Floristic turnover in Iceland from 15 to 6 Ma – extracting biogeographical signals from fossil floral assemblages

Aim This study aims to document the floristic changes that occurred in Iceland between 15 and 6 Ma and to establish the dispersal mechanisms for the plant taxa encountered. Using changing patterns of dispersal, two factors controlling floristic changes are tested. Possible factors are (1) climate change, and (2) the changing biogeography of Iceland over the time interval studied; that is, the presence or absence of a Miocene North Atlantic Land Bridge. Location The North Atlantic. Methods Species lists of fossil plants from Iceland in the time period 15 to 6 Ma were compiled using published data and new data. Closest living analogues were used to establish dispersal properties for the fossil taxa. Dispersal mechanisms of fossil plants were then used to reconstruct how Iceland was colonized during various periods. Results Miocene floras of Iceland (15–6 Ma) show relatively high floristic turnover from the oldest floras towards the youngest; and few taxa from the oldest floras persist in the younger floras. The frequencies of the various dispersal mechanisms seen in the 15‐Ma floras are quite different from those recorded in the 6‐Ma floras, and there is a gradual change in the prevailing mode of dispersal from short‐distance anemochory and dyschory to long‐distance anemochory. Two mechanisms can be used to explain changing floral composition: (1) climate change, and (2) the interaction between the dispersal mechanisms of plants and the increasing isolation of proto‐Iceland during the Miocene. Main conclusions Dispersal mechanisms can be used to extract palaeogeographic signals from fossil floras. The composition of floras and dispersal mechanisms indicate that Iceland was connected both to Greenland and to Europe in the early Middle Miocene, allowing transcontinental migration. The change in prevalence of dispersal modes from 15 to 6 Ma appears to reflect the break‐up of a land bridge and the increasing isolation of Iceland after 12 Ma. Concurrent gradual cooling and isolation caused changes in species composition. Specifically, the widening of the North Atlantic Ocean prevented taxa with limited dispersal capability from colonizing Iceland, while climate cooling led to the extinction of thermophilous taxa.     

CHROMOSOME NUMBERS OF CAMPANULACEAE. II. THE LOBELIA TUPA COMPLEX OF CHILE

Gametic chromosome numbers are reported for 27 collections representing the four species of the Lobelia tupa complex (Campanulaceae, Lobelioideae) in Chile; all are n = 21. This represents the first report of chromosome numbers for L. bridgesii Hook. & Arn., L. excelsa Bonpl., and L. polyphylla Hook. & Arn., and confirms previous reports of this number in L. tupa L. As the basic chromosome number of Lobelioideae is x = 7, these species are interpreted as hexaploids. Higher polyploids are extremely rare among Lobelioideae; most of those previously reported have been either sporadic individuals or populations within an otherwise diploid or tetraploid species, or occasional species within an otherwise diploid and tetraploid lineage. This is the first report of an entire complex of lobelioid species that is uniformly hexaploid. This suggests that the Chilean endemics are relatively derived within Lobelia, and offers some support for the monophyly of the complex.     

Electrostatic effects on modification of charged groups in the active site cleft of subtilisin by protein engineering

The dielectric constant in the active site cleft of subtilisin from Bacillus amyloliquefaciens has been probed by mutating charged residues on the rim and measuring the effect on the pKa value of the active site histidine (His64) by kinetics. Mutation of a negatively charged surface residue, which is 12 to 13 A from His64, to an uncharged one Asp----Ser99) lowers the pKa of the histidine by up to 0.4 unit at low ionic strength (0.005 to 0.01 M). This corresponds to an apparent dielectric constant of about 40 to 50 between Asp99 and His64. The mutation is in an external loop that is known to tolerate a serine at position 99 from homologies with subtilisins from other bacilli. The environment between His64 and Asp99 is predominantly protein. Another charged residue that is at a similar distance from His64 (14 to 15 A) and is also in an external loop that is known to tolerate a serine residue is Glu156, at the opposite side of the active site. There is only water in a direct line between His64 and Glu156. Mutation of Glu----Ser156 also lowers the pKa of His64 by up to 0.4 unit at low ionic strength. This change again corresponds to an apparent dielectric constant of about 40 to 50. The pKa values were determined from the pH dependence of kcat/KM for the hydrolysis of peptide substrates, with a precision of typically +/- 0.02 unit. The following suggests that the changes in pKa are real and not artefacts of experimental conditions: Hill plots of the data for pKa determination have gradients (h) of -1.00(+/- 0.02), showing that there are negligible systematic deviations from theoretical ionization curves involving a monobasic acid: the pH dependence for the hydrolysis of two different substrates (succinyl-L-alanyl-L-alanyl-L-prolyl-L-phenylalanyl p-nitroanilide and benzoyl-L-valyl-L-glycyl-L-arginyl p-nitroanilide) gives identical results so that the pKa is independent of substrate; the pH dependence is unaffected by changing the concentration of enzyme, so that aggregation is not affecting the results; the shift in pKa is masked by high ionic strength, as expected qualitatively for ionic shielding of electrostatic interactions.     

Sequence dependent conformations of oligomeric DNA's in aqueous solutions and in crystals

In order to determine the sequence dependence of the conformation of deoxynucleotides, Raman spectra have been obtained for the following oligodeoxynucleotides in aqueous salt solutions and in crystals: d(CpG)(I), d(TGCGCGCA)(II), d(CACGCGTG)(III), d(CGTGCACG)(IV), d(CGCATGCG)(V), d(ACGCGCGT)(VI), d(CGCGTACGCG)(VII), d(CGCACGTGCG)(VIII) and d(CGTGCGCACG)(IX), d(GCTATAGC) (X), d(GCATATGC) (XI), d(GGTATACC) (XII) and d(GGATATCC) (XIII). The normal B type conformation is observed for all the oligomer DNA's at low salt (0.1-1.0 M NaCl) concentration in the temperature range of 0-25 degrees C. It was considered possible that all of the first nine oligomers could go into the Z form in aqueous high salt (5.0-6.0 M NaCl) solutions, and under these conditions the last four were considered candidates to go into the A form. The B-type conformation was found to exist in high salt solutions for (I), (IV), (V), (VI), (X), (XI) and (XIII); the Z or partial Z conformation appears in high salt solution for the oligomers, (II), (III), (VII), (VIII) and (IX); an A or partial A conformation appears in high salt solution for (XII). In the crystalline state, (IV), (VIII), (X), and (XI) stay in the B-form and all of the other oligomers adopt the complete Z-form except for (XII) which crystallizes in the A form. In both the crystal and in aqueous solutions, the identification of the conformation genus was made by means of Raman spectroscopy. In the crystal of (I), grown at pH7.0, guanosine is found to be in C3'-endo/syn conformation and cytidine in C2'-endo/anti, which may be taken as the ideal building block of the typical Z conformation. At pH4, (I) crystallizes in a conformation similar to the B genus. A study of the thermally induced B to Z transition has been carried out for (II) and (III). Based on the analysis of Raman spectra of the alternating pyrimidine-purine oligomers which might be expected to go into the Z form, the tendency for these oligonucleotides to adopt the Z form can be ranked as: d(CGCGCGCG) greater than (II) greater than (III) greater than (V) approximately (VI) greater than (IV) for octamers and (VII) greater than (VIII) greater than (IX) for the decamers. Similarly, those oligomers which might have a tendency to go into the A form could be ranked as (XII) greater than (XIII) approximately (X) greater than (XI). These data should provide help in formulating rules for predicting the sequence dependence of the B to A and B to Z transitions. Some possible rules are explored, but precautions should be taken.     

Factors controlling initial and long-term ATP regeneration catalyzed by immobilized chromatophores

Photosynthetic ATP regeneration was measured in open reactors using immobilized Chromatophores from Rhodopseudomonas capsulata. The influence of several factors on both initial and long-term ADP photophosphorylation was studied. The effect of phosphate salts and of bovine serum albumin on the organelle activity yield was studied. Photophosphorylation was initiated either with ADP or regenerated ATP and the roles of these nucleotides were compared. Different photoreactor configurations were tested for the production of a phosphorylated compound and a flat reactor selected. The presence of inorganic pyrophosphate in the reaction medium was shown to improve the synthesis of ATP 1.4 times. Using the optimal conditions described here, the total G-6P production was 50-fold higher than in batch reactors.     

Characterization and in Vivo Cloning of prlC, a Suppressor of Signal Sequence Mutations in Escherichia coli K12

The prlC gene of E. coli was originally identified as an allele, prlC1, which suppresses certain signal sequence mutations in the genes for several exported proteins. We have isolated six new alleles of prlC that also confer this phenotype. These mutations can be placed into three classes based on the degree to which they suppress the lamB signal sequence deletion, lamBs78. Genetic mapping reveals that the physical location of the mutations in prlC correlates with the strength of the suppression, suggesting that different regions of the gene can be altered to yield a suppressor phenotype. We also describe an in vivo cloning procedure using lambda placMu9H. The procedure relies on transposition and illegitimate recombination to generate a specialized transducing phage that carries prlC1. This method should be applicable to any gene for which there is a mutant phenotype.     

Characterization of specific fluorenylmethyloxycarbonyl-containing calmodulin adducts by spectroscopy and phosphodiesterase stimulation

A reagent (I, N⁴-(9-fluorenylmethyloxycarbonyl-4-amino-1-oxyl-4-succinimidyloxycarbonyl-2,2,6,6-tetramethylpiperidine)) that acylates calmodulin specifically at lysines 75 and 148 was recently described (Jackson and Puett, 1984). Chromatographic procedures are described that permit purification to apparent homogeneity of a 1 : 1 and a 2 : 1 adduct characterized by modification at just Lys 75 or at Lys 75 and Lys 148, respectively. These adducts are suitable for detailed characterization in an effort to provide information on calmodulin structure-function relationships. The adducts were incapable of, or exhibited low potency (e.g., 0.1% that of calmodulin) in, stimulating the activity of an activatable bovine brain cyclic nucleotide phosphodiesterase (3,5-cyclic AMP 5-nucleotidehydrolase, EC 3.1.4.17) preparation. Electron paramagnetic resonance (EPR) spectroscopy of the adducts yielded rotational correlation times of approximately 3–6 nsec, in agreement with the expected value for a hydrated protein of this molecular weight (5–7 nsec). Thus, the nitroxide reporter group appears to monitor closely the motion of the protein, and there is no evidence of a major conformational change in the derivative relative to calmodulin. Interestingly, removal of the fluorenylmethyloxycarbonyl portion from the 1 : 1 adduct to give a deprotected 1 : 1 adduct resulted in apparent greater mobility of the probe, since the rotational correlation coefficient was found to be 1 nsec. Circular dichroic spectra were obtained over the wavelength interval 200–250 nm on the two adducts and on the deprotected 1 : 1 adduct. These derivatives, like calmodulin, exhibited a Ca²⁺-mediated increase in helicity, and the spectra of the adducts in the presence of a chelating agent and in the presence of saturating Ca²⁺ were similar to those obtained for calmodulin. Thus, the adducts have secondary structures similar to the native protein. Proton nuclear magnetic resonance spectra were determined in the aromatic region (6–8 ppm) for the deprotected 1 : 1 adduct before and after reduction of the nitroxide with ascorbate. The nitroxide had little effect on the chemical shifts of the two tyrosines and the single histidine relative to calmodulin, although the histidine C4 resonance was markedly altered by the addition of ascorbate. In order to explore in greater detail the tertiary structure of the 1 : 1 adduct, a reagent similar to I, but not paramagnetic, was synthesized. This compound II, -N-(9-fluorenylmethyloxycarbonyl)alanine N-hydroxysuccinimide ester, like I, forms a 1 : 1 adduct at Lys 75 and a 2 : 1 adduct at Lys 75 and Lys 148. Proton NMR spectra of adducts with II were not complicated by the relaxation effects arising from adducts with I; thus more definitive assignments could be made to the upfield resonances, including the fluorene protons. Again, it was possible to conclude that adduct formation had no major effect on the tertiary structure of the protein as monitored by chemical shifts associated with various residues. We conclude that modification of just Lys 75, a residue in the long connecting helix of calmodulin, does not lead to major changes in protein conformation but does interfere with the ability of calmodulin to stimulate an activatable form of bovine brain cyclic nucleotide phosphodiesterase.     

Opportunistic zygomycotic infections

This is a literature review of 361 opportunistic fungal infections caused by the Zygomycetes. The clinical and laboratory diagnosis, pathogenesis, management, treatment, and outcome of infection are discussed. The Zygomycetes are a group of opportunistic fungi (orders Mucorales and Entomophthorales) which cause severe infections which may be fatal. Early clinical recognition, prompt diagnostic procedures, control of underlying disease and treatment with high doses of amphotericin B and aggressive surgery increases survival in an otherwise lethal infection.     

Arterial supply of the choriocapillaris of anuran amphibians (Rana temporaria,Rana esculenta)

Summary The pattern of the vascular supply to the choroid of the frog eye was studied in toto with the use of the injection-replication-SEM technique. The choroid of anuran amphibians is composed mainly of the choriocapillaris. In both species studied (Rana temporaria, Rana esculenta), an independent arterial supply to the choriocapillaris supplemented that from the ciliary arteries. This additional vascular route arises from the optic artery, a separate branch of the arteria infundibularis superficialis. The optic artery, accompanied by its vein within the vascular sheath of the optic nerve, joins the rich arterial capillary network of the choriocapillaris and supplies the posterior pole of the ocular bulb. The superficial capillary network displays a dense collar around the entrance of the optic nerve into the eye and is composed of a circular meshwork of small capillaries, several layers deep. More peripherally, however, it becomes single layered. This capillary network, as a whole, establishes numerous connections with the adjacent choriocapillaris at the posterior pole of the ocular bulb. In anuran amphibians the complex arrangement of both arterial systems supporting the choriocapillaris may be regarded as a more complete equivalent of the short posterior ciliary arteries of mammals.