for estimating total canopy respiration is proposedand discussed. ¹Contributions from JIBP-PT No. 36. The essential parts of thispaper were presented by KIRA to the IBP symposium The BiologicalBasis of Productivity held at Varna, Bulgaria, on April 4, 1968. (Received August 15, 1968; )     

Comparison of contemporary mating patterns in continuous and fragmented Eucalyptus globulus native forests

While habitat fragmentation is a central issue in forest conservation studies in the face of broad-scale anthropogenic changes to the environment, its effects on contemporary mating patterns remain controversial. This is partly because of the inherent variation in mating patterns which may exist within species and the fact that few studies have replication at the landscape level. To study the effect of forest fragmentation on contemporary mating patterns, including effective pollen dispersal, we compared four native populations of the Australian forest tree, Eucalyptus globulus . We used six microsatellite markers to genotype 1289 open-pollinated offspring from paired fragmented and continuous populations on the island of Tasmania and in Victoria on mainland Australia. The mating patterns in the two continuous populations were similar, despite large differences in population density. In contrast, the two fragmented populations were variable and idiosyncratic in their mating patterns, particularly in their pollen dispersal kernels. The continuous populations showed relatively high outcrossing rates (86–89%) and low correlated paternity (0.03–0.06) compared with the fragmented populations (65–79% and 0.12–0.20 respectively). A greater proportion of trees contributed to reproduction in the fragmented ( de/d ≥ 0.5) compared with the continuous populations ( de/d  =   0.03–0.04). Despite significant inbreeding in the offspring of the fragmented populations, there was little evidence of loss of genetic diversity. It is argued that enhanced medium- and long-distance dispersal in fragmented landscapes may act to partly buffer the remnant populations from the negative effects of inbreeding and drift.     

Adult eclosion rhythm of the Indian meal moth Plodia interpunctella: response to various thermocycles with different means and amplitudes

In addition to photoperiod, thermoperiod (or thermocycle) might be an important Zeitgeber for entraining the circadian oscillator controlling adult eclosion rhythm in the Indian meal moth Plodia interpunctella Hübner (Lepidoptera: Pyralidae). This is confirmed by exposing larvae receiving diapause‐preventing treatments to various thermocycles with different means and amplitudes of temperature. The thermocycles investigated in the present study are TC 8 : 16 h, TC 12 : 12 h, TC 16 : 8 h and TC 20 : 4 h, where T and C represent thermophase (30 °C) and cryophase (20 °C), respectively. For all thermocycles, the peak of adult eclosion rhythm occurs at around the mid‐thermophase. This indicates that the larvae use both ‘temperature‐rise’ and ‘temperature‐fall’ signals to adjust the eclosion phase in each thermocycle. The absence (DD) or presence (LL) of light affects this time‐keeping system slightly under the given thermocycle. The rhythmic adult eclosion noted after exposure of larvae to 30 °C DD for 14 days is recorded in the thermocycles (TC 12 : 12 h, DD; mean temperature = 25 °C) with different amplitudes of 27.5/22.5 °C, 26.5/23.5 °C and 25.5/24.5 °C. The peak in adult eclosion advances in time as the amplitude of the temperature cycle decreases. In the temperature cycle of 25.5/24.5 °C, a peak occurs at the end of the cryophase, 2 h before the temperature‐rise. The adult eclosion rhythm is also observed under various thermocycles (TC 12 : 12 h, DD) consisting of different temperature levels (30 to 20 °C) with different amplitudes. It is found that the temporal position of the peak advances significantly when the amplitude of the thermocycle becomes lower.     

Cell to Cell Adhesion Systems in Xenopus laevis, the South African Clawed Frog I. Detection of Ca2+ Dependent and Independent Adhesion Systems in Adult and Embryonic Cells

The reaggregation kinetics of frog ( Xenopus laevis ) cells prepared using several different dissociation procedures were monitored. Two distinct modes of cell adhesion were revealed, one mediated by Ca²⁺ dependent cell-cell adhesion system (CDS) and the other mediated by Ca²⁺ independent one (CIDS).
CDS was detected in frog embryonic (two-cell stage to neurula) cells and in cells of epithelial cell lines (A6 and A8), while CIDS was detected only in A6 cells. Frog CDS was resistant to 0.01% tryptic digestion in the presence of 1 mM Ca²⁺ and CIDS was resistant to dilute (0.0001%) tryptic digestion. Cells with both mechanisms were prepared by dissociation with 1 mM EDTA and both mechanisms were absent in cells dissociated with 0.01% trypsin and 1 mM EDTA. Frog CDS and CIDS functioned in a temperature dependent and independent manner, respectively. Properties of frog CDS and CIDS revealed in this study were the same as those of mammalian or avian CDS and CIDS, respectively. Frog cells with CDS cross-adhered to mammalian epithelial (F9) cells but not to fibroblastic (V79) cells Ca²⁺ dependently. Monoclonal antibody ECCD-1 raised against mouse epithelial CDS blocked aggregation of frog A8 cells.
These results suggest the similarity between frog CDS and mammalian epithelial type CDS.     

Origins of Laboratory Mice Deduced from Restriction Patterns of Mitochondrial DNA

To determine the origins of laboratory mice, the restriction patterns of mitochondrial DNAs (mtDNAs) from various strains were compared with those of relevant subspecies and/or races of mus musculus . In most strains and substrains of laboratory mice examined (50/55), the cleavage patterns were identical to those of the European subspecies M. m. domesticus . Those that varied include two sublines of NZB, the strain NZC, and the Japanese strain RR. The NZB and NZC patterns were identical to that of the European subspecies M. m. brevirostris , which itself has restriction patterns similar to M. m. domesticus . On the other hand, the RR pattern was identical to M. m. molossinus -like mice trapped in Western China and slightly different from Japanese M. m. molossinus . These findings suggest that the strains NZB and NZC stemmed from a European founder stock which differed from the ancestral stocks of other laboratory strains and that the ancestral mice of the RR strain had been transported from China to Japan. Therefore, most laboratory strains of mice are derived from the European subspecies M. m. domesticus while M. m. brevirostris and M. m. molossinus have made minor contributions. M. m. musculus does not appear to have made any contribution.