Reactivation and Dedifferentiation of Differentiated Murine Erythroleukemic Cell Nuclei

A murine erythroleukemic cell line, 745 A4-TG, deficient in hypoxanthine-guanine-phosphoribosyl transferase, can be induced with 3 mM hexamethylene bisacetamide to yield at least 50% of cells undergoing irreversible erythroid differentiation and finally losing capacity for cell divisions. The effects of such induced differentiation of 745 A4-TG on its ability to form viable and proliferating hybrids when fused with 3T3 1T22 fibroblasts were investigated. We found that when the induced 745 A4-TG cells were used, more continuously proliferating hybrids were obtained than could be accounted for by the residual uninduced cells which remained in these induced preparations. This suggests that some of the induced 745 A4-TG cells, when fused with 3T3 1T22 reverted from the induced phenotype of a limited capacity for cell proliferation to an uninduced state of continuous proliferation. This observation was further confirmed with the use of fully differentiated 745 A4-TG cells, which were obtained after selection with a bromodeoxyuridine suicide treatment to eliminate the uninduced and the partially differentiated cells in the preparations. When these selected, fully differentiated cells, as characterized by their lack of proliferation capacity and thymidine kinase activity, were fused with 3T3 1T22 (also deficient in thymidine kinase), it was found that not only were viable hybrid colonies obtained in a selection medium, which precluded the proliferation of either parental cells, but these hybrids continued to proliferate for more than two months in selection medium. These data thus confirmed that some fully differentiated erythroleukemic nucleus components in the hybrids were reactivated to regain capacity for cell proliferation and to dedifferentiate to synthesize thymidine kinase for survival in the selection medium. The lack of hemoglobin synthesis by these hybrids also indicates dedifferention of these murine erythroleukemic components in the hybrids.     

LIGHT ABSORPTION BY PLANTS AND ITS IMPLICATIONS FOR PHOTOSYNTHESIS

The preceding account has attempted to examine the interactions between light absorption and photosynthesis, with reference to both unicellular and multicellular terrestrial and aquatic plants. There are, however, some notable plant groups to which no direct reference has been made, e.g. mosses, liverworts and lichens. Although many have similar optical properties to terrestrial vascular plants (Gates, 1980) and apparently similar photosynthetic responses (see Green & Snelgar, 1982; Kershaw, 1984) they may possess subtle, as yet unknown differences. For instance, the lichen thallus has a high surface reflectance although the transmittance is virtually zero (Gates, 1980; Osborne, unpublished results). It is envisaged, however, that differences in optical properties between species will reflect differences in degree not kind. Although not all variation in photosynthesis is due to differences in light absorption a number of accounts suggest that this is a contributing factor. Variations in leaf absorptance have been found to account for most of the variation in leaf photosynthesis at low J_is (see Ehleringer & Björkman, 1978a; Osborne & Garrett, 1983). There is, however, little direct experimental evidence on light absorption and photosynthesis in either microalgal species or aquatic macrophytes. We also do not know over what range of incident photon flux densities photosynthesis is determined largely by changes in light absorption. Plants growing under natural conditions also experience large diurnal and seasonal fluctuations in J_i, unlike species grown under laboratory conditions. The occurrence of transitory peaks in J_i tends to overshadow the fact that the average J_i is often lower than the J₁ required to saturate photosynthesis, i.e. 1500–2000 μmol m^-2 s^-1, depending on the growth treatment. Using the data of Monteith (1977) and I W m²= 5 μmol m^-2 s^-1, and with photosynthetically active radiation 50% of total solar radiation, the daily mean value for Britain is approximately 450 μmol m^-2 s^-1, with a maximum in June of 1000μmol m^-2 s^-1 and a minimum during the winter of 75 μmol m^-2 s^-1. Such values could be even lower on shaded understory leaves and considerably lower for aquatic species. Based on average values of net photosynthesis for a terrestrial plant leaf, light saturation would only be expected in June while for most of the year the average values would lie largely on the light-limited portion of the photosynthesis light response curve. Although the daily average values in tropical climates may be higher during the winter months, they are remarkably similar throughout the world for the respective summers in the northern and southern hemispheres, because the increased daylength at high latitudes compensates for the lower J_is. The expected lower dark respiration rates during the winter may also partially offset the effects of a lower light level. There is therefore a trade-off between high J_is for a short period of time against a lower J_i for a longer period of time. We might expect different photosynthetic responses to these two very different conditions. Importantly, a low J_i with a long daylength may enable a plant to photosynthesize at or near its maximum photon efficiency for most of the day. Although the response of the plant to fluctuations in J_i is complicated because it is affected by the previous environmental conditions, this may indicate that light absorption has a much greater significance under natural conditions, particularly for perennial species. The bias in many laboratories towards research on terrestrial vascular plants also tends to ignore the fact that a number of multicellular and unicellular aquatic species survive in very low light environments. Furthermore, the direct extrapolation of photosynthetic responses from measurements on single leaves to those of whole plants is clearly erroneous. Although this is obvious, many physiological ecologists have attributed all manner of things to the photosynthetic responses of ‘primary’ leaves. Most researchers have ignored problems associated with composite plant tissues and internal light gradients. Clearly caution is required in interpreting the photosynthesis light-response curve of multicellular tissues based on biochemical features alone. Also, the importance of cell structure on light absorption and photosynthesis has generally been ignored and attributed solely to the effects of structural features on CO₂ diffusion. In doing so the work of two or three generations of plant physiologists has been ignored. Haberlandt (1914) at the turn of the century probably first implicated the role of cell structure in leaf optics, and Heath (1970) stressed that in order to completely understand the role of light in photosynthesis we need to know the flux incident on the chloroplast itself. Even this suggestion may need modification because of the capacity of the internal chloroplast membranes for scattering light. It is worth emphasizing the importance of light gradients within tissues and their role in regulating photosynthesis, particularly at light saturation. Measurements of light gradients are fraught with problems because of experimental difficulties and the majority (few) are based on reflectance and transmittance measurements. Seyfried & Fukshansky (1983) have shown that light incident on the lower surface of a Cucurbita cotyledon produced a larger light gradient than light incident from above, indicating the importance of the spatial arrangement of the tissues with respect to the light source. Also, light incident on the lower surface of leaves of Picea sitchensis was less ‘effective’ in photosynthesis than light from above (Leverenz & Jarvis, 1979). Clearly, two tissues could have the same gross absorptance but different photosynthetic rates because of differences in the internal light environment. Fisher & Fisher (1983) have recently found asymmetries in the light distribution within leaves, which they related to asymmetries in photosynthetic products due to differences in solar elevation. Such modifications in light distribution could be important for a number of solar-tracking species. Changes in light absorption are brought about by a whole gamut of physiological, morphological and behavioural responses which serve to optimize the amount of light absorbed. Perhaps the simplest way of regulating the amount of light absorbed is by restricting growth either to particular times of the year or to conditions when the light climate is favourable. We are still largely ignorant of many details of these modifications. In particular, differences in tissue structure such as the size and number of vacuoles or the effects of organelles on the scattering component of the internal light environment of photosynthetic tissues are not understood. A better understanding of the interaction of light with plants in aquatic systems is also required. It is unfortunate that light-absorptance measurements are not routinely made in photosynthetic studies, and this is quite clearly a neglected area of study. That these measurements are not made is even more surprising, since techniques have been available for over sixty years (Ulbricht, 1920). Absorptance measurements are of particular importance in the photosynthetic adaptation of microalgae, where only a small proportion of the incident photon flux density is absorbed. For multicellular species more detailed information is required on internal light gradients and their variability. Light-absorptance measurements are also important in any study relating kinetic data on CO₂ fixation to in vivo photosynthesis, especially when there are large variations in the morphology and structure of the photosynthetic organ.     

Codon usage in the vertebrate hemoglobins and its implications

A study of codon usage in vertebrate hemoglobins revealed an evolutionary trend toward elevated numbers of CpG codon boundary pairs in mammalian hemoglobin alpha genes. Selection for CpG codon boundaries countering the generally observed CpG suppression is strongly suggested by these data. These observations parallel recently published experimental results that indicate that constitutive expression of the human alpha-globin gene appears to be determined by regulatory information encoded within the structural gene. The possibility is raised that, in the absence of selection, CpG decay can be used to date the evolutionary origin of a mammalian alpha pseudogene from its active alpha gene.      

Evolution of crystallins: expression of lens-specific proteins in the blind mammals mole (Talpa europaea) and mole rat (Spalax ehrenbergi)

The mole (Talpa europaea; Insectivora) and the mole rat (Spalax ehrenbergi; Rodentia) both have degenerated eyes as a convergent adaptation to subterranean life. The rudimentary eye lenses of these blind mammals no longer function in a visual process. The crystallin genes, which display a lens-specific expression pattern, were studied in these blind mammals and in related species with normal eyes by hybridizing their genomic DNAs with probes obtained from cDNA clones for alpha A-, alpha B-, and beta Bp-crystallins from calf and gamma 3- crystallin from the rat. For all crystallin genes examined, the hybridization signals of mole and mole rat genomic DNA were comparable, respectively, with those of shrew and of rat and mouse, normal-vision representatives of the orders Insectivora and Rodentia. The expression of the crystallins at the protein level was tested by using antiserum specific for alpha-crystallin in immunofluorescence reactions on lens sections of mole and mole rat eyes and by using antisera against the beta- and gamma-crystallins on sections of the mole eye. All antisera gave positive fluorescence reactions exclusively with lens tissue of these blind mammals, indicating that the crystallins are still normally expressed despite the fact that these lenses have had no function in a visual process in these mammals for at least many million years. These findings apparently imply that some unknown selective advantage has conserved the crystallin genes and their expression after the loss of normal function of the lenses.      

Role of Peroxidase when Hydroxyproline-rich Protein in Plant Cell Walls is increased by Ethylene

Ethylene may control the growth of plant cells by regulating hydroxylation of specific wall proteins.     

Cell Separation and Anatomy of Abscission in the Oil Palm, Elaeis guineensis Jacq.

Shedding of the fruit of the oil palm takes place in two co-ordinatedstages. The first, a cell separation event at a pre-defined,positionally differentiated abscission zone at the base of thefruit, is followed by further cell separation in peripheraltissue at the junction with the rudimentary androecial ringand the tepals. The position of the second separation is determinedby the age and ripeness of the fruit and the degree of pressureto which it is subjected; it is also dependent upon completionof the first stage. Implications of this unusual two stage separationprocess are discussed. Key words: Abscission, cell separation, anatomy, oil palm, Elaeis guineensis     

The evolutionary history of the amylase multigene family in Drosophila pseudoobscura

In Drosophila pseudoobscura, the amylase (Amy) multigene family is contained within a series of inversions, or gene arrangements, on the third chromosome. The Standard (ST), Santa Cruz (SC), and Tree Line (TL) inversions are central to the phylogeny of arrangements, and have clusters of other arrangements derived from them. The gene arrangements belonging to each of these three clusters have a characteristic number of Amy genes, ranging from three in ST to two in SC to one in TL. This distribution pattern can reflect a history of either duplications or deletions, although the data available in the past did not permit a decision between these alternatives. We provide unambiguous evidence that three Amy genes were present before the divergence of the ST, SC, and TL arrangements. Thus, the current status of the Amy multigene family is the result of deletions in the TL and SC arrangements, which created three new pseudogenes: TL Amy2-psi, TL Amy3-psi, and SC Amy3- psi. Analysis of pseudogene sequences revealed that, in the SC and ST arrangements, pseudogene evolution has been retarded, most likely due to the homogenization effect of gene conversion. Finally, by determining the original copy number, we have reconstructed the evolutionary history of the Amy multigene family and linked it with the evolution of the central gene arrangements.