Characterization of Trichodesmium spp. by genetic techniques

The genetic diversity of Trichodesmium spp. from natural populations (off Bermuda in the Sargasso Sea and off North Australia in the Arafura and Coral Seas) and of culture isolates from two regions (Sargasso Sea and Indian Ocean) was investigated. Three independent techniques were used, including a DNA fingerprinting method based on a highly iterated palindrome (HIP1), denaturing gradient gel electrophoresis of a hetR fragment, and sequencing of the internal transcribed spacer (ITS) of the 16S-23S rDNA region. Low genetic diversity was observed in natural populations of Trichodesmium spp. from the two hemispheres. Culture isolates of Trichodesmium thiebautii, Trichodesmium hildebrandtii, Trichodesmium tenue, and Katagnymene spiralis displayed remarkable similarity when these techniques were used, suggesting that K. spiralis is very closely related to the genus TRICHODESMIUM: The largest genetic variation was found between Trichodesmium erythraeum and all other species of Trichodesmium, including a species of KATAGNYMENE: Our data obtained with all three techniques suggest that there are two major clades of Trichodesmium spp. The HIP1 fingerprinting and ITS sequence analyses allowed the closely related species to be distinguished. This is the first report of the presence of HIP1 in marine cyanobacteria.     

INK4a-deficient human diploid fibroblasts are resistant to RAS-induced senescence

The CDKN2A tumour suppressor locus encodes two distinct proteins, p16(INK4a) and p14(ARF), both of which have been implicated in replicative senescence, the state of permanent growth arrest provoked in somatic cells by aberrant proliferative signals or by cumulative population doublings in culture. Here we describe primary fibroblasts from a member of a melanoma-prone family who is homozygous for an intragenic deletion in CDKN2A. Analyses of the resultant gene products imply that the cells are p16(INK4a) deficient but express physiologically relevant levels of a frameshift protein that retains the known functions of p14(ARF). Although they have a finite lifespan, the cells are resistant to arrest by oncogenic RAS. Indeed, ectopic expression of RAS and telomerase (hTERT) results in outgrowth of anchorage-independent colonies that have essentially diploid karyotypes and functional p53. We find that in human fibroblasts, ARF is not induced demonstrably by RAS, pointing to significant differences between the proliferative barriers implemented by the CDKN2A locus in different cell types or species.     

The transposable elements of the Drosophila melanogaster euchromatin: a genomics perspective

Background

Transposable elements are found in the genomes of nearly all eukaryotes. The recent completion of the Release 3 euchromatic genomic sequence of Drosophila melanogaster by the Berkeley Drosophila Genome Project has provided precise sequence for the repetitive elements in the Drosophila euchromatin. We have used this genomic sequence to describe the euchromatic transposable elements in the sequenced strain of this species.

Results

We identified 85 known and eight novel families of transposable element varying in copy number from one to 146. A total of 1,572 full and partial transposable elements were identified, comprising 3.86% of the sequence. More than two-thirds of the transposable elements are partial. The density of transposable elements increases an average of 4.7 times in the centromere-proximal regions of each of the major chromosome arms. We found that transposable elements are preferentially found outside genes; only 436 of 1,572 transposable elements are contained within the 61.4 Mb of sequence that is annotated as being transcribed. A large proportion of transposable elements is found nested within other elements of the same or different classes. Lastly, an analysis of structural variation from different families reveals distinct patterns of deletion for elements belonging to different classes.

Conclusions

This analysis represents an initial characterization of the transposable elements in the Release 3 euchromatic genomic sequence of D. melanogaster for which comparison to the transposable elements of other organisms can begin to be made. These data have been made available on the Berkeley Drosophila Genome Project website for future analyses.     

Dinitrogen fixation in the world's oceans

The surface water of themarine environment has traditionally beenviewed as a nitrogen (N) limited habitat, andthis has guided the development of conceptualbiogeochemical models focusing largely on thereservoir of nitrate as the critical source ofN to sustain primary productivity. However,selected groups of Bacteria, includingcyanobacteria, and Archaea canutilize dinitrogen (N₂) as an alternativeN source. In the marine environment, thesemicroorganisms can have profound effects on netcommunity production processes and can impactthe coupling of C-N-P cycles as well as the netoceanic sequestration of atmospheric carbondioxide. As one component of an integrated Nitrogen Transport and Transformations project, we have begun to re-assess ourunderstanding of (1) the biotic sources andrates of N₂ fixation in the world'soceans, (2) the major controls on rates ofoceanic N₂ fixation, (3) the significanceof this N₂ fixation for the global carboncycle and (4) the role of human activities inthe alteration of oceanic N₂ fixation. Preliminary results indicate that rates ofN₂ fixation, especially in subtropical andtropical open ocean habitats, have a major rolein the global marine N budget. Iron (Fe)bioavailability appears to be an importantcontrol and is, therefore, critical inextrapolation to global rates of N₂fixation. Anthropogenic perturbations mayalter N₂ fixation in coastal environmentsthrough habitat destruction and eutrophication,and open ocean N₂ fixation may be enhancedby warming and increased stratification of theupper water column. Global anthropogenic andclimatic changes may also affect N₂fixation rates, for example by altering dustinputs (i.e. Fe) or by expansion ofsubtropical boundaries. Some recent estimatesof global ocean N₂ fixation are in therange of 100–200 Tg N (1–2 × 10¹⁴ g N)yr^–1, but have large uncertainties. Theseestimates are nearly an order of magnitudegreater than historical, pre-1980 estimates,but approach modern estimates of oceanicdenitrification.     

Growth variations in the bivalve<Emphasis Type="Italic"> Mya truncata</Emphasis>: a tool to trace changes in the Frisian Front macrofauna (southern North Sea)?

Annual monitoring of the benthic fauna living at the Frisian Front (southern North Sea) has shown a tenfold decrease in the dominant brittlestar Amphiura filiformis in 1993–1995. In search of evidence that this decline was caused by a change in benthic food supply, we analysed variations in the shell growth of the bivalve Mya truncata from the Frisian Front during the period of interest. For this purpose, the widths of the internal growth bands in the chondrophore of M. truncata were standardised and assigned to calendar years. Averaging the yearly band width in the period 1985–2000 among 25 individuals revealed low growth rates in 1986 and 1992. Growth of M. truncata quickly recovered after 1992, while A. filiformis densities remained at low levels. Moreover, the 1986 dip in M. truncata growth had no equivalent in A. filiformis density. We conclude that there is no direct coupling between fluctuations in density of A. filiformis and variations in growth of M. truncata. The data we collected during this study on the size and spatial distribution of M. truncata are discussed in the light of plans for the protection and conservation of long-lived benthic organisms in the North Sea. Communicated by E. Rachor     

Scavenging of reactive oxygen species by a novel glucurinated flavonoid antioxidant isolated and purified from spinach

NAO is a natural water soluble antioxidant that was isolated and purified from spinach leaves. Using HPLC, NMR, and CMR spectroscopy, the main components were identified as flavonoids and p-coumaric acid derivatives. The NAO was found to be a very effective antioxidant in several in vivo and in vitro biological systems. In the present study, the antioxidant activity of the novel antioxidant glucurinated flavonoid (GF) isolated and characterized from NAO, is compared to well-known antioxidants. In addition, the direct free radical scavenging properties of the purified component GF were studied using the electron spin resonance (ESR) technique. GF and NAO were found to be superior to EGCG and NAC and to the Vitamin E homologue Trolox in inhibiting reactive oxygen species (ROS) formation in the autooxidation system of linoleic acid and in fibroblasts exposed to metal oxidation. GF and NAO were found to inhibit the ESR signal intensity of DMPO-O(2) radical formation during the riboflavin photodynamic reaction. 10 mM GF caused approximately 90% inhibition in the intensity of the ESR signal, while NAO at a concentration of 60 microg/ml caused an inhibition of about 50%. Using the Fenton reaction, GF and NAO were found to inhibit DMPO-OH radical formation. A concentration of 2 mM GF caused a 70% inhibition in the intensity of the DMPO-OH radical ESR signal, while propyl gallate at the same concentration caused only 50% inhibition. Furthermore, both GF and NAO also inhibited the (1)O(2) dependent TEMPO radical generated in the photoradiation TPPS4 system. About 80% inhibition was obtained by 4 mM GF. The results obtained indicate that the natural antioxidants derived from spinach may directly affect the scavenging of ROS and, as a consequence, may be considered as effective sources for combating oxidative damage.     

Physiological adaptations of the gut in the Lake Magadi tilapia, Alcolapia grahami, an alkaline- and saline-adapted teleost fish

We describe the gut physiology of the Lake Magadi tilapia (Alcolapia grahami), specifically those aspects associated with feeding and drinking while living in water of unusually high carbonate alkalinity (titratable base=245 mequiv l(-1)) and pH (9.85). Drinking of this highly alkaline lake water occurs at rates comparable to or higher than those seen in marine teleosts. Eating and drinking take place throughout the day, although drinking predominates during hours of darkness. The intestine directly intersects the esophagus at the anterior end of the stomach forming a 'T', and the pyloric sphincter, which comprises both smooth and striated muscle, is open when the stomach is empty and closed when the stomach is full. This unique configuration (a functional trifurcation) allows imbibed alkaline water to bypass the empty stomach, thereby avoiding a reactive mixing with acidic gastric fluids, and minimizes interference with a full stomach. No titratable base was present in the stomach, where the mean pH was 3.55, but the intestine was progressively more alkaline (foregut 6.96, midgut 7.74, hindgut 8.12, rectum 8.42); base levels in the intestinal fluid were comparable to those in lake water. The gut was highly efficient at absorbing water (76.6%), which accompanied the absorption of Na(+) (78.5%), titratable base (80.8%), and Cl(-) (71.8%). The majority of Na(+), base and water absorption occurred in the foregut by an apparent Na(+) plus base co-transport system. Overall, more than 70% of the intestinal flux occurred via Na(+) plus base co-transport, and less than 30% by Na(+) plus Cl(-) co-transport, a very different situation from the processes in the intestine of a typical marine teleost.     

Cyanobacterial diversity in geographically related and distant host plants of the genus Gunnera

The diversity among 45 cyanobacterial isolates from 11 different Gunnera species originating from different geographical areas was examined. By means of polymerase chain reaction (PCR) fingerprinting with short tandemly repeated repetitive (STRR) sequences as primers, ten groups of symbiotic cyanobacteria and five unique isolates not belonging to a particular group were identified. Most groups were restricted to one geographical area, indicating a limited distribution of related cyanobacterial strains. An extensive cyanobacterial diversity was found both within and between the 11 different Gunnera species. Within a particular plant and even within the same stem gland, more than one cyanobacterial strain at a time could be present. These results indicate a low specificity in Gunnera-Nostoc symbiosis.