Immunoelectron microscopic localization of fibronectin in the smooth muscle layer of mouse small intestine

We studied the ultrastructural distribution of fibronectin in the smooth muscle layer of mouse small intestine with affinity-purified antibodies using the immunogold technique. Fibronectin was present over the pericellular area extending from the cell membrane to the extracellular matrix beyond the basal lamina. Distribution of the glycoprotein over the pericellular area was heterogeneous, i.e., it was localized more abundantly in the narrow space between smooth muscle cells, the gaps having a width of 60-80 nm where the two dense bands in adjacent cells matched each other. Such localization suggests that fibronectin contributes to cell adhesion. Within the basement membrane, gold label was localized both in lamina lucida and lamina densa, more densely in the latter than in the former. Fibronectin was also co-distributed with collagen fibers in the extracellular matrix. Within smooth muscle cells, gold particles were observed on rough endoplasmic reticulum and secretory vesicle-like structures. These results suggest that smooth muscle cells synthesize fibronectin and secrete it as a component of the basal lamina and extracellular matrix.     

Effect of retinoic acid on in vitro proliferation activity and glycosaminoglycan synthesis of mesenchymal cells from palatal shelves of mouse fetuses

To clarify the mechanism by which retinoid causes cleft palate, we investigated the effect of retinoic acid (RA) on proliferation activity and glycosaminoglycan (GAG) synthesis in mouse fetuses palatal mesenchymal (MFPM) cells. MFPM cells were incubated for 1-11 days with various concentrations of RA to examine its effect on growth rate. Also, confluent cultures were incubated with [3H]glucosamine or [35S]sulfate in the presence of various concentrations of RA to investigate the effect of RA on GAG synthesis. RA remarkably inhibited the growth of MFPM cells in a dose-dependent manner. RA also inhibited the synthesis of GAGs, with sulfated GAGs being more severely affected than hyaluronic acid. These data suggest that the inhibition of proliferation activity and GAG synthesis of palatal mesenchymal cells might be involved in the induction of cleft palate by retinoic acid.     

Spacing and kinship in the Formosan squirrel living in different habitats

Summary Spacing and kinship of the Formosan squirrel, Callosciurus erythraeus thaiwanensis, were studied in two different habitats. One, native habitat in the woods of Kenting, southern Formosa, was rich in available food throughout the year and had several species of predators. The other, a site in Kamakura, central Japan where squirrels had been introduced, had relatively scanty food and few potential predators. 1. Home ranges among males and between sexes overlapped extensively in both habitats. 2. Females occupied exclusive home ranges in Kamakura but had small overlapping home ranges in Ken-ting. 3. Most males disappeared from their natal areas at 1 year old in both habitats (86% in Kamakura and 93% in Ken-ting), but less females disappeared (36% in Kamakura and 35% in Ken-ting). 4. In Kamakura, daughters settled adjacent to the mother or inherited the home range of the mother, but never shared the mother's home range. In Ken-ting, 35% of daughters shared the home range with their mothers. 5. Tolerance among female kin in Ken-ting was probably facilitated by the richness of available food throughout the year, and functioned to reduce predation risk via alarm calling and mobbing.     

The usefulness of CHAPS as a non-cytotoxic stabilizing agent in purification of growth factors

Among several detergents, a zwitterionic detergent, 3-[(3-cholamidopropyl)dimethylammonio]-1-propane sulfonate (CHAPS), was found to be least cytotoxic for cultured mammalian cells. CHAPS improved the activity recovery and elution profile of crude and purified fibroblast growth factors (FGFs) during chromatographies. Diluted preparations of FGFs were stabilized by CHAPS against the loss during storage. Amino acid sequence analysis was not disturbed by CHAPS. CHAPS was removable by reversed-phase high-performance liquid chromatography. These results indicate that CHAPS is useful as a non-cytotoxic stabilizing agent in purification of various kinds of bioactive polypeptides.Abbreviations -MEM Alpha Modification of Eagle's Minimal essential medium - CHAPS 3-[(3-cholamidopropyl)dimethylammonio]-1-propane sulfonate - CHAPSO 3-[(3-cholamidopropyl)dimethylammonio]-2-hydroxy-1-propane sulfonate - CS Calf Serum - EGF Epidermal Growth Factor - FGF Fibroblast Growth Factor - HPLC High-Performance Liquid Chromatography - NGF Nerve Growth Factor - NOG 1-O-n-octyl--D-glucopyranoside - NP-40 Nonidet P-40 - PBS Phosphate-Buffered Saline - SB 12 3-(dodecylmethylammonio)-1-propane sulfonate - SDS Sodium Dodecyl Sulfate - TGF- and Transforming Growth Factor type and      

Heterogeneous distribution of the precursor of type I and type III collagen and fibronectin in the rough endoplasmic reticulum of palatal mesenchymal cells of the mouse embryo cultured in ascorbate-depleted medium

Summary In order to examine the intracellular distribution of precursors of type I and type III collagen and fibronectin in the palatal mesenchymal (MEPM) cells of the mouse embryo cultured under ascorbate-deficient conditions, immuno-electron-microscopic studies were carried out by use of affinity purified antibodies for these proteins. MEPM cells were obtained from the palatal shelves of 14-day-old mouse fetuses and cultured for 3–7 days in medium, either with or without 50 ng/dish/day ascorbic acid. Results obtained were as follows: (1) Although the rough endoplasmic reticulum (rER) of MEPM cells cultured for 5 days in ascorbate-supplemented medium was flattened, that in cells cultured in ascorbate-deficient medium had a distended or vesicular appearance. (2) Vesicular or distended rER showed heterogeneous staining for both type I and type III collagen, namely, some parts of rER showed positive staining for both types of collagen, while others showed negative staining. (3) Both type I and type III collagen showed codistribution in the same vesicular rER. (4) Vesicular rER showed negative or very faint labelling for fibronectin. These results may suggest regional differences in the function of rER.     

Intra- and interspecific variation in body size ofProtohermes (Megaloptera: Corydalidae)

The life history of three populations ofProtohermes grandis and two populations ofProtohermes immaculatus (Megaloptera: Corydalidae) was compared. In general, the larvae lived in stream riffles for 2 years and the adults appeared in summer. Adult body size differed between these closely related species and also between the populations ofP. grandis. Dwarfism occurred inP. immaculatus, a species that is endemic to the small, isolated island, Amami Island. The population ofP. grandis on Yaku Island, located between Amami Island and the mainland Kyushu, had an intermediate body size between that ofP. immaculatus and the mainland population ofP. grandis. Despite being an insular population,P. grandis on Tsushima Island had a similar body size to mainlandP. grandis. In these populations with large adults, some larvae lived in the streams for 3 years. The size distribution of benthic animals, which are the prey available toProtohermes larvae, differed between the streams studied. The density of large prey was lowest on Amami Island, intermediate on Yaku Island, and highest on the mainland and Tsushima Island. Different size distributions of available prey may be caused by the differences of benthic fauna; most of Ecdyonuridae and Ephemerellidae (large mayflies) and Perlidae (large stoneflies) were not found on Amami and Yaku Islands. Thus, there is a tendency to dwarfism in the populations ofProtobermes inhabiting streams where the density of large prey is low.     

Cloning, Nucleotide Sequences and Differential Expression of the nifH and nifH-Like (frxC) Genes from the Filamentous Nitrogen-Fixing Cyanobacterium Plectonema boryanum

The frxC gene, found in liverwort chloroplast DNA, encodes aprotein of unknown function. The deduced amino acid sequenceof the protein shows significant homology to that of ni-trogenaseFe-protein encoded by the nifH gene. We have cloned the frxCand nifH genes from the nitrogen-fixing cyanobacterium Plectonemaboryanum, using frxC- and nifH-specific probes, and have determinedtheir nucleotide sequences. The amino acid sequence deducedfrom the frxC gene of P. boryanum exhibits 83% homology to thatof the protein encoded by the/rxCgene from liverwort, whereasit exhibits only 34% homology to that encoded by the nifH genefrom the same organism, namely, P. boryanum. Northern blot analysisshowed that the frxC gene was transcribed more actively undernitrogenase-repressed conditions than under nitrogenase-inducedconditions, suggesting that the FrxC protein has a functiondistinct from nitrogen fixation. These results, together withthe phylogenetic relationship between the nifH and frxC genes,indicate that the frxC and nifH genes are derived from a commonancestral gene but have evolved independently to encode proteinswith different functions. (Received April 27, 1991; Accepted August 12, 1991)     

Sequence of three members and expression of a new major subfamily of glutelin genes from rice

Three members have been isolated of an additional glutelin gene subfamily, named subfamily B, consisting of about five members per haploid rice genome. Restriction fragment length polymorphism analysis showed major differences between Japonica and Indica lines, indicating the divergence of the subfamily since the split between the two varieties. While corresponding exons of the subfamily B showed 80 to 88% nucleotide sequence homology, those exons were only 60–65% homologous to those of the glutelin A subfamily [15, 19, 24], distinguishing them from the subfamily A. Intron position and derived polypeptide structure, in addition to the nucleotide sequence, confirm the subfamily B members as glutelins. Analysis of RNA from seeds of different stages of development showed that the subfamily B members were expressed at the same time as those of subfamily A, demonstrating coordinated regulation of the two subfamilies.