394delTT: a Nordic cystic fibrosis mutation

In a systematic screening for mutations in the gene encoding the cystic fibrosis transmembrane regulator among Danish cystic fibrosis (CF) patients, we identified a mutation in exon 3 (394delTT); this mutation was found to be relatively common in Denmark. We therefore screened for 394delTT in Sweden and Norway, where it turned out to be the second most frequent mutation, accounting for 4% of all CF mutations. It also occurs with a high frequency in Finland, but has not been found in larger surveys of mutations in the CFTR gene. Thus, 394delTT seems to be a specific Nordic CF mutation.     

Comparison of phylogenetic relationships based on phospholipid fatty acid profiles and ribosomal RNA sequence similarities among dissimilatory sulfate-reducing bacteria

Abstract Twenty-five isolates of dissimilatory sulfate-reducing bacteria were clustered based on similarity analysis of their phospholipid ester-linked fatty acids (PLFA). Of these, 22 showed that phylogenetic relationships based on the sequence similarity of their 16S rRNA directly paralleled the PLFA relationships. Desulfobacter latus and Desulfobacter curvatus grouped with the other Desulfobacter spp. by 16S rRNA comparison but not with the PLFA analysis as they contained significantly more monoenoic PLFA than the others. Similarly, Desulfovibrio africanus clustered with the Desulfovibrio spp. by 16S rRNA but not with them when analyzed by PLFA patterns because of higher monoenoic PLFA content. Otherwise, clustering obtained with either analysis was essentially congruent. The relationships defined by PLFA patterns appeared robust to shifts in nutrients and terminal electron acceptors. Additional analyses utilizing the lipopolysaccharide-lipid A hydroxy fatty acid patterns appeared not to shift the relationships based on PLFA significantly except when completely absent, as in Gram-positive bacteria. Phylogenetic relationships between isolates defined by 16S rRNA sequence divergence represent a selection clearly different from the multi-enzyme activities responsible for the PLFA patterns. Determination of bacterial relationships based on different selective pressures for various cellular components provides more clues to evolutionary history leading to a more rational nomenclature.     

Analysis of a primer-independent GTF-I from Streptococcus salivarius

Abstract A glucosyltransferase (GTF) gene, designated gtfL , from Streptococcus salivarius was cloned and expressed in Escherichia coli and its nucleotide sequence determined. The GTF-L enzyme catalysed the synthesis of water-insoluble glucan in a primer-independent manner. The nucleotide sequence and derived amino acid sequence of GTF-L were similar in size and domain structure to previously sequenced glucosyltransferases. However, a 464-bp region of high variability was identified which could be selectively amplified from strains of S. salivarius by the polymerase chain reaction and could therefore form the basis for species identification. No sequence-specific motifs related to the solubility and linkage of the glucan product or its need for a dextran primer could be ascertained.     

Electron Flow to the Intersystem Chain from Stromal Components and Cyclic Electron Flow in Maize Chloroplasts, as Detected in Intact Leaves by Monitoring Redox Change of P700 and Chlorophyll Fluorescence

The functional pool size of electrons in the intersystem chainof the chloroplasts of maize was estimated to be about 25 perP700 by the redox change in P700 with single- and multiple-turnoverlights under far-red light in intact leaves. This is about twicethe pool size observed in C₃ plants. Furthermore, the stromalpool size of electrons that can be donated to P700⁺ after actinicillumination was larger in maize leaves than in leaves of C₃plants, giving a maximum value of 225 electrons per P700. Maizeleaves showed an increase in the yield of modulated Chl fluorescenceafter turning off of actinic light, which confirms the donationof electrons in the dark to the intersystem chain from the stromaldonors that accumulated during actinic illumination. We proposethat the mesophyll chloroplasts are responsible for a high levelof electron-donating activity to the intersystem chain fromstromal donors such as triose phosphates and malate with NADPHas an intermediate. The level of P700⁺ under strong far-redlight was decreased after actinic illumination, suggesting theoperation of an actinic light-triggered cyclic electron flowin chloroplasts of the bundle sheath cells. (Received August 14, 1992; Accepted October 13, 1992)     

The Casparian Strip of Clivia miniata Reg. Roots: Isolation,Fine Structure and Chemical Nature*

The root endodermis of Clivia miniata Reg. was successfully isolated using the cell wall degrading enzymes cellulase and pectinase. The enzymes did not depolymerize those regions of the primary cell walls of anticlinal endodermal root cells where the Casparian strips were located. Since the endodermis of C. miniata roots remained in its primary developmental state over the whole root length, endodermal isolates essentially represented Casparian strips. Thus, sufficient amounts of isolated Casparian strips could be obtained to allow further detailed investigations of the isolates by microscopic, histochemical and analytical methods. Scanning electron microscopy revealed the reticular structure of the Casparian strips completely surrounding the central cylinder of the roots. Whereas in younger parts of the root only the anticlinal cell walls of the endodermis remained intact in the isolates, in older parts of the root the periclinal walls also restricted enzymatic degradation due to the deposition of lignin. Extracts of the isolates with organic solvents did not reveal any wax-like substances which might have been deposited within the cell wall forming a transport barrier, as is the case with cutin and suberin. However, several histochemical and analytical methods (elemental analysis and FTIR spectroscopy) showed that the chemical nature of the Casparian strips of C. miniata roots can definitely be a lignified cell wall. These findings are in complete agreement with studies carried out at the beginning of this century on the chemical nature of the Casparian strips of several other plant species. The implications of these results concerning apoplasmatic transport of solutes and water across Casparian strips are discussed.     

Satellite DNA sequences in the New World primateCebus apella (Platyrrhini,Primates)

Two satellite DNAs, designated CapA and CapB, were isolated from the neotropical primate,Cebus apella. The satellites exhibit nonoverlapping distributions onC. apella chromosomes. CapA is a major component of interstitial regions of constitutive heterochromatin, a very large block of heterochromatin comprising most of the long arm of chromosome 11, and some telomeres. The CapA monomer has a length of about 1500 bp and appears recently to have undergone an amplification episode in theC. apella genome. CapA-like sequences are probably present in members of the family Cebidae (to whichC. apella belongs), but not in members of the family Callitrichidae (marmosets). CapB sequences can be detected at the centromeres of manyC. apella chromosomes, and similar sequences are present in all neotropical primates. The 342 bp CapB monomer shares 60%–64% sequence identity with several alpha satellite sequences of human origin. Because of its structure, sequence, and location, it appears that CapB is the New World primate homolog of Old World primate alpha satellite DNA.     

Modulation of SPARC Expression during Butyrate-Induced Terminal Differentiation of Cultured Human Keratinocytes: Regulation via a TGF-β-Dependent Pathway

Expression of SPARC (secreted protein acidic and rich in cysteine), a 43-kDa extracellular matrix-associated glycoprotein involved in tissue remodeling, was quantitated during normal human keratinocyte (NHK) growth in culture and as a function of sodium n-butyrate (NaB)-induced differentiation to mature enucleate cornified envelopes (CEs). Low levels of SPARC expression were observed in the basal-like cells of control NHKs, with isolated cells showing intense SPARC expression on the ventral surface. After addition of NaB, SPARC expression increased and the pattern of expression shifted to one involving predominantly suprabasal cells (i.e., spinous cells, pre-CEs, and mature CEs). Dense deposits of SPARC often surrounded the mature CEs. Flow cytometric analysis indicated that approximately 13% of NHKs expressed SPARC within 24 h of seeding into culture. This fraction of SPARC⁺ cells increased with time and peaked immediately postconfluence (31.3 ± 6.3% SPARC⁺). Cellular SPARC expression then decreased to baseline levels during entrance into plateau phase growth. SPARC was detectable in all phases of the cell cycle. SPARC levels were more intense and heterogeneous within the G₂/M and G₁ phases while S phase cells exhibited relatively homogeneous, low intensity, SPARC expression. During NaB-induced NHK differentiation, SPARC intracellular content increased prior to the onset of CE formation (i.e., 2 days after its addition) followed by a period of extracellular accumulation which coincided with the time of maximal CE generation (i.e., Days 4 and 5 after NaB addition). Correlation of cell size with anti-SPARC immunoreactivity revealed a predominance of SPARC expression in cells with a suprabasal phenotype. NHKs cultured on fibronectin (FN), an established modulator of epidermal cell maturation in vitro, showed a similar response to NaB. In general, however, the level of NaB-induced SPARC expression was considerably reduced in FN cultures correlating with a lower efficiency of CE formation. Induced SPARC expression was, in large part, dependent on autocrine transforming growth factor-β (TGF-β) production since incubation in the presence of NaB + neutralizing antibodies to TGF-β inhibited both the expression of SPARC by 72% and development of mature CEs.     

The Tol-Pal system of Acinetobacter baumannii is important for cell morphology,antibiotic resistance and virulence

International Microbiology - Acinetobacter baumannii is an opportunistic human pathogen that has become a global threat to healthcare institutions. This Gram-negative bacterium is one of the most...     

Female reproductive fluid concentrations affect sperm performance of alternative male phenotypes in an external fertilizer

There is growing evidence that the female reproductive fluid (FRF) plays an important role in cryptic female choice through its differential effect on the performance of sperm from different males. In a natural spawning event, the male(s) may release ejaculate closer or further away from the spawning female. If the relative spatial proximity of competing males reflects the female pre-mating preference towards those males, then favoured males will encounter higher concentrations of FRF than unpreferred males. Despite this being a common situation in many external fertilizers, whether different concentrations of FRF can differentially influence the sperm performance of distinct male phenotypes (favoured and unfavoured by the female) remains to be elucidated. Here, we tested this hypothesis using the grass goby (Zosterisessor ophiocephalus), a fish with distinct territorial-sneaker reproductive tactics and female pre-mating preference towards territorial males, that consequently mate in an advantaged position and whose sperm experience higher concentrations of FRF. Our findings revealed a differential concentration-dependent effect of FRF over sneaker and territorial sperm motility only at low concentrations (i.e. at the distance where sneakers typically ejaculate), with increasing FRF concentrations (i.e. close to the eggs) similarly boosting the sperm performance of both sneaker and territorial males. The ability to release sperm close to the eggs is a prerogative of territorials, but FRF can likewise advantage the sperm of those sneakers that are able to get closer, allowing flexibility in the direction of female post-mating choice.     

Identification of Products from Oxidation of Uric Acid Induced by Hydroxyl Radicals

The aim of the present study was to separate and characterise products formed by oxidation of uric acid by hydroxyl radicals with a view to probing for these products in vivo in clinical contexts. Aerated solutions of 200 μM uric acid, or its oxidation products, allantoin or parabanic acid, were exposed to gamma radiolysis, (52.0 Gy/min), as a source of HO- radicals, at pH 3.4 and 7.4. Aliquots were taken every 5 minutes for 20 minutes and oxidation products were separated by HPLC and analysed with a diode array detector. Identities of oxidation products were confirmed on the basis of similarity of retention times and absorbance spectra and peak purity parameters of known standards. Hydroperoxides were measured by tri-iodide formation in the 20 minute sample. Exposure of uric acid to such HO fluxes produced a net loss of the parent compound with formation of a complex mixture of products with allantoin and parabanic acid being the predominant products at pH 3.4. The rate of uric acid degradation at physiological pH was slower and the distribution of oxidation products was different. A small but significant amount of uric acid hydroperoxide was detected at both pHs. A mechanism for uric acid oxidation under these conditions is presented.