Direct identification of the putative surface IgM receptor-associated molecule encoded by murine B cell-specific mb-1 gene

The B cell-specific mb-1 gene was recently reported to encode a putative surface glycoprotein with CD3-like structural properties. Hombach et al. suggested and presented evidence to show that this mb-1 gene encodes the 34-kDa membrane glycoprotein (B34 or IgM-alpha) associated with IgMR molecule. To identify the mb-1 gene product directly in B cells, affinity-purified MB-1-specific antibody was prepared by immunization of rabbits with synthetic MB-1 oligopeptide. Immunoprecipitation in combination with two-dimensional diagonal gel electrophoresis analysis revealed that this antibody detected a B cell-specific surface glycoprotein that is very similar to the IgM-alpha (B34) protein described by Hombach et al. However, MB-1 protein exists usually as the monomeric form on the surface of B cells, in contrast to IgM-alpha, which was detected as the dimeric (IgM-alpha/IgM-alpha or IgM-alpha/Ig-beta) protein. We also found that MB-1 protein is already expressed on the sIgM- pre-B cell lymphoma, which might suggest an alternative functional role of this B cell-specific MB-1 protein in B cell differentiation. The molecular identity of MB-1 protein and IgM-alpha (B34) is discussed.     

Statistical Studies Using AIC Method to Decide the Question of "Break" or "Straight" in Arrhenius Plots of Water Proton NMR Relaxation Times in Chilling-Sensitive Vigna and Insensitive Pisum Seedlings

The question of "break" or "straight" in Arrhenius plots forthe temperature dependency of NMR relaxation times (T₁) of waterprotons in etiolated intact seedlings for chilling-sensitivetwo Vigna species and chilling-insensitive Pisum was statisticallystudied using the Akaike's Information Criterion (AIC), a versatileprocedure for statistical model identification. Among sevenmodels, the most appropriate was based on the following assumptions:individual k (preparation numbers) two-half lines connectingat the break point (TCB) and imposing no restrictions for thegradient and the break point (Model 7). The worst two modelswere one straight line or one TCB (Model 1 or 4). Thus, thedata obtained from replicated preparations should not be treatedas a whole but as individual in each sample case. Break pointsdetermined with Model 7 ranged around 11-5°C for V. radiataand 17-10°C for V. mungo. The implication of the occurrenceof "break" for Pisum clearly differed from Vigna judging bythe model fitness based on the AIC values. The question of "break"or "straight" in Arrhenius plots is therefore fairly dependenton the validity of the model selection in the statistical analysis,and the AIC method is a useful procedure for the resolutionof the problem of the "use" or "misuse" of Arrhenius plots inplant physiology. (Received November 2, 1988; Accepted January 17, 1989)     

In vitro induction of HLA-restricted cytotoxic T lymphocytes against autologous Epstein-Barr Virus transformed B lymphoblastoid cell line

Cytotoxic T lymphocytes (CTL) against autologous EBV-transformed B lymphoblastoid cell line (LCL) were induced in vitro by culturing peripheral blood lymphocytes (PBL) of healthy donors together with mitomycin C-treated autologous LCL for 6 days. The cytotoxic cells developed only from the E-rosette-positive fraction but not from the negative fraction of PBL. These CTL killed autologous LCL but not PWM-stimulated autologous PBL. In addition, the CTL killed allogeneic LCL when at least 1 of the HLA-A antigens was identical with that of the LCL of CTL donor. However, identity of HLA-B and HLA-C antigens was not enough for a significant killing of allogeneic LCL. The specificity of the CTL was also confirmed by a cold target inhibition test. These results indicated that the CTL induced specifically recognized EBV-transformed cells with HLA restriction.     

Isolation of Drosophila flightless mutants which affect myofibrillar proteins of indirect flight muscle

Summary A large number of dominant flightless mutants of Drosophila were chemically induced, and their thorax proteins were examined by means of two-dimensional gel electrophoresis (O'Farrell 1975). Among them, 26 lines were found to have deficiency or reduction of some of myofibrillar proteins in indirect flight muscle (IFM). The gel patterns of the mutants could be classified into eleven groups. In general, more than a few polypeptides were either absent or reduced in each mutant line. Although the mutations affect myofibrillar proteins in apparently complex and diverse ways, logical correlations were found among the changes. There are pairs of proteins which always change together when a number of mutants are compared. There are also many pairs in which presence of one protein is necessary, but not sufficient for presence of the other. This suggests that absence of one component leads to disappearance or reduction of others which are either spatially or functionally related to the former. The correlation is possibly due to a hierarchy of the proteins in the myofibrillar assembly processes.Chromosomal loci of eleven typical mutants were examined, and it was found that most of them are located in two small regions of the second and the third chromosomes. IFM myofibrils of these mutants are either abnormal or absent in homozygotes as well as in heterozygotes.     

Induction and regulation of immunoglobulin expression in a murine pre-B cell line, 70Z/3. I. Cell cycle-associated induction of sIgM expression and kappa-chain synthesis in 70Z/3 cells by LPS stimulation

Stimulation of a murine pre-B cell line, 70Z/3, with LPS induced sIgM expression and an increase of L kappa-chain mRNA within 12 hr. In synchronized 70Z/3 cells, the cell division cycle was 10 to 11 hr and LSP-stimulation did not affect the cell division cycle. LPS-signals given at G1/S boundary induced sIgM expression in the G2 to M-phase. On the other hand, LPS-signals given after M-phase did not induce sIgM expression. Inhibition of cell division with demecorcin did not affect sIgM expression in the M-phase. Induction of intracellular kappa-chain synthesis was also observed in the G2 to M-phase when the LPS-stimulation was provided at G1/S boundary, but LPS-signals given after the M-phase did not induce de novo synthesis of kappa-chain. These results showed that LPS-induced sIgM expression and synthesis of kappa-chain were cell cycle-related events and sIgM expression was associated wih de novo synthesis of kappa-chain.     

Effect of NaCl addition on nanosecond O2 escaping reaction of myoglobin: evidences for the transition of myoglobin dynamic structure at 20 degrees C.

We studied the nanosecond (ns) geminate O2 escape reaction from the protein interior of myoglobin (Mb) to the solvent phase in the temperature range of 5-40 degrees C containing 0-0.1 M NaCl. In the flash photolysis experiments, we found that both the rate constant, kout, and its Arrhenius plot changed upon the variation of the NaCl concentration. In particular, it was noteworthy that the Arrhenius plot of kout dramatically changed in its slope, keeping the break at 20 degrees C, upon the addition of NaCl, indicating that the thermodynamic parameters such as an enthalpy of activation (delta H not equal to) and an entropy of activation (delta S not equal to) are different between above and below 20 degrees C, and that they are further altered upon the NaCl addition to the sample solution. From these results, we suggested that the Mb dynamic structure in the ns geminate O2 escape reaction is sensitively regulated by the interaction of the protein surface and the salt. The present study also showed that an inconsistency of the Arrhenius plot of kout between Chatfield et al. ((1990) J. Am. Chem. Soc. 112, 4680-4687) and us ((1990) J. Biol. Chem. 265, 18823-18828) is probably due to the difference in the solution condition.     

Titration of botulinum antitoxin of low levels by the score method

Botulinum antitoxin is commonly titrated by injecting a mixture of toxin and antitoxin into mice and by utilizing deaths as a marker to measure the amount of unneutralized toxin. We attempted to titrate antitoxin by converting the severity of symptoms (notably palsy) and time-to-death in days into scores. In neutralization tests with toxin levels at 5.9 LD50 and 23.5 LD50, a linear relationship was obtained for antitoxin dose in a range between 0.03 to 0.003 IU/ml. Statistical analysis showed that homogeneity of variance or slope was not denied for the scores obtained on any day from the first to the fourth days after injection, demonstrating that this method can titrate accurately antitoxin of such a low level as 0.003 IU/ml within 4 days after injection.     

Chromosomal assignment in mouse of matrix Gla protein and bone Gla protein genes.

Matrix Gla protein (MGLAP) and bone Gla protein (BGLAP) are calcium-binding, vitamin K-dependent proteins produced by cells of the osteoblastic lineage. Sequence homology suggests that the genes for these proteins evolved from a common ancestor. Somatic whole cell hybrids and karyotypically simple microcell hybrids were used to map Mglap to mouse Chromosome 6 and Bglap to mouse Chromosome 3. Human MGLAP has previously been mapped to chromosome 12p, a region with homology to mouse Chromosome 6, and human BGLAP has been mapped to chromosome 1q, a region with homology to mouse Chromosome 3. It appears that BGLAP is the third calcium-binding protein that maps to human chromosome 1q and mouse Chromosome 3.     

Effects of thiamine and pyridoxine on the content and composition of sterols in Saccharomyces carlsbergensis 4228

The level of sterols in $\text{S. carlsbergensis}$ 4228 cells grown aerobically on a synthetic medium fortified with thiamine was significantly low compared with that in the control cells. The levels of free and esterified sterols in the thiamine-cells were 60% and 10% of the corresponding sterol levels in the control cells, respectively. Analysis by gas-liquid chromatography of non-saponifiable lipids extracted from the cells revealed that the amounts of squalene, lanosterol and two unidentified sterols were higher than those in the control cells and that ergosterol and zymosterol, major sterols in the control cells, were not present. These effects of thiamine on the content and composition of sterols were abolished by the addition of pyridoxine to the medium.     

The isolation of metallothionein and its protective role in cadmium poisoning

People that have been subjected to cadmium poisoning show marked calcified tissue and kidney disturbances. In rats fed a cadmium-containing, low-calcium-vitamin D-deficient diet, the major portion of the cadmium accumulated in the liver and kidneys. Despite the fact that only a small amount (2.8 ppm) of cadmium completely inhibits the in vitro enzymic 1-hydroxylation reaction of 25-hydroxycholecalciferol, the in vivo 1-hydroxylation proceeded without appreciable inhibition even in the rats loaded with large amounts of oral cadmium. No light-microscopic morphological changes could be found in the kidneys of cadmium-fed rats. Most of the cadmium that accumulated in the kidneys was in a form bound to the protein, metallothionein, and therefore was not toxic to that organ. On the other hand, only 20% of the cadmium present in bone appears to be protein bound. The data strongly suggest that the protective effect of metallothionein in the kidney is serendipitous when involved in cadmium poisoning and that cadmium ion acts directly on bone rather than by an indirect action through a functional disturbance of the kidney.