The reserve proteins in the cells of mature cotyledons ofLupinus albus var.Lucky

Summary The nuclear DNA content of cotyledonary cells of two lupin seeds (L₁ and L₂) with markedly different total protein content, were investigated by scanning cytophotometry. Both seeds had polyloid nuclei with DNA levels varying between 8 C and 64 C, the majority being either 16 C or 32 C. The highest DNA levels were found in the abaxial and central cotyledonary zones of both seeds; seed L₂ had a higher ploidy level than L₁. It is shown that the volume of condensed chromatin (chromocenters) increased proportionally with the DNA content of the nucleus. A comparison was made between the distribution of protein, previously determined byLe Gal andRey (1986) and the DNA throughout the cotyledon. The L₂ seed, which has the highest total protein and the highest protein content per cell, also exhibited the greatest DNA content per cell. For both seeds, the r-value for association of DNA and protein content per cell was highly significant (0.98).     

Localization of sequences regulating ancestral and acquired sites of esterase6 activity in Drosophila melanogaster

We have broadly defined the DNA regions regulating esterase6 activity in several life stages and tissue types of D. melanogaster using P- element-mediated transformation of constructs that contain the esterase6 coding region and deletions or substitutions in 5' or 3' flanking DNA. Hemolymph is a conserved ancestral site of EST6 activity in Drosophila and the primary sequences regulating its activity lie between -171 and -25 bp relative to the translation initiation site: deletion of these sequences decrease activity approximately 20-fold. Hemolymph activity is also modulated by four other DNA regions, three of which lie 5' and one of which lies 3' of the coding region. Of these, two have positive and two have negative effects, each of approximately twofold. Esterase6 activity is present also in two male reproductive tract tissues; the ejaculatory bulb, which is another ancestral activity site, and the ejaculatory duct, which is a recently acquired site within the melanogaster species subgroup. Activities in these tissues are at least in part independently regulated: activity in the ejaculatory bulb is conferred by sequences between -273 and -172 bp (threefold decrease when deleted), while activity in the ejaculatory duct is conferred by more distal sequences between -844 and -614 bp (fourfold decrease when deleted). The reproductive tract activity is further modulated by two additional DNA regions, one in 5' DNA (-613 to -284 bp; threefold decrease when deleted) and the other in 3' DNA (+1860 to +2731 bp; threefold decrease when deleted) that probably overlaps the adjacent esteraseP gene. Collating these data with previous studies suggests that expression of EST6 in the ancestral sites is mainly regulated by conserved proximal sequences while more variable distal sequences regulate expression in the acquired ejaculatory duct site.      

Optimization of transverse alternating field electrophoresis for strain identification of Leuconostoc oenos

Genomic DNA of several strains oof oenological lactic bacteria belonging to the species Lactobacillus plantarum, Leuconostoc oenos and Pediococcus pentosaceus was digested by the rare-cutting endonucleases ApaI and SmaI. The restriction products were separated by transverse alternating field electrophoresis (TAFE). The size of the genome of L. oenos estimated by adding the molecular size of the ApaI fragments was on average 1320 kb. A strong polymorphism was observed between the strains, which could be easily differentiated except for two industrial strains of L. oenos. A simple modification of the TAFE apparatus is proposed to improve the separation of the DNA fragments. Correspondence to: J.-N. Hallet     

The IS4 family of insertion sequences: evidence for a conserved transposase motif

The eight IS 231 variants characterized so far (IS 231 A-F, V and W) display similar transposases with an overall 40% identity. Comparison with all the proka-ryotic transposable elements sequenced so far revealed that the IS231 transposases share two conserved regions with those of 35 other insertion sequences of wide origins. These insertion sequences, defining the IS4 family, have a common bipartite organization of their ends and are divided into two similarity groups. Interestingly, the transposase domains conserved within this family display similarities with the well known integrase domain shared by transposases of the IS3 and IS15 families, and integrases of retroelements. This domain is also found in IS30- related elements and Tn7 TnsB protein. Amino acid residues conserved throughout all these prokaryotic and eukaryotic mobile genetic elements define a major transposase/integrase motif, likely to play an important role in the transposition process.     

Intraspecific diversity in the fungal speciesChaunopycnis alba: Implications for microbial screening programs

Intraspecific variation among 84 isolates of the anamorphic fungusChaunopycnis alba from 26 different geographical locations was analyzed by investigating optimal growth temperatures, differences in the production of secondary metabolites and presence or absence of the cyclosporin synthetase gene. The genetic diversity was assessed using random amplified polymorphic DNA (RAPD). Analysis of these data showed high genetic, metabolic and physiological diversity within this species. Isolates from the Antarctic represented the most homogeneous group withinC. alba and together with isolates from the Arctic these polar strains differed from alpine, temperate and tropical strains by low optimal growth temperatures and by low production of secondary metabolites. Isolates from tropical climes were characterized by high optimal growth temperatures and by the production of comparatively diverse metabolite spectra. Most of the isolates that were similar in the combination of their physiological and metabolic characters were also genetically related. Isolates from different geographical origins did not show many similarities, with the exception of the cyclosporin A-producing isolates, and large diversity could be observed even within a single habitat. This leads us to the suggestion that for pharmaceutical screening programs samples should be collected from a diversity of different geographical and climatic locations. For the selection of strains for screening the RAPD assay seems to be the most powerful tool. It reflected the highest intraspecific diversity and the results corresponded well with the other characteristics.     

The highly thermostable arginine repressor of Bacillus stearothermophilus: gene cloning and repressor–operator interactions

We report here the cloning of the arginine repressor gene argR of Bacillus stearothermophilus and the characterization and purification to homogeneity of its product. The deduced amino acid sequence of the 16.8-kDa ArgR subunit shares 72% identity with its mesophilic homologue AhrC of Bacilus subtilis . Sequence analysis of B. stearothermophilus ArgR and comparisons with mesophilic arginine repressors suggest that the thermostable repressor comprises an N-terminal DNA-binding and a C-terminal oligomerization and arginine-binding region. B. stearothermophilus ArgR has been overexpressed in E. coli and purified as a 48.0-kDa trimeric protein. The repressor inhibits the expression of a B. stearothermophilus argC–lacZ fusion in E. coli cells. In the presence of arginine, the purified protein binds tightly and specifically to the argC operator, which largely overlaps the argC promoter. The purified B. stearothermophilus repressor proved to be very thermostable with a half-life of approximately 30 min at 90°C, whereas B. subtilis AhrC was largely inactivated at 65°C. Moreover, ArgR operator complexes were found to be remarkably thermostable and could be formed efficiently at up to 85°C, well above the optimal growth temperature of the moderate thermophile B. stearothermophilus . This pronounced resistance of the repressor–operator complexes to heat treatment suggests that the same type of regulatory mechanism could operate in extreme thermophiles.