Molecular drift of the bride of sevenless (boss) gene in Drosophila

DNA sequences were determined for three to five alleles of the bride-of- sevenless (boss) gene in each of four species of Drosophila. The product of boss is a transmembrane receptor for a ligand coded by the sevenless gene that triggers differentiation of the R7 photoreceptor cell in the compound eye. Population parameters affecting the rate and pattern of molecular evolution of boss were estimated from the multinomial configurations of nucleotide polymorphisms of synonymous codons. The time of divergence between D. melanogaster and D. simulans was estimated as approximately 1 Myr, that between D. teissieri and D. yakuba as approximately 0.75 Myr, and that between the two pairs of sibling species as approximately 2 Myr. (The boss genes themselves have estimated divergence times approximately 50% greater than the species divergence times.) The effective size of the species was estimated as approximately 5 x 10(6), and the average mutation rate was estimated as 1-2 x 10(-9)/nucleotide/generation. The ratio of amino acid polymorphisms within species to fixed differences between species suggests that approximately 25% of all possible single-step amino acid replacements in the boss gene product may be selectively neutral or nearly neutral. The data also imply that random genetic drift has been responsible for virtually all of the observed differences in the portion of the boss gene analyzed among the four species.      

Are short normal children at a disadvantage? The Wessex growth study.

OBJECTIVE: To examine whether short stature through childhood represents a disadvantage at around 12 years. DESIGN: Longitudinal non-intervention study of the physical and psychological development of children recruited from the community in 1986-7 after entry into primary school at age 5-6 years; this is the second psychometric assessment made in 1994-5 after entry into secondary school at age 11-13 years. SETTING: Southampton and Winchester health districts. SUBJECTS: 106 short normal children (< 3rd centile for height when recruited) and 119 controls of average stature (10th-90th centile). MAIN OUTCOME MEASURES: Psychometric measures of cognitive development, self concept development, behaviour, and locus of control. RESULTS: The short children did not differ significantly from the control children on measures of self esteem (19.4 v 20.2), self perception (104.2 v 102.4), parents'' perception (46.9 v 47.0), or behaviour (6.8 v 5.3). The short children achieved significantly lower scores on measures of intelligence quotient (IQ) (102.6 v 108.6; P < 0.005), reading attainment (44.3 v 47.9; P < 0.002), and basic number skills (40.2 v 43.5; P < 0.003) and displayed less internalisation of control (16.6 v 14.3; P < 0.001) and less satisfaction with their height (P < 0.0001). More short than control children, however, came from working class homes (P < 0.05). Social class was a better predictor than height of all measures except that of body satisfaction. Attainment scores were predicted by class and IQ together rather than by height. Height accounted for some of the variance in IQ and locus of control scores. CONCLUSIONS: These results provide only limited support for the hypothesis that short children are disadvantaged, at least up until 11-13 years old. Social class seems to have more influence than height on children''s psychological development.     

Identification and DNA sequence of fixZ, a nifB-like gene from Rhizobium leguminosarum.

Previously, several mutants which nodulated peas but which failed to fix nitrogen were isolated following Tn5 mutagenesis of pRL 1JI, a symbiotic plasmid of Rhizobium leguminosarum. Two of these alleles, fix52::Tn5 and fix137::Tn5 were in a region of pRL 1JI which hybridized to a probe that contained the nifA gene and the amino-terminal region of the nifB gene of Klebsiella pneumoniae. The nitrogen fixation defect of the fix52::Tn5 mutant strain was corrected by a 2.0kb fragment of the corresponding wild-type DNA cloned in a wide host-range plasmid. The DNA sequence of this region revealed an open reading frame corresponding to the gene within which the fix52::Tn5 allele was located. The polypeptide corresponding to this open reading frame had a deduced molecular weight of 39,936 and the gene was termed fixZ. The deduced amino acid sequence of the fixZ gene product contained two clusters of cysteine residues, suggesting that the protein may contain an iron-sulphur cluster. The sequence of the fixZ polypeptide was very similar to the sequence of the K. pneumoniae nifB gene (provided by W. Arnold and A. Pühler) which is required for the synthesis of the FeMo-cofactor of nitrogenase. It was shown that the previously observed hybridization was due to homology between the amino terminal regions of fixZ and nifB. Upstream from fixZ was found another open reading frame whose 5' terminus was not established, but within which was located the fix137::Tn5 allele. This gene was termed fixY. The deduced amino acid sequence of the sequenced part of fixY showed similarity to that of the regulatory nifA gene of K. pneumoniae (provided by W. J. Buikema and F. M. Ausubel). Thus in R. leguminoarum the fix genes that correspond to the nifA and nifB genes are in the same relative orientation as in K. pneumoniae.     

The contractile basis of amoeboid movement: V. The control of gelation, solation, and contraction in extracts from dictyostelium discoideum

Motile extracts have been prepared from Dictyostelium discoideum by homogenization and differential centrifugation at 4 degrees C in a stabilization solution (60). These extracts gelled on warming to 25 degrees Celsius and contracted in response to micromolar Ca++ or a pH in excess of 7.0. Optimal gelation occurred in a solution containing 2.5 mM ethylene glycol-bis (β-aminoethyl ether)N,N,N',N'-tetraacetate (EGTA), 2.5 mM piperazine-N-N'-bis [2-ethane sulfonic acid] (PIPES), 1 mM MgC1(2), 1 mM ATP, and 20 mM KCI at ph 7.0 (relaxation solution), while micromolar levels of Ca++ inhibited gelation. Conditions that solated the gel elicited contraction of extracts containing myosin. This was true regardless of whether chemical (micromolar Ca++, pH >7.0, cytochalasin B, elevated concentrations of KCI, MgC1(2), and sucrose) or physical (pressure, mechanical stress, and cold) means were used to induce solation. Myosin was definitely required for contraction. During Ca++-or pH-elicited contraction: (a) actin, myosin, and a 95,000-dalton polypeptide were concentrated in the contracted extract; (b) the gelation activity was recovered in the material sqeezed out the contracting extract;(c) electron microscopy demonstrated that the number of free, recognizable F-actin filaments increased; (d) the actomyosin MgATPase activity was stimulated by 4- to 10-fold. In the absense of myosin the Dictyostelium extract did not contract, while gelation proceeded normally. During solation of the gel in the absense of myosin: (a) electron microscopy demonstrated that the number of free, recognizable F- actin filaments increased; (b) solation-dependent contraction of the extract and the Ca++-stimulated MgATPase activity were reconstituted by adding puried Dictyostelium myosin. Actin purified from the Dictyostelium extract did not gel (at 2 mg/ml), while low concentrations of actin (0.7-2 mg/ml) that contained several contaminating components underwent rapid Ca++ regulated gelation. These results indicated : (a) gelation in Dictyostelium extracts involves a specific Ca++-sensitive interaction between actin and several other components; (b) myosin is an absolute requirement for contraction of the extract; (c) actin-myosin interactions capable of producing force for movement are prevented in the gel, while solation of the gel by either physical or chemical means results in the release of F-actin capable of interaction with myosin and subsequent contraction. The effectiveness of physical agents in producting contraction suggests that the regulation of contraction by the gel is structural in nature.     

Different effects of inhibitors on two mutants of Escherichia coli K12 affected in the Fo portion of the adenosine triphosphatase complex.

The effects of the inhibitors dicyclohexyl-carbodiimide (DCCD), bathophenanthroline and tertiary octylcatechol, on some enzyme activities in membranes from strains of Escherichia coli carrying mutations in the uncB or uncC genes have been studied. Membranes prepared from uncC mutants retain a normal DCCD-sensitive Mg2+-stimulated adenosine triphosphatase (Mg-ATPase) activity whereas in uncB mutants this enzyme activity is insensitive to DCCD. The membrane-bound Mg-ATPase activity from the uncC mutant strain, as compared with that from the normal strain, is only partially sensitive to the inhibitors bathophenanthroline or tertiary-octylcatechol. Both of these inhibitors stimulate the membrane-bound Mg-ATPase from uncB mutant strains. A DCCD-insensitive Mg-ATPase activity is found in the cytoplasmic fraction following cell disruption of either the uncB or the uncC mutants. The lipophilic chelators bathophenanthroline and tertiary-octylcatechol stimulate the activity of the 'soluble' Mg-ATPase in the uncB mutant but partially inhibit the activity in the uncC mutant. The NADH oxidase activities in membranes from both mutant and normal strains are strongly inhibited by tertiary-octylcatechol and bathophenanthroline but not by DCCD.     

Molecular genetics of mutants of Rhizobium leguminosarum which fail to fix nitrogen

Summary Mutants of Rhizobium leguminosarum which failed to fix nitrogen within nodules on peas were isolated following the insertion of the transposon Tn5 into pRL1JI, a Rhizobium plasmid known to carry the genes for nitrogenase. The sites of the Tn5 insertions were identified by restriction endonuclease mapping of cloned fragments of DNA from the mutant strains. One group of mutants was located within 4 kilobases of the structural genes for nitrogenase and another was located about 30 kilobases from this region. Two mutants from the first group, one of which appeared to be affected in a nitrogenase gene, induced nodules that contained bacterioids, but the number of plant cells containing bacteroids was less than in a normal nodule. Another group of mutants, which was located about 30 kilobases from the nitrogenase genes failed to form bacterioids. Electron microscopy of the nodules induced by these mutants indicated that there was a defect in their release from infection threads.     

Genetic complementation between two mutant unc alleles (unc A401 and unc D409) affecting the Fl portion of the magnesium ion-stimulated adenosine triphosphatase of Escherichia coli K12.

A new mutant strain of Escherichia coli in which phosphorylation is uncoupled from electron transport was isolated. A genetic-complementation analysis, using partial diploid strains, showed that the new mutant allele, uncD409, is in a gene distinct from the other previously identified genes uncA, uncB and uncC. A strain carrying the uncd409 allele has no Mg2+ ion-stimulated adenosine triphosphatase activity and is therefore phenotypically similar to strains carrying the uncA401 mutant allele. Complementation between the uncA401 and the uncD409 alleles occurred, as indicated by growth of partial diploid strains on succinate and their growth yields on limiting concentrations of glucose. Complementation was confirmed by using membranes prepared from the above partial diploids. Such membranes were found to have Mg2+-stimulated adenosine triphosphatase activity, ATP-dependent transhydrogenase activity ADP-induced atebrin-fluorescence quenching and low but significant amounts of oxidative phosphorylation.     

Prostaglandin involvement in contractions evoked in rabbit detrusor by field stimulation and by adenosine 5'-triphosphate

Previous reports suggest that in rabbit urinary bladder both noncholinergic nonadrenergic excitatory responses and the contraction produced by adenosine 5'-triphosphate (ATP) are antagonized by indomethacin. We have attempted by further indirect testing on isolated detrusor strips to determine what role prostaglandins (PGs) might play in these processes. The second part of the biphasic contractile response to ATP was reduced to about 30% of control by PG synthesis inhibitors but the initial phase of the ATP response and te contraction produced by the beta, gamma-methylene analogue of ATP were unaffected. At concentrations that did not affect the response to acetylcholine but greatly suppressed the response to arachidonic acid, indomethacin antagonized the contraction evoked by field stimulation by about 30% at 1-2 Hz (largely noncholinergic and nonadrenergic). SC 19220, a putative PG receptor blocker, also produced about 25% reduction in the response to field stimulation but with only about 50% reduction in the response to arachidonic acid, PGE2, or PGF2alpha, SC 19220 also antagonized the frequency-response curve in atropine-treated strips. These findings lead us to suggest that beside maintaining tone and spontaneous activity in the bladder PGs mediate the slow tonic phase of the ATP response and may contribute to facilitatory modulation of noncholinergic nonadrenergic excitatory transmission.