Kimiko Asano

Claudins in Caenorhabditis elegans: Their Distribution and Barrier Function in the Epithelium

Claudins ( approximately 23 kDa) with four transmembrane domains are major cell adhesion molecules working at tight junctions in vertebrates, where the intercellular space is tightly sealed (reviewed in ). We examined here the possible occurrence of claudin-like proteins in invertebrates, which do not bear typical tight junctions. Close blast searching of the C. elegans genome database identified four claudin-related, approximately 20-kDa integral membrane proteins (CLC-1 to -4), which showed sequence similarity to the vertebrate claudins. The expression and distribution of CLC-1 was then examined in detail by GFP technology as well as by immunofluorescence microscopy. CLC-1 was mainly expressed in the epithelial cells in the pharyngeal region of digestive tubes and colocalized with AJM-1 at their intercellular junctions. Then, to examine the possible involvement of CLC-1 in the barrier function, we performed RNA interference in combination with a tracer experiment: in CLC-1-deficient worms, the barrier function of the pharyngeal portion of the digestive tubes appeared to be severely affected. CLC-2 was expressed in seam cells in the hypodermis, and it also appeared to be involved in the hypodermis barrier. These findings indicated that multiple species of the claudin homologs, which are involved in the barrier function of the epithelium, exist in C. elegans.

Changes in the rat liver mitochondrial DNA upon aging

During experiments on the molecular basis of morphological and functional changes observed in rat liver mitochondria upon aging, we found that the buoyant density profile of mitochondrial DNA (mtDNA) shows a wide distribution pattern especially in the lighter region than that of young rat liver mtDNA. The heterogeneous pattern may be partly recovered to become similar to that of young rat liver mtDNA by treatment with proteinase K. Therefore, it is quite likely that mtDNA of old rat liver contains firmly bound protein(s) or peptides. During the morphological observation of mtDNA by electron microscopy, we found that mtDNA of old rat had a novel property, that is, the ability to attach to negatively charged mica in the absence of magnesium ions, although their morphological features showing circular 5 microns contour length form did not change. Further, mtDNA gained resistance against EcoRI digestion during aging. This property was not shared by the DNA from young animal, and might be due to the binding protein(s).

Quantitation of changes in mitochondrial DNA during aging and regeneration of rat liver using non-radioactive DNA probes

By using DNA probes prepared from cloned cells which contain mitochondrial DNA (mtDNA) sequences in plasmids, the specific detection of mtDNA became possible in the presence of large excess of nuclear DNA by DNA-DNA hybridization. For this purpose, we prepared mtDNA probes labeled with non-radioactive substrate, which allowed a wider possibility of application. This method revealed that the contents of mtDNA in rat liver are strikingly decreased during aging. Furthermore, it was observed that although mtDNA content increased upon partial hepatectomy even in old rats, it decreased to the pre-operation level rather rapidly within 1 week after reaching peak in regenerated liver.

Adenosine-3′, 5′-cyclic monophosphate receptor protein in adrenal cortical mitochondria

Abstract The binding of adenosine-3′, 5′-cyclic monophosphate to bovine adrenal cortical mitochondrial subfraction was studied. The binding was mainly in a mitochondrial soluble fraction. The binding properties of this fraction are different from that of cell supernatant.