Ultrastructure of chloroplasts in'Marimo'(Cladophora aegagropila, Chlorophyta), and changes after exposure to light

This study compares the ultrastructure of the inner, dark-habituated cells of the green ‘Cladophora-ball’, or Marimo, to that of similar cells at the surface. Cells not exposed to light possess fewer, but larger and more irregular, chloroplasts than do the outer cells. Unexposed chloroplasts have a pyrenoid matrix lacking starch sheaths and containing unusually thick granal stacks. Despite prolonged exposure to darkness, the chloroplasts contain small starch grains. After exposure to light, such chloroplasts divide, become smaller and take on the appearance of those in the outer layer cells. Within 48 h, all of the chloroplasts develop substantial starch grains and the pyrenoids are surrounded by starch sheaths. This response is consistent with previous reports of the recovery of photosynthetic activity in inner cells exposed to light.     

A Digital Image Processing Technique for the Analysis of Particle Movements: Its Application to Organelle Movements during Mitosis in Adiantum Protonemata

A digital image processing technique was developed for the simultaneousdetection of changes in organelle movements in different regionsof a cell from the protonemata of the fern, Adiantum. The speedof particle movements at a chosen position in a series of dynamicimages that had been recorded by a time-lapse video system wasdetermined in terms of standard error of brightness changeswith a gray scale level. Using this new method and microscopy we could distinguish 3different regions during mitosis in terms of organelle movementpatterns. Organelles located outside of the nucleus were inmovement until the nucleolus disappeared at prophase, whereasorganelles in the boundary region between the nucleus and cytoplasmbecame active after prophase. The organelles located outsidethe nucleus again began to move rapidly after chromosome separation.The nuclear pole region showed movement throughout mitosis. ³ Present address: Tbaraki Satellite Communication Center, KokusaiDenshin Denwa Co. Ltd., Takahagi, Ibaraki 318, Japan. ⁴ Present address: Biology Department, Faculty of Science, TokyoMetropolitan University, Fukazawa, Tokyo 158, Japan. (Received September 3, 1982; Accepted December 23, 1982)     

Stimulation of cAMP formation by prostaglandin D2 in primary cultures of bovine adrenal medullary cells: identification of the responsive population as fibroblasts

In primary cultures of bovine adrenal medulla, chromaffin cells responded to prostaglandin (PG) E2 by stimulating phosphoinositide metabolism (Yokohama et al. (1988) J. Biol. Chem. 263, 1119-1122). In contrast, nonchromaffin cells were found to respond to PGD2 by elevating their intracellular cAMP level. The formation of cAMP was detected at as low as 0.1 nM PGD2 and increased more than 100-fold over the basal level at 0.1 microM, and the response was specific for PGD2 (greater than PGE1 greater than PGE2 greater than PGF2 alpha = PGI2). The magnitude of cAMP formation and its specificity to PGD2 were retained throughout a 40-day culture period. Based on the inhibitory effect of cis-4-hydroxy-L-proline, an inhibitor of collagen synthesis, on cAMP formation, morphology, and immunoreactivity of cells to anti-collagen type I antiserum, the responsive cells were identified as fibroblasts. These results taken together demonstrate that the adrenal medulla is composed of chromaffin and nonchromaffin cells, which respond to PGE2 and PGD2, respectively, by two different signal transduction pathways. The cAMP formation by PGD2 was also observed in fibroblasts from bovine embryonic trachea among cell lines tested, suggesting that some populations of fibroblasts responsive to PGD2 exist in various tissues and may discriminate the signal from that of PGE1 or PGE2.     

The generation and consolidation of a radial array of cortical microtubules in developing guard cells of Allium cepa L.

The initiation and development of a radial array of microtubules (MTs) in guard cells of A. cepa was studied using immunofluorescence microscopy of tubulin in isolated epidermal layers. Soon after the completion of cytokinesis, MTs originate in the cortex adjacent to a central strip of the new, anticlinically oriented ventral wall separating the two guard cells. Cortical MTs extend from the mid-region of the central strip toward the cell edge where the ventral wall joins the inner periclinal wall. They then spread in a fan-like formation along the periclinal wall and gradually extend along the lateral and end walls as well. Many MTs criss-cross at various angles as they arc past the edge formed by the junction of the ventral and periclinal walls, but they do not terminate there, indicating that, contrary to previous report, the edge is not involved in MT initiation. Instead, the mid-region of the central strip appears to function as a planar MT-organizing zone. Initially, MTs radiate from this zone through the inner cytoplasm as well as the cortex. During cell expansion, however, the cortical MTs increasingly predominate and consolidate into relatively thick, long bundles, while the frequency of non-cortical MTs diminishes. The apparent density of MTs per unit surface area is maintained as the cells expand and gradually flex into an elliptical shape. The guard cells eventually separate completely at the pore site. The entire process is accomplished within about 12 h.Abbreviations DIC differential interference contrast - GC guard cell - MT microtubule To whom correspondence should be addressed.     

Solution structure of microtubule-associated protein light chain 3 and identification of its functional subdomains

Microtubule-associated protein (MAP) light chain 3 (LC3) is a human homologue of yeast Apg8/Aut7/Cvt5 (Atg8), which is essential for autophagy. MAP-LC3 is cleaved by a cysteine protease to produce LC3-I, which is located in cytosolic fraction. LC3-I, in turn, is converted to LC3-II through the actions of E1- and E2-like enzymes. LC3-II is covalently attached to phosphatidylethanolamine on its C terminus, and it binds tightly to autophagosome membranes. We determined the solution structure of LC3-I and found that it is divided into N- and C-terminal subdomains. Additional analysis using a photochemically induced dynamic nuclear polarization technique also showed that the N-terminal subdomain of LC3-I makes contact with the surface of the C-terminal subdomain and that LC3-I adopts a single compact conformation in solution. Moreover, the addition of dodecylphosphocholine into the LC3-I solution induced chemical shift perturbations primarily in the C-terminal subdomain, which implies that the two subdomains have different sensitivities to dodecylphosphocholine micelles. On the other hand, deletion of the N-terminal subdomain abolished binding of tubulin and microtubules. Thus, we showed that two subdomains of the LC3-I structure have distinct functions, suggesting that MAP-LC3 can act as an adaptor protein between microtubules and autophagosomes.     

Dimeric transthyretin variant assembles into spherical neurotoxins

Familial amyloidotic polyneuropathy is a hereditary autosomal-dominant disease in which the deposited transthyretin fibrils are derived from amyloidogenic mutation. We investigated structure and stability of a human Ser112Ile transthyretin variant and showed that the Ser112Ile variant exists as a dimer having nonnative tertiary structure at physiological pH. In addition, the dimeric Ser112Ile assembles into a spherical aggregate and exerts cytotoxicity in a human neuroblastoma cell line. Our results suggest the importance of an unstable dimeric structure in forming spherical aggregates that will induce cell death.     

Design,synthesis, and evaluation of novel inhibitors for wild-type human serine racemase

Most of the endogenous free d-serine (about 90%) in the brain is produced by serine racemase (SR). d-Serine in the brain is involved in neurodegenerative disorders and epileptic states as an endogenous co-agonist of the NMDA-type glutamate receptor. Thus, SR inhibitors are expected to be novel therapeutic candidates for the treatment of these disorders. In this study, we solved the crystal structure of wild-type SR, and tried to identify a new inhibitor of SR by in silico screening using the structural information. As a result, we identified two hit compounds by their in vitro evaluations using wild-type SR.Based on the structure of the more potent hit compound 1, we synthesized 15 derivatives and evaluated their inhibitory activities against wild-type SR. Among them, the compound 9C showed relatively high inhibitory potency for wild-type SR. Compound 9C was a more potent inhibitor than compound 24, which was synthesized by our group based upon the structural information of the mutant-type SR.     

Chromium(III) ion and thyroxine cooperate to stabilize the transthyretin tetramer and suppress in vitro amyloid fibril formation

Transthyretin (TTR) amyloid fibril formation, which is triggered by the dissociation of tetrameric TTR, appears to be the causative factor in familial amyloidotic polyneuropathy and senile systemic amyloidosis. Binding of thyroxine (T(4)), a native ligand of TTR, stabilizes the tetramer, but the bioavailability of T(4) for TTR binding is limited due to the preferential binding of T(4) to globulin, the major T(4) carrier in plasma. Here, we show that Cr(3+) increased the T(4)-binding capacity of wild-type (WT) and amyloidogenic V30M-TTR. Moreover, we demonstrate that Cr(3+) and T(4) cooperatively suppressed in vitro fibril formation due to the stabilization of WT-TTR and V30M-TTR.     

Polyglutamine tract-binding protein-1 binds to U5-15kD via a continuous 23-residue segment of the C-terminal domain

Polyglutamine tract-binding protein-1 (PQBP-1) is a nuclear protein that interacts with various proteins, including RNA polymerase II and the spliceosomal protein U5-15kD. PQBP-1 is known to be associated with X-linked mental retardation in which a frameshift mutation in the PQBP-1 gene occurs. In the present study, we demonstrate that PQBP-1 binds to U5-15kD via a continuous 23-residue segment within its C-terminal domain. Intriguingly, this segment is lost in the frameshift mutants of PQBP-1 associated with X-linked mental retardation. These findings suggest that the frameshift mutations in the PQBP-1 gene lead to expression of mutants lacking the ability to interact with U5-15kD.