Somatic Embryogenesis in Cultured Carrot Cells

Somatic embryogenesis in cultured plant cells is an ideal system for investigating the whole process of differentiation and development from single cells to whole plants, and especially the molecular mechanism of expression of totipotency. This review reports recent progress the studies on somatic embryogenesis.     

Protein Synthesis during Photocontrolled Progression of the Cell Cycle in Single-celled Protonemata of the Fern Adiantum Capillus-veneris

Protein synthesis during photoinduced, synchronous progression of the cell cycle in single-celled protonemata of the fern Adiantum capillus-veneris was studied by tracer techniques. Nuclei of the protonemata were labelled with ³H-thymidine during spore germination so that the amount of ³H incorporated into the TCA-insoluble fraction of the cells could be used as a measure of the cell number in each sample. The rate of the incorporation of ¹⁴C-amino acids into TCA-insoluble materials was not significantly varied at different stages of the cell cycle or by treatment with blue light. Extracts of cells labelled with ³⁵S-methionine at various times after the transfer from red light condition (G₀) to darkness (G₁ to S) were analyzed by two-dimensional gel electrophoresis. At least 3 of about 200 spots showed significant changes in intensity on fluorograms. Spot A (molecular weight 20,000, isoelectric point 6.3) was detectable only in early G₁, whereas spot B (molecular weight 19,500, isoelectric point 6.3) was found only in the late G₁ and S phases. When the cells were exposed to blue light before the dark incubation, the times of disappearance of spot A and appearance of spot B were advanced depending upon the progression of the cell cycle but not upon the clock time.     

Factors controlling rates of nonphotochemical transformation of Pisum phytochrome in vitro

Nonphotochemical transformations of the far-red absorbing formof phytochrome, such as its decay or dark reversion to Pr, werestudied with solutions obtained from etiolated pea epicotyltissues at various steps of purification. At pH 7.8, the rateof dark Pfr reversion became significantly faster after thecrude extract was purified by gel filtration, but that of wellpurified solutions was quite low. Decay of Pfr was not seenduring any purification step at an alkaline pH, but it occurredin the acidic range of pH even in the presence of sulfhydrylcompounds. The rate of Pfr reversion was also influenced bypH; it increased with an increasing pH. Dark reversion of Pisum Pfr was confirmed to proceed in a short,rapid initial phase followed by a slow phase. (Received September 9, 1970; )     

Structure of the “Peroxy-Y Base” from Liver tRNA<Superscript>Phe</Superscript>

WE wish to report that reconstituted sperm whale myoglobin prepared by the method of Breslow¹ (except that pH 2 was found sufficient to remove all the haem) (I) crystallizes² in a different habit from those prepared by the method of Rossi-Fanelli et al.³ (II) using haemin of Sigma lot 77B-0220 and our own ⁵⁷Fe photoporphyrin preparation and the native myoglobin (III). Although all three form type A³ monoclinic prisms, the best developed plane is [001] for II and III, it is [100] for I. There seems to be great interest in reconstituted haemoproteins^4,5, so it is important that crystallization habit may be a sensitive test for subtle changes in protein structures.     

Potential fragmentation and loss of thermal habitats for charrs in the Japanese archipelago due to climatic warming

1. The upper thermal limits of the present distributions of two charr species, Dolly Varden, Salvelinus malma , and white-spotted charr, S. leucomaenis , in streams of the Japanese archipelago were examined using groundwater temperature as an index of thermal condition. The lower limits of the altitudinal distributions of Dolly Varden and white-spotted charr were delineated, respectively, by 8 and 16 °C groundwater isotherms.
2. The potential impact of future climatic warming on the geographical distribution, habitat extent and population fragmentation of each species was predicted at both the full archipelago and individual catchment levels.
3. For Dolly Varden, analysis at the full archipelago level indicated a loss of 27.6, 67.2, 79.6 and 89.6% of the current geographical range, respectively, for a 1, 2, 3 and 4 °C increase in mean annual air temperature. The present distribution area of white-spotted charr would likewise reduce by 4.1, 20.5, 33.8 and 45.6%, respectively.
4. Based on the analyses of three individual catchments, one for Dolly Varden and two for white-spotted charr, the lower habitat boundaries for the two charr species could be expected to rise increasingly to higher elevations in each catchment as warming proceeded. As a consequence, there would be large reductions in mean habitat area, with increasing habitat fragmentation followed by localized extinctions of the two species.     

Primers Designed for Amplification of Echinococcus multilocularis DNA Amplify the DNA of Encephalitozoon-Like Spores in the Polymerase Chain Reaction

ABSTRACT. Microsporidian spores were developed from cells which were grown in vitro from a human liver lesion which was due to larval Echinococcus multilocularis . The microsporidian spores developed in the same fashion as an Encephalitozoon cuniculi . The Encephalitozoon -like spores were completely separated on Percoll gradients. The separated spores contained DNA capable of amplification by two different primer sets designed for the polymerase chain reaction (PCR) of E. multilocularis DNA. However, the cell DNA from which microsporidium developed was thoroughly insensitive to the PCR using the E. multilocularis primer sets. The results strongly suggested that Encephalitozoon should be taken into consideration, when DNA isolated from larval E. multilocularis is analyzed.     

Potential fragmentation and loss of thermal habitats for charrs in the Japanese archipelago due to climatic warming

1. The upper thermal limits of the present distributions of two charr species, Dolly Varden, Salvelinus malma , and white-spotted charr, S. leucomaenis , in streams of the Japanese archipelago were examined using groundwater temperature as an index of thermal condition. The lower limits of the altitudinal distributions of Dolly Varden and white-spotted charr were delineated, respectively, by 8 and 16 °C groundwater isotherms.
2. The potential impact of future climatic warming on the geographical distribution, habitat extent and population fragmentation of each species was predicted at both the full archipelago and individual catchment levels.
3. For Dolly Varden, analysis at the full archipelago level indicated a loss of 27.6, 67.2, 79.6 and 89.6% of the current geographical range, respectively, for a 1, 2, 3 and 4 °C increase in mean annual air temperature. The present distribution area of white-spotted charr would likewise reduce by 4.1, 20.5, 33.8 and 45.6%, respectively.
4. Based on the analyses of three individual catchments, one for Dolly Varden and two for white-spotted charr, the lower habitat boundaries for the two charr species could be expected to rise increasingly to higher elevations in each catchment as warming proceeded. As a consequence, there would be large reductions in mean habitat area, with increasing habitat fragmentation followed by localized extinctions of the two species.     

Cycloheximide Insensitive Amoebo-Flagellate Transformation in Starved Amoebae of Physarum polycephalum

The requirement of protein synthesis for amoebo-flagellate transformation of Physarum polycephalum was re-examined. When amoebae were grown on nutrient agar in association with live food bacteria and harvested in mid-exponential phase of growth, it took ca. 2 hours for half the cells to form flagella after suspension in phosphate buffer. The transformation was completely inhibited by 5 μg/ml cycloheximide. To the contrary, when the amoebae in mid-exponential phase were starved for 3 hr on non-nutrient agar and then suspended in phosphate buffer, the duration required for this process was shortened to ca. 8 min and it was not inhibited by up to 100 μg/ml cycloheximide. A similar result was obtained using bactobolin, another inhibitor of protein synthesis. When amoebae were starved on non-nutrient agar containing 5 μg/ml cycloheximide, however, the starvation effect described above was not observed. The results indicate that protein(s) necessary for the transformation might be synthesized during the starvation period, and that the amoebo-flagellate transformation may or may not require concomitant protein synthesis depending upon preculture conditions.     

EFFECTS OF PHYTOCHROME AND BLUE-NEAR ULTRAVIOLET LIGHT-ABSORBING PIGMENT ON DURATION OF COMPONENT PHASES OF THE CELL CYCLE IN ADIANTUM GAMETOPHYTES

Single-celled protonemata of the fern Adiantum capillus-veneris, kept under continuous red light, grew with a very low rate of cell division, and the cell cycle was arrested in the early G₁ phase. Cell division was induced by transferring the protonemata to the dark after various light treatments, and the duration of component phases in the cell cycle was determined by a continuous-labelling technique with 3H-thymidine. Blue light irradiation greatly reduced the duration of the G₁ phase but did not affect that of other phases. The greater the fluence of blue light, the shorter was the duration of G₁ phase was observed. In contrast, a brief exposure of red-light-grown protonemata to far-red light given immediately before the dark incubation showed no effect on the duration of G₁ S and M phases but significantly extended that of the G₂ phase. The effect of far-red light on the G₂ phase was reversed by red light, and the effects of red and far-red light were repeatedly reversible. The progression in the M phase was shown by means of a time-lapse video system to be not at all influenced by any pre-irradiation described above.     

BIOASSAY OF ANTHERIDIOGEN IN LYGODIUM JAPONICUM

Antheridia were induced by exogenously applied GA₃ at concentrations between 10⁻⁶ and 3 × 10⁻⁴ M in very young filamentous protonemata of Lygodium japonicum grown in darkness; the longer the dark preculture of protonemata, the lower was the sensitivity of the protonemata to GA₃. Antheridial initials were discernible after 36 hr of GA₃ treatment in the most sensitive protonemata, and the timing of antheridial initiation was delayed with increasing protonemal age.
This quantitative response of the protonemata provided the basis for a new method of assaying gibberellins in terms of the degree of antheridial formation. According to this method, all the gibberellins tested and one of their precursors were active in inducing antheridia in the protonemata, and the activity spectrum of the gibberellins was as follows: GA₇>GA₄>GA₉>GA₃>GA₅>GA₁>GA₈.
The amounts of antheridiogen contained in conditioned media were measured by the present bioassay. A semi-logarithmic relation was shown between the percentage of antheridial formation and the concentration of conditioned medium within a certain dilution range. The amounts of antheridiogen secreted by the prothallia were quantitatively compared by transferring samples onto fresh media for a short period of time.