The relative cellular levels of CP2a and CP2b potentiates erythroid cell-specific expression of the α-globin gene by regulating the nuclear localization of CP2c

CP2b activates α-globin expression in an erythroid cell-specific manner, through interaction with CP2c and PIAS1. Although CP2a is identical to CP2b except for lacking an exon encoding additional 36 amino acids and has the intrinsic DNA binding and transactivation properties, it does not exert any role in α-globin expression. Investigation of subcellular localization of exogenous CP2 proteins revealed that CP2a and CP2b were exclusively localized in the cytosol and nucleus, respectively. The CP2b-specific exon was in charge of the nuclear localization of CP2b. Interestingly, subcellular localization of CP2c was either in the nucleus or cytosol depending on the relative level of CP2a and CP2b although CP2c intrinsically localized in the cytosol in the absence of CP2a/CP2b. Finally, dramatic increment of hemoglobin expression was correlated with nuclear translocation of CP2c during MEL cell differentiation. Our data suggest that CP2b potentiate erythroid cell-specific α-globin expression by recruiting CP2c into the nucleus.     

Random mutagenesis in the large extrinsic loop E and transmembrane α-helix VI of the CP 47 protein of Photosystem II

The intrinsic chlorophyll-protein CP 47 is a component of Photosystem II which functions in both light-harvesting and oxygen evolution. Using the Escherichia coli mutator strain XL-1 Red, we introduced mutations at 14 sites in the large extrinsic loop E of CP 47 and its adjacent transmembrane -helix VI. Four mutant cell lines were recovered in which the histidyl residues 455H, 466H and 469H were altered. The cell lines H455T, H455Y, H469Y, and the double mutant F432L,H466R exhibited phenotypes that supported the identification of the histidyl residues 455H, 466H and 469H as chlorophyll ligands. Four additional mutant cell lines were recovered which contained mutations at positions 448R in the large extrinsic loop of CP 47. These mutants, R448K, R448Q, R448S, and R448W, exhibited variable phenotypes ranging from moderate alteration of photoautotrophic growth and oxygen evolution rates to a complete inhibition of these parameters. Those mutants exhibiting photoautotrophic growth and oxygen evolution capability under standard conditions were unable to grow photoautotrophically or evolve oxygen when grown at low chloride concentrations. Finally, a mutant cell line exhibiting a substitution at position 342G was recovered. The mutant G342D exhibited moderate alterations of photoautotrophic growth and oxygen evolution. In addition to these alterations, mutants were recovered in which deletions and insertions (leading to frame shifts) and stop codons were introduced. These mutants uniformly lacked the ability to either grow photoautotrophically or evolve oxygen.     

Functional analysis of the iron-stress induced CP 43′ polypeptide of PS II in the cyanobacterium Synechococcus sp. PCC 7942

Under conditions of iron-stress, the Photosystem II associated chlorophyll a protein complex designated CP 43, which is encoded by the isiA gene, becomes the major pigment-protein complex in Synechococcus sp. PCC 7942. The isiB gene, which is located immediately downstream of isiA, encodes the protein flavodoxin, which can functionally replace ferredoxin under conditions of iron stress. We have constructed two cyanobacterial insertion mutants which are lacking (i) the CP 43 apoprotein (designated isiA ^–) and (ii) flavodoxin (designated isiB ^–). The function of CP 43 was studied by comparing the cell characteristics, PS II functional absorption cross-sections and Chl a fluorescence parameters from the wild-type, isiA ^– and isiB ^– strains grown under iron-stressed conditions. In all strains grown under iron deprivation, the cell number doubling time was maintained despite marked changes in pigment composition and other cell characteristics. This indicates that iron-starved cells remained viable and that their altered phenotype suggests an adequate acclimation to low iron even in absence of CP 43 and/or flavodoxin. Under both iron conditions, no differences were detected between the three strains in the functional absorption crossection of PS II determined from single turnover flash saturation curves of Chl a fluorescence. This demonstrates that CP 43 is not part of the functional light-harvesting antenna for PS II. In the wild-type and the isiB ^– strain grown under iron-deficient conditions, CP 43 was present in the thylakoid membrane as an uncoupled Chl-protein complex. This was indicated by (1) an increase of the yield of prompt Chl a fluorescence (F_o) and (2) the persistence after PS II trap closure of a fast fluorescence decay component showing a maximum at 685 nm.Abbreviations Chl chlorophyll - CP 43, CP 47 and CP 43 Chl a binding protein complexes of indicated molecular mass - DCMU 3-(3,4-dichlorophenyl)-1,1-dimethylurea - F_m and F_m fluorescence when all PS II reaction centers are dosed in dark- and light-acclimated cells, respectively - F_o fluorescence when all PS II reaction centers are open in dark acclimated cells - F_v variable fluorescence after dark acclimation (F_m–F_o)     

Amino acid deletions in the cytosolic domains of the chlorophyll a-binding protein CP47 slow QA − oxidation and/or prevent the assembly of Photosystem II

The Photosystem II (PSII) core antenna chlorophyll a-binding protein, CP47, contains six membrane-spanning -helices separated by five hydrophilic loops: A–E. To identify important hydrophilic cytosolic regions, oligonucleotide-directed mutagenesis was employed to introduce short segment deletions into loops B and D, and the C-terminal domain. Four strains carrying deletions of between three and five residues were created in loop B. Two strains, with deletions adjacent to helices II and III, did not assemble PSII; however, the mutants (F123–D125) and (R127–S131) remained photoautotrophic with near wild-type levels of assembled reaction centers. In contrast, all deletions introduced into loop D, connecting helices IV and V, failed to assemble significant levels of PSII and were obligate photoheterotrophic mutants. However, deletions in the C-terminal domain did not prevent the assembly of PSII reaction centers although the mutant (S471–T473), with a deletion adjacent to helix VI, exhibited retarded Q_A ^– oxidation kinetics and the PSII-specific herbicide, atrazine, bound less tightly in the (S471–T473) and (F475–D477) strains. Deletions in the C-terminal domain also created mutants with large protein aggregates that were recognized by an antibody raised against the PSII reaction center D1 protein. Low-temperature fluorescence emission spectra of photoautotrophic strains carrying deletions in either the C-terminal domain or loop B did not provide evidence for impaired energy transfer from the phycobilisomes to the PSII reaction center. The data therefore suggest an important structural role for loop D in the assembly of PSII and a potential interaction between the C-terminal domain of CP47 and the PSII reaction center that, when perturbed, results in photoinduced protein aggregates involving the D1 protein.     

Kinetic Fluorescence Spectral Analysis of Core Antennas CP43 and CP47 of Photosystem Ⅱ with Ultrafast Time-resolved Technology

Ultrafast time-resolved fluorescence experiments have been performed with core antennas CP43 and CP47 of PS Ⅱ. Their dynamic fluorescence spectra were obtained with excitation wavelength 514.5 nm. For CP43, the emission spectrum was found to be from 640 to 780 nm with a peak at ～680 nm and the lifetime of fluorescence was 3.54 ns. For CP47, the emission spectrum was from 630 to 775 nm with a peak at ～691 nm and the fluorescence lifetime was 3.22 ns. The fluorescence emission efficiencies of Chl a in CP43 and CP47 were calculated to be approximately 38.3% and 40.6%, respectively. The energy transfer from β-Car to Chl a in CP43 and CP47 was discussed. The rates of energy transfer from β-Car to Chl a were measured to be about 9.6×1011 s-1 and 1.3×1012 s-1 and energy transfer efficiencies 47.5% and 66.5% respectively. The edge-edge distances between β-Car and Chl a in CP43 and CP47 were estimated to be ～0.110 nm and ～0.085nm respectively.     

Effect of cortisone on the developmental pattern of the neutral and the acid β-galactosidases of the small intestine of the rat

1. The developmental pattern and effect of cortisone on acid beta-galactosidase and neutral beta-galactosidase were studied in postnatal rats by a recently proposed method for their independent determination. 2. After birth the acid beta-galactosidase activity increases in the ileum, whereas it decreases slightly in the jejunum. On day 16 after birth the activity in the ileum decreases and in 20-day-old rats activity in both parts of the intestine decreases to adult values. In suckling animals the activity in the ileum exceeds the jejunal activity severalfold and in adult animals the activity in the jejunum is slightly higher than that in the ileum. 3. Neutral beta-galactosidase activity is high after birth and decreases in both jejunum and ileum after day 20 after birth. In 12-20-day-old rats activity in both parts is essentially the same, but in adult animals jejunal activity exceeds ileal activity four-to five-fold. 4. Cortisone (0.5, 2.0 or 5.0mg/100g body wt. daily for 4 days) does not influence the activity of either enzyme in 60-day-old rats. Acid beta-galactosidase activity is decreased after cortisone treatment in 8-, 12-, 16-and 18-day-old rats, with sensitivity to cortisone increasing with the approach of weaning. No effect of cortisone on acid beta-galactosidase is seen in 8-day-old rats. Neutral beta-galactosidase activity is increased in the ileum of 8-, 12-, 16- and 18-day old rats, but only in the jejunum of 8-and 12-day-old rats.     

Association of the subunits of the calcium-independent receptor of α-latrotoxin

CIRL-1 also called latrophilin 1 or CL belongs to the family of adhesion G protein-coupled receptors (GPCRs). As all members of adhesion GPSR family CIRL-1 consists of two heterologous subunits, extracellular hydrophilic p120 and heptahelical membrane protein p85. Both CIRL-1 subunits are encoded by one gene but as a result of intracellular proteolysis of precursor, mature receptor has two-subunit structure. It was also shown that a minor portion of the CIRL-1 receptor complexes dissociates, producing the soluble receptor ectodomain, and this dissociation is due to the second cleavage at the site between the site of primary proteolysis and the first transmembrane domain. Recently model of independent localization p120 and p85 on the cell surface was proposed. In this article we evaluated the amount of p120-p85 complex still presented on the cellular membrane and confirmed that on cell surface major amount of mature CIRL-1 presented as a p120-p85 subunit complex.     

A 13C CP/MAS NMR spectroscopy and AFM study of the structure of Glucagel™, a gelling β-glucan from barley

The structure of Glucagel™, a mixed-linked (1→3), (1→4)-β-d-glucan extracted from barley, was examined using ¹³C CP/MAS NMR spectroscopy and atomic force microscopy (AFM). Results from ¹³C CP/MAS NMR spectroscopy showed that Glucagel™ contained regions with two distinct conformations. In some of the regions the β-glucan chains associated to form a unique conformation, the A-conformation, while in the other regions the β-glucan chains were in an amorphous conformation. Dilute solutions of Glucagel™ were prepared for imaging by dissolving Glucagel™ in water at 90 °C. If the dilute solution was immediately deposited onto mica and the surface dried, then no fine detail was seen in the AFM image. However, when dilute solutions of Glucagel™ were left for several days before being deposited onto the mica surface, individual fibres could be clearly imaged. These results suggested that in gels formed from Glucagel™, junction zones occur because of the interaction of two β-glucan chains in the A-conformation.     

Phylogeography of the harlequin beetle-riding pseudoscorpion and the rise of the Isthmus of Panamá

Molecular and geological evidence indicates that the emergence of the Isthmus of Panamá influenced the historical biogeography of the Neotropics in a complex, staggered manner dating back at least 9 Myr bp. To assess the influence of Isthmus formation on the biogeography of the harlequin beetle-riding pseudoscorpion, Cordylochernes scorpioides, we analysed mitochondrial COI sequence data from 71 individuals from 13 locations in Panamá and northern South America. Parsimony and likelihood-based phylogenies identified deep divergence between South American and Panamanian clades. In contrast to low haplotype diversity in South America, the Panamanian Cordylochernes clade is comprised of three highly divergent lineages: one clade consisting predominantly of individuals from central Panamá (PAN A), and two sister clades (PAN B1 and PAN B2) of western Panamanian pseudoscorpions. Breeding experiments demonstrated a strictly maternal mode of inheritance, indicating that our analyses were not confounded by nuclear-mitochondrial pseudogenes. Haplotype diversity is striking in western Atlantic Panamá, where all three Panamanian clades can occur in a single host tree. This sympatry points to the existence of a cryptic species hybrid zone in western Panamá, a conclusion supported by interclade crosses and coalescence-based migration rates. Molecular clock estimates yield a divergence time of approximately 3 Myr between the central and western Panamanian clades. Taken together, these results are consistent with a recent model in which a transitory proto-Isthmus enabled an early wave of colonization out of South America at the close of the Miocene, followed by sea level rise, inundation of the terrestrial corridor and then a second wave of colonization that occurred when the Isthmus was completed approximately 3 Myr bp.     

Did the evolution of the phytoplankton fuel the diversification of the marine biosphere?

We discuss the possible links between the fossil record of marine biodiversity, nutrient availability and primary productivity. The parallelism of the fossil records of marine phytoplankton and faunal biodiversity implicates the quantity (primary productivity) and quality (stoichiometry) of phytoplankton as being critical to the diversification of the marine biosphere through the Phanerozoic. The relatively subdued marine biodiversity of the Palaeozoic corresponds to a time of relatively low macronutrient availability and poor food quality of the phytoplankton as opposed to the diversification of the Modern Fauna through the Mesozoic–Cenozoic. Increasing nutrient runoff to the oceans through the Phanerozoic resulted from orogeny, the emplacement of Large Igneous Provinces (LIPs), the evolution of deep-rooting forests and the appearance of more easily decomposable terrestrial organic matter that enhanced weathering. Positive feedback by bioturbation of an expanding benthos played a critical role in evolving biogeochemical cycles by linking the oxidation of dead organic matter and the recycling of nutrients back to the water column where they could be re-utilized. We assess our conclusions against a recently published biogeochemical model for geological time-scales. Major peaks of marine diversity often occur near rising or peak fluxes of silica, phosphorus and dissolved reactive oceanic phosphorus; either major or minor ⁸⁷Sr/⁸⁶Sr peaks; and frequently in the vicinity of major (Circum-Atlantic Magmatic Province) and minor volcanic events, some of which are associated with Oceanic Anoxic Events. These processes appear to be scale-dependent in that they lie on a continuum between biodiversification on macroevolutionary scales of geological time and mass extinction.     

Imperfect Batesian mimicry—the effects of the frequency and the distastefulness of the model

Batesian mimicry is the resemblance between unpalatable models and palatable mimics. The widely accepted idea is that the frequency and the unprofitability of the model are crucial for the introduction of a Batesian mimic into the prey population. However, experimental evidence is limited and furthermore, previous studies have considered mainly perfect mimicry (automimicry). We investigated imperfect Batesian mimicry by varying the frequency of an aposematic model at two levels of distastefulness. The predator encountered prey in a random order, one prey item at a time. The prey were thus presented realistically in a sequential way. Great tits (Parus major) were used as predators. This experiment, with a novel signal, supports the idea that Batesian mimics gain most when the models outnumber them. The mortalities of the mimics as well as the models were significantly dependent on the frequency of the model. Both prey types survived better the fewer mimics there were confusing the predator. There were also indications that the degree of distastefulness of the model had an effect on the survival of the Batesian mimic: the models survived significantly better the more distasteful they were. The experiment supports the most classical predictions in the theories of the origin and maintenance of Batesian mimicry.     

Microspectrophotometry of the photoreceptors of palaeognathous birds — the emu and the tinamou

Summary With the aid of a microspectrophotometer the visual pigments and oil globules in the retina of the emu (Dromiceius novae-hollandiae), the brushland tinamou (Nothoprocta c. cinerascens) and the Chilean tinamou (Nothoprocta perdicaria sanborni) were characterized. All three of these palaeognathous birds contain in their rods a typical rhodopsin with _max near 500 nm. Each of these birds has cones containing iodopsin-like visual pigments with _max in the 560–570 nm spectral region. No unequivocal evidence was obtained for the presence of cone pigments other than this iodopsin-like pigment, although one cell thought to be a cone, and containing a visual pigment with _max near 498 nm, was observed in the retina of the brushland tinamou. The oil globule systems of the three palaeognathous species are identical to each other and are much simpler than is typical for neognathous birds in that only two different types of globule are present, one with _T50 at 508 nm and another with _T50 at 568 nm. Comparison of the data with observations made on neognathous species indicates (1) that palaeognathous birds probably have poorer color discrimination capabilities than neognathous birds and (2) that the tinamou is more closely related to the ratites than to the galliform species.This study was supported, in part, by NIH Grant No. EY01839 (A.J. Sillman), NIH Grant No. EY00323 (W.N. McFarland) and NSF Grant No. 78-07657 (E.R. Loew). The authors thank E. Clinite, R. Dunford, C. Murphy, R. Riis and D. Weathers for their valuable assistance. Thanks also go to R.E. Burger for his gift of the emus.