Substance-P-like immunoreactive subepithelial nerve plexuses in the labial mucosa of the mouse

Substance P (SP)-like immunoreactivity was examined in the lower labial mucosa of the mouse by using the whole-mount technique. The density and design of subepithelial nerve plexuses containing SP differed depending on portions of the lower labial mucosa.     

The role of 5'-methylthioadenosine on rat calvaria cell differentiation.

The role of 5'-methylthioadenosine (MTA), formed during the process of polyamine biosynthesis, on differentiation of osteoprogenitor cells was assessed by its effects on alkaline phosphatase (ALP) activity, bone nodule formation and osteopontin contents of cultured rat calvaria (RC) cells. These three markers were stimulated by exogenous MTA and were depressed by 5'-difluoromethylthioadenosine (DFMTA), a synthetic inhibitor of MTA phosphorylase, which cleaves MTA to adenine and 5-methylthioribose-1-phosphate. 5-Methylthioribose and 2-keto-4-methylthiobutyrate, metabolites of 5-methylthioribose-1-phosphate, had no effects on ALP activity and bone nodule formation in the presence or absence of DFMTA. On the other hand, adenine enhanced ALP activity, bone nodule formation and osteopontin contents in mineralized nodules and also partially reversed DFMTA-induced inhibition of these three markers. MTA, its metabolites and DFMTA did not affect the growth of RC cells under these culture conditions. These results suggest that adenine formed from MTA is important in the differentiation of RC cells.     

Concentrations of essential elements after repeated administrations of tin and selenium

To study effects of simultaneous administration of tin (Sn) and selenium (Se) on concentrations of several essential elements, mice were injected with either SnCl2 (ip) or Na2SeO3 (sc), alone or both compounds at a daily dose of 5 mumol/kg each for 12 consecutive days. Mice were sacrificed at 20 h after the last injection and concentrations of Sn, Se, Na, Ca, Zn, P, Fe, K, and Mg in the liver, kidney, spleen, pancreas, testis, seminal vesicle, lung, femoral muscle, and femoral bone were determined. In the control mice, Sn and Se concentrations were the highest in bone (0.69 micrograms Sn and 6.93 micrograms Se/g dry wt). Administered Sn was found to accumulate in all organs except the testis. Among the essential elements determined, Na was the most affected in terms of concentration in the organs and Mg was the least affected element in these organs. Among the organs tested, each elemental concentration in the pancreas was most affected. Simultaneous injections of Sn and Se appeared to keep the correlation coefficients between elements similar to those found in the control mice.     

Analysis of an ABA-responsive rice gene promoter in transgenic tobacco

We have analyzed in transgenic tobacco the expression of a chimeric gene containing 5 sequences of the rice rab-16B gene fused to the -glucuronidase (GUS) reporter gene. This construct, a translational fusion (–482 to +184) including 14 amino acids of the RAB-16B protein, is expressed only in zygotic and pollen-derived embryos. In zygotic embryos, GUS activity begins to accumulate 10 days after flowering (daf), and increases until seed maturation at 25 daf. Immunological measurements of endogenous abscisic acid (ABA) accumulation in these seeds showed a close parallel between hormone levels and GUS activity. However, GUS activity could not be reproducibly induced by treatment of immature embryos with ABA (10 M). Neither GUS activity nor GUS mRNA could be detected in leaves of transgenic tobacco even after ABA treatment. In contrast, GUS activity could be induced to high levels in pollen-derived embryos by treatment with ABA. Our results show that 482 bp of 5 sequences of the rice rab-16B promoter can confer in transgenic tobacco developmentally regulated expression in embryos but not ABA-responsive expression in vegetative tissues.     

Preparation of biologically active Ascaris suum mitochondrial tRNAMet with a TV-replacement loop by ligation of chemically synthesized RNA fragments.

Ascaris suum mitochondrial tRNA Met lacking the entire T stem was prepared by enzymatic ligation of two chemically synthesized RNA fragments. The synthetic tRNA could be charged with methionine by A.suum mitochondrial extract, although the charging activity was considerably low compared with that of the native tRNA, probably due to lack of modification. Enzymatic probing of the synthetic tRNA showed a very similar digestion pattern to that of the native tRNA Met, which has already been concluded to take an L-shape-like structure [Watanabe et al. (1994) J. Biol. Chem., 269, 22902-22906]. These results suggest that the synthetic tRNA possesses almost the same conformation as the native one, irrespective of the presence or absence of modified residues. The method of preparing the bizarre tRNA used here will provide a useful tool for elucidating the tertiary structure of such tRNAs, because they can be obtained without too much difficulty in the amounts necessary for physicochemical studies such as NMR spectroscopy.     

MOLECULAR PHYLOGENETIC ANALYSIS OF rbcL IN THE PRYMNESIOPHYTA1

The nucleotide sequences of rbcL genes encoding the large subunit of ribulose-1,5-bisphosphate carboxylase/oxygenase (Rubisco) were determined from six species of Prymnesiophyta to clarify their phylogenetic relationships. Molecular phylogenetic trees were constructed using PAUP (Phylogenetic Analysis Using Parsimony). These analyses suggest that the Prymnesiophyta, except for the Pavlovales, area relatively stable monophyletic group. Pleurochrysis carterae, included in the Isochrysidales, is a sister species of a monophyletic group consisting of other members of the Isochrysidales, Gephyrocapsa oceanica and Emiliania huxleyi, members of the Coccosphaerales, Calyptrosphaera sphaeroidea and Umbilicosphaera sibogae var. foliosa, and a member of the Prymnesiales, Chrysochromulina hirta. The nucleotide sequence of rbcL from G. oceanica was identical to that from E. huxleyi within the region examined. Our trees show that G. oceanica and E. huxleyi are more closely related to C. hirta than to U. sibogae, C. sphaeroidea, and P. carterae. These results suggest that orders in the Prymnesiophyceae, including the above-mentioned genera, should be redefined.     

Inhibition of endoplasmic reticulum (ER)-to-Golgi transport induces relocalization of binding protein (BiP) within the ER to form the BiP bodies.

Immunofluorescence staining of yeast cells with anti-binding protein (BiP) antibodies shows uniform staining of the endoplasmic reticulum (ER). We have found that overproduction of Sec12p, an ER membrane protein, causes a change of BiP distribution within the cell. Upon induction of Sec12p by the GAL1 promoter, the staining pattern of BiP turns into bright dots scattering in the cell, whereas the staining of Sec12p remains to be the typical ER figure. Overproduction of other ER membrane proteins, HMG-CoA reductase or Sed4 protein, does not induce such relocalization of BiP. Pulse-chase experiments and electron microscopy have revealed that the overproduction of Sec12p inhibits protein transport from the ER to the Golgi apparatus. When the transport is arrested by one of the sec mutations that block the ER-to-Golgi step at the restrictive temperature, the BiP staining also changes into the punctate pattern. In contrast, the sec mutants that block later or earlier steps of the secretory pathway do not induce such change of BiP localization. These observations indicate that relocalization of BiP is caused by the inhibition of ER-to-Golgi transport. Using immunoelectron microscopy, we have found that the punctate staining is because of the accumulation of BiP in the restricted region of the ER, which we propose to call the "BiP body." This implicates existence of ER subdomains in yeast. A vacuolar protein, proteinase A, appears to colocalize in the BiP body when the ER-to-Golgi transport is blocked, suggesting that the BiP body may have a role as the site of accumulation of cargo molecules before exit from the ER.