Formation of Recombinants Between Nontransmissible Drug-Resistance Determinants and Transfer Factors

Noninfectious drug-resistance determinants acquired conjugal transmissibility by the formation of recombinants with transfer factors, suggesting the origin of R factors.     

Some observations on the solitary male among Japanese monkeys

The social behavior pattern of a solitary male at Koshima was studied by means of radio-telemetry. The relationship between the solitary males and the troop was estimated from radio-tracking data of the former's location and movement, and by direct observation of the latter at each corresponding hour.For most of day, the solitary male stayed within a distance of about 20 to 150 m from the central part of the troop, occasionally approaching it. His movement also was synchronized with that of the troop. For two nights, the solitary male slept at places which were about 200 m from the sleeping sites of the troop and faced them across the beach. The relationship between the solitary male and the troop did not seem to be strongly antagonistic.It can be assumed that the solitary male was moving according to certain pre-determined relationships or social contacts with the troop. The example of this solitary male shows the existence of the solitary male that follows and maintains contact with the troop, even outside the copulatory season.This study was sponsored by Scientific Research Grant No. 91620 of the Ministry of Education to the Japan Monkey Centre.     

Regulatory role of CD4/L3T4 molecules in IL-2 production by affecting intracellular Ca2+ concentration of T cell clone stimulated with soluble anti-CD3

To elucidate the role of CD4 molecule in T cell activation, the effect of anti-CD4 on T cell IL-2 production was examined by using an alloreactive Th clone. The alloreactive T cell used in the present experiments produced IL-2 in response to soluble anti-CD3 epsilon-chain (anti-CD3) without accessory cell or insoluble antibody carrier. The IL-2 production was suppressed by the addition of anti-CD4 in cultures. An intracellular free Ca2+ concentration ([Ca2+]i) of the T cell clone was elevated by anti-CD3 stimulation, but the elevation was suppressed in the presence of anti-CD4. When the clone was stimulated in Ca2(+)-free medium, the elevation of [Ca2+]i was not observed. When Ca2+ influx was induced by calcium ionophore A23187 or ionomycin, the clone produced IL-2 in response to anti-CD3 in the presence of anti-CD4. When polyclonal T cell line or several other alloreactive T cell clones were examined for their anti-CD3 response, essentially the same results as mentioned above were obtained. Taken together, these results suggest that the slow and sustained elevation of [Ca2+]i is an essential signal for IL-2 production of T cells, and that anti-CD4 suppresses the IL-2 production by interfering the [Ca2+]i elevation. The significance of CD4 molecules in murine T cell activation was discussed.     

Construction of Libraries for Methylation Sites by In-gel Competitive Reassociation (IGCR)

The in-gel competitive reassociation (IGCR) procedure was successfullyapplied to construct a comprehensive library enriched in DNAfragments containing C^5mCGG sequences from mouse liver and braingenomic DNA. For IGCR, methylation-insensitive restriction enzyme(Msp I) digests were used as target DNA and methylation-sensitiverestriction enzyme (Hpa II) digests as competitor DNA. Southernblot analysis indicated that 60 to 70% of the clones in thelibrary were derived from the methylated sites and overall enrichmentwas 200- to 1000-fold. IGCR was further applied to constructa library for the sites differentially methylated between brainand liver DNA. In the library, approximately 20% of the HpaII sites exhibited different degrees of methylation betweenthese tissues.     

Analysis of the replication region of a mycobacterial plasmid, pMSC262.

We determined the nucleotide sequence of a DNA fragment which contains the replication region of pMSC262, a Mycobacterium scrofulaceum plasmid used to construct the Mycobacterium-Escherichia coli shuttle vector. The complete sequence of the fragment contained 2,504 bp with an overall G+C content of 69.8%. By deletion analysis, we found that the minimum length required for plasmid replication in M. bovis BCG was about 1.6 kb. Within this region, several open reading frames (ORFs) and a putative replication origin (ori) were identified by computer analysis. One of the ORFs, ORF2, which encodes a putative 28.9-kDa basic protein with characteristics of DNA-binding proteins, appeared to be involved in replication of the plasmid in BCG. By separation of ORF2 and the putative ori region, it was revealed that the relative locations of ORF2 and the putative ori region are likely important for replication in BCG. No DNA or amino acid homologies were found between this replication region and that of pAL5000, another mycobacterial plasmid used for vector plasmid construction. In addition, we found that this replicon did not lead to replication in E. coli and was compatible in BCG with pAL5000-derived vector plasmid pYUB75 (R. G. Barletta, D. D. Kim, S. B. Snapper, B. R. Bloom, and W. R. Jacobs, J., J. Gen. Microbiol. 138:23-30, 1992).     

Theste13+ gene encoding a putative RNA helicase is essential for nitrogen starvation-induced G1 arrest and initiation of sexual development in the fission yeastSchizosaccharomyces pombe

When the fission yeastSchizosaccharomyces pombe is starved for nitrogen, the cells are arrested in the G1 phase, enter the G0 phase and initiate sexual development. Theste13 mutant, however, fails to undergo a G1 arrest when starved for nitrogen and since this mutant phenotype is not suppressed by a mutation in adenylyl cyclase (cyr1), it would appear thatste13 ⁺ either acts independently of the decrease in the cellular cAMP level induced by starvation for nitrogen, or functions downstream of this controlling event. We have used functional complementation to clone theste13 ⁺ gene from anS. pombe genomic library and show that its disruption is not lethal, indicating that, while the gene is required for sexual development, it is not essential for cell growth. Nucleotide sequencing predicts thatste13 ⁺ should encode a protein of 485 amino acids in which the consensus motifs of ATP-dependent RNA helicases of the DEAD box family are completely conserved. Point mutations introduced into these consensus motifs abolished theste13 ⁺ functions. The predicted Ste13 protein is 72% identical to theDrosophila melanogaster Me31B protein over a stretch of 391 amino acids. ME31B is a developmentally regulated gene that is expressed preferentially in the female germline and may be required for oogenesis. Expression of ME31B cDNA inS. pombe suppresses theste13 mutation. These two evolutionarily conserved genes encoding putative RNA helicases may play a pivotal role in sexual development.     

The kinetics of end-product inhibition of l-lactate production from xylose and glucose by Lactococcus lactis IO-1

Summary The relative contributions of lactate inhibition and the generation of sterile (undividing) cells to the low xylose utilisation rate of Lactococcus lactis IO-1 was investigated. The lactate inhibition constant of xylose grown cells was shown to be 9.3 times more than that of glucose grown cells. However, the sterile cell production rate and LDH inactivation rate of the xylose cultures were at least 10 times less than the glucose cultures. Thus, it is suggested that the slower substrate consumption rate in xylose medium is caused mainly by the large inhibition constant for the end product.     

Methenamine-Silver Staining: a Simple and Sensitive Staining Method for Senile Plaques and Neurofibrillary Tangles

An improved methenamine-silver impregnation method is presented which exhibits sensitivity for amyloid substances comparable to that of anti-β protein immunostaining. In optimally treated sections, this technique stained both β-amyloid deposits and neurofibrillary tangles, which are known to have a β-pleated structure. This simple procedure allows a large number of sections to be stained for routine examination.