Ancient Mitochondrial DNA Reveals the Origin of Sus scrofa from Rebun Island, Japan

The Kabukai A site (5 to 8C A.D.) of the Okhotsk cultural area is on Rebun Island, a small island near the coast, north–northwest of Hokkaido, Japan. Specimens of Sus scrofa, called the Sakhalin pig, were discovered in five cultural layers at the Kabukai A site. Ancient DNA was extracted from the remains of 42 Sakhalin pig bones. Thirty-nine nucleotide sequences of the 574-bp mitochondrial DNA control region, estimated to have originated from at least 21 individuals, were amplified and analyzed phylogenetically. Nine distinct haplotypes (A1, A2, A3, B1, B2, C1, C2, D1, and D2) from this site were classified into four haplotype groups (A, B, C, and D) by parsimonious network analysis. Phylogenetic analysis of 9 ancient and 55 modern haplotypes indicated that the population of Sakhalin pigs at the Kabukai A site belonged to two distinct clusters; haplotype groups A and B formed a cluster comprised only of themselves, and haplotype groups C and D belonged to the cluster of one of the two genetic groups of Japanese wild boars uniquely distributed in the western part of Japan, including one northeast Mongolian wild boar. Analysis of the haplotype distribution among three archaeological sites and their historical transitions among the five layers reflecting the cultural periods at the Kabukai A site suggests that the Sakhalin pig populations were introduced from Sakhalin island and the Amur River basin in the northeastern Eurasian continent together with some cultural influences. Received: 18 April 2000 / Accepted: 24 November 2000     

H+-ATPase defect in Corynebacterium glutamicum abolishes glutamic acid production with enhancement of glucose consumption rate

A mutant of Corynebacterim glutamicum ('Brevibacterium flayum') ATCC14067 with a reduced H+-ATPase activity, F172-8, was obtained as a spontaneous neomycin-resistant mutant. The ATPase activity of strain F172-8 was reduced to about 25% of that of the parental strain. Strain F172-8 was cultured in a glutamic-acid fermentation medium containing 100 g/l of glucose using ajar fermentor. It was found that glucose consumption per cell during the exponential phase was higher by 70% in the mutant than in the parent. The respiration rate per cell of the mutant also increased to twice as much as that of the parent. However, the growth rate of the mutant was lower than that of the parent. Under those conditions, the parent produced more than 40 g/l glutamic acid, while the mutant hardly produced any glutamic acid. Instead the mutant produced 24.6 g/l lactic acid as the main metabolite of glucose. Remarkably, the accumulation of pyruvate and pyruvate-family amino acids, i.e., alanine and valine, was detected in the mutant. On the other hand, the parent accumulated alpha-ketoglutaric acid and a glutamate-family amino acid, proline, as major by-products. It was concluded that the decrease in the H+-ATPase activity caused the above-mentioned metabolic changes in strain F172-8, because a revertant of strain F172-8, R2-1, with a H+-ATPase activity of 70% of that of strain ATCC14067, showed a fermentation profile similar to that of the parent. Sequence analyses of the atp operon genes of these strains identified one point mutation in the gamma subunit in strain F172-8.     

The DEFECTIVE IN ANTHER DEHISCENCE1 Gene Encodes a Novel Phospholipase A1 Catalyzing the Initial Step of Jasmonic Acid Biosynthesis, Which Synchronizes Pollen Maturation, Anther Dehiscence, and Flower Opening in Arabidopsis

The Arabidopsis mutant defective in anther dehiscence1 (dad1) shows defects in anther dehiscence, pollen maturation, and flower opening. The defects were rescued by the exogenous application of jasmonic acid (JA) or linolenic acid, which is consistent with the reduced accumulation of JA in the dad1 flower buds. We identified the DAD1 gene by T-DNA tagging, which is characteristic to a putative N-terminal transit peptide and a conserved motif found in lipase active sites. DAD1 protein expressed in Escherichia coli hydrolyzed phospholipids in an sn-1–specific manner, and DAD1–green fluorescent protein fusion protein expressed in leaf epidermal cells localized predominantly in chloroplasts. These results indicate that the DAD1 protein is a chloroplastic phospholipase A1 that catalyzes the initial step of JA biosynthesis. DAD1 promoter::β-glucuronidase analysis revealed that the expression of DAD1 is restricted in the stamen filaments. A model is presented in which JA synthesized in the filaments regulates the water transport in stamens and petals.     

Radioimmunoassay of nicotine.

A sensitive and specific radioimmunoassay of nicotine was developed using antisera raised against 6-(p-aminobenzamido) nicotine coupled to bovine serum albumin. Inhibition studies with various nicotine analogues revealed that the antisera are highly specific for both the N-methylpyrrolidine ring and the pyridine ring of nicotine, and especially for the structural changes of the former. The use of these antisera in an assay of nicotine in biological fluids is desirable, since the pyrrolidine ring of nicotine is first metabolized in vivo and antibodies must, therefore, discriminate nicotine from other nicotine metabolites.     

Intra- and interbreed genetic variations of mitochondrial DNA major non-coding regions in Japanese native dog breeds [Canis familiaris)

Mitochondrial DNA (mtDNA) major non-coding regions were amplified from 73 dogs of eight Japanese native dog breeds and from 21 dogs of 16 non-Japanese dog breeds by the polymerase chain reaction and their DNA sequences were determined. A total of 51 nucleotide positions within the non-coding region (969–972 base pairs) showed nucleotide variations of which 48 were caused by transition. These nucleotide substitutions were abundant in the region proximate to tRNA^Pro. In addition to the nucleotide substitutions, the dog mtDNA D-loop sequences had a heteroplasmic repetitive sequence (TACACGTÀCG) involving size variation. The DNA sequences of the non-coding region were classified into four different groups by phylogenetic analysis and the deepest branchpoints of this dog phylogeny was calculated to about 100 000 years before the present. Phylogenetic analysis showed that Japanese native dog breeds could not be clearly delimited as distinct breeds. Many haplotypes found in members of some clustering groups were seen in each dog breed, and interbreed nucleotide differences between Japanese dog breeds were almost the same as the intrabreed nucleotide diversities.     

Dimerization of the RelA protein of Escherichia coli.

The RelA protein of Escherichia coli is a ribosome-associated (p)ppGpp synthetase that is activated by amino acid deprivation. It was recently reported that the activity of RelA is regulated by oligomerization mediated by the C-terminal domain of RelA. The oligomerization of RelA is further characterized in this study. The C-terminal domain consisting of amino acids 455-744, designated 'RelA, formed homooligomers as well as heterooligomers with RelA as demonstrated by copurification of RelA and 'RelA and by an affinity blotting assay. Glutaraldehyde-induced cross-linking indicated that the oligomer was a dimer. The functional analysis of 'RelA was based on a combination of yeast two-hybrid analysis, the determination of the effects of overexpression of 'RelA derivatives on the stringent response, and the cellular localization of the overexpressed 'RelA derivatives. These studies indicated that two regions, designated 'RelA-1 (amino acids 455-538) and 'RelA-2 (amino acids 550-682), were involved in dimerization. The involvement of one of these two regions, RelA-2, is consistent with a previous site-directed mutagenesis study. In addition to dimerization, 'RelA-2 apparently contained the main ribosome-binding domain of RelA. The third region, 'RelA-3 (amino acids 682-744), was not involved in either dimerization or ribosome binding. The overexpression of 'RelA-1 and 'RelA-2, but not 'RelA-3, inhibited the stringent response. These results support the previously proposed model which suggests a role for oligomerization in the regulation of (p)ppGpp synthetase.     

Sundasalanx(Sundasalangidae) is a progenetic clupeiform, not a closely‐related group of salangids (Osmeriformes): mitogenomic evidence

Partitioned Bayesian analysis of whole mitochondrial genome sequences from 30 basal teleosts confidently placed Sundasalanx (Sundasalangidae) within the Clupeiformes, not Osmeriformes as originally thought. This study represents the first demonstration of the phylogenetic position of Sundasalanx on a phylogenetic tree.