Regulation of ascorbate oxidase expression in pumpkin by auxin and copper

Ascorbate oxidase expression in pumpkin (Cucurbita spp.) tissues was studied. Specific ascorbate oxidase activities in pumpkin leaf and stem tissues were about 2 and 1.5 times that in the fruit tissues, respectively. In seeds, little ascorbate oxidase activity was detected. Northern blot analyses showed an abundant ascorbate oxidase mRNA in leaf and stem tissues. Fruit tissues had lower levels of ascorbate oxidase mRNA than leaf and stem tissues. Ascorbate oxidase mRNA was not detected in seeds. Specific ascorbate oxidase activity gradually increased during early seedling growth of pumpkin seeds. The increase was accompanied by an increase in ascorbate oxidase mRNA. When ascorbate oxidase activity in developing pumpkin fruits was investigated, the activities in immature fruits that are rapidly growing at 0, 2, 4, and 7 d after anthesis were much higher than those in mature fruits at 14 and 30 d after anthesis. The specific activity and mRNA of ascorbate oxidase markedly increased after inoculation of pumpkin fruit tissues into Murashige and Skoog's culture medium in the presence of an auxin such as 2,4-dichlorophenoxyacetic acid (2,4-D) but not in the absence of 2,4-D. In the presence of 10 mg/L of 2,4-D, ascorbate oxidase mRNA was the most abundant. Thus, ascorbate oxidase is induced by 2,4-D. These results indicate that ascorbate oxidase is involved in cell growth. In pumpkin callus, ascorbate oxidase activity could be markedly increased by adding copper. Furthermore, immunological blotting showed that the amount of ascorbate oxidase protein was also increased by adding copper. However, northern blot analyses showed that ascorbate oxidase mRNA was not increased by adding copper. We suggest that copper may control ascorbate oxidase expression at translation or at a site after translation.     

Functional analyses of the Dof domain, a zinc finger DNA-binding domain, in a pumpkin DNA-binding protein AOBP

AOBP, a DNA-binding protein in pumpkin, contains a Dof domain that is composed of 52 amino acid residues and is highly conserved in several DNA-binding proteins of higher plants. The Dof domain has a significant resemblance to Cys2/Cys2 zinc finger DNA-binding domains of steroid hormone receptors and GATA1, but has a longer putative loop where an extra Cys residue is conserved. We show that the Dof domain in AOBP functions as a zinc finger DNA-binding domain and suggest that the Cys residue uniquely conserved in the putative loop might negatively regulate the binding to DNA.     

High-throughput classification of images of cells transfected with cDNA clones

The sequence of the human genome has been determined. The next task is to determine the function of the genes. Classifying cellular forms of proteins encoded by human cDNA clones is a primary step toward understanding the biological role of proteins and their coding genes. We report here our ongoing work on an automatic system to facilitate this classification. Our system handles the transfection, incubation, acquisition of microscopic images of the cells, and the classification of forms there appearing in the images. Our system correctly classified proteins by their forms at a rate of 90% in feasibility studies.     

Pathological steps of cancer-related lymphedema: histological changes in the collecting lymphatic vessels after lymphadenectomy

Introduction

To date, an electron microscopy study of the collecting lymphatic vessels has not been conducted to examine the early stages of lymphedema. However, such histological studies could be useful for elucidating the mechanism of lymphedema onset. The aim of this study was to clarify the changes occurring in collecting lymphatic vessels after lymphadenectomy.

Methods

The study was conducted on 114 specimens from 37 patients who developed lymphedema of the lower limbs after receiving surgical treatment for gynecologic cancers and who consulted the University of Tokyo Hospital and affiliated hospitals from April 2009 to March 2011. Lymphatic vessels that were not needed for lymphatico venous anastomosis surgery were trimmed and subsequently examined using electron microscopy and light microscopy.

Results

Based on macroscopic findings, the histochemical changes in the collecting lymphatic vessels were defined as follows: normal, ectasis, contraction, and sclerosis type (NECST). In the ectasis type, an increase in endolymphatic pressure was accompanied by a flattening of the lymphatic vessel endothelial cells. In the contraction type, smooth muscle cells were transformed into synthetic cells and promoted the growth of collagen fibers. In the sclerosis type, fibrous elements accounted for the majority of the components, the lymphatic vessels lost their transport and concentrating abilities, and the lumen was either narrowed or completely obstructed.

Conclusions

The increase in pressure inside the collecting lymphatic vessels after lymphadenectomy was accompanied by histological changes that began before the onset of lymphedema.     

Outstanding Performance of Hole‐Blocking Layer‐Free Perovskite Solar Cell Using Hierarchically Porous Fluorine‐Doped Tin Oxide Substrate

Perovskite solar cells (PSCs) are of great interest in current photovoltaic research due to their extraordinary power conversion efficiency of ≈20% and boundless potentialities. The high efficiency has been mostly obtained from TiO₂‐based PSCs, where TiO₂ is utilized as a hole‐blocking, mesoporous layer. However, trapped charges and the light‐induced photocatalytic effect of TiO₂ seriously degrade the perovskite and preclude PSCs from being immediately commercialized. Herein, a simplified PSC is successfully fabricated by eliminating the problematic TiO₂ layers, using instead a fluorine‐doped tin oxide (FTO)/perovskite/hole–conductor/Au design. Simultaneously, the sluggish charge extraction at the FTO/perovskite interface is overcome by modifying the surface of the FTO to a porous structure using electrochemical etching. This surface engineering enables a substantial increase in the photocurrent density and mitigation of the hysteretic behavior of the pristine FTO‐based PSC; a remarkable 19.22% efficiency with a low level of hysteresis is obtained. This performance is closely approaching that of conventional PSCs and may facilitate their commercialization due to improved convenience, lower cost, greater stability, and potentially more efficient mass production.     

A neutralizable epitope is induced on HGF upon its interaction with its receptor cMet

A new conformational neutralizable epitope is created on heptocyte growth factor (HGF), when it interacts with its receptor, cMet. By immunizing rabbits with HGF-cMet complex, we successfully generated a monoclonal antibody (SFN68) that inhibits HGF-cMet interaction, and blocks the biological function mediated by HGF. To define the epitope, we screened out an epitope-mimicking peptide, KSLSRHDHIHHH, from a phage display of combinatorial peptide library. In molecular mimicry this peptide bound to cMet and inhibited HGF-cMet interaction. No humoral response was induced to this epitope-mimicking peptide when immunization was done with HGF alone.     

Carcinoma of the Lower Uterine Segment (LUS): Clinicopathological Characteristics and Association with Lynch Syndrome

Endometrial cancer arises from the uterine body and fundus in many cases, but can also originate from the lower region of the uterine body through the upper region of the cervix. Such tumors are referred to as carcinoma of the lower uterine segment (LUS) or isthmus, and account for 3-6.3% of all cases of endometrial cancer. This relatively low incidence has permitted performance of only small-scale studies, but the clinical and pathological characteristics of carcinoma of the LUS in all these reports have differed from those of other endometrial cancers. Generally, endometrial cancer is classified into estrogen-dependent endometrioid adenocarcinoma (designated as type I), and non-endometrioid types that are less associated with estrogen and include poorly differentiated adenocarcinoma (type II). In some reports, carcinoma of the LUS has been found to have type II characteristics. Carcinoma of the LUS has also been associated with Lynch syndrome, a hereditary disease with frequent development of colorectal, endometrial, and ovarian cancers. Lynch syndrome is thought to be induced by mismatch repair gene mutation. The frequency of Lynch syndrome in cases of general endometrial cancer is 1-2%. In contrast, the frequency in patients with carcinoma of the LUS is much higher, with up to 29% of cases diagnosable with Lynch syndrome and a high frequency of hMSH2 mutation found in one study. This suggests that further investigation of the clinical and pathological characteristics of carcinoma of the LUS and the association with Lynch syndrome is required through performance of a large-scale survey.     

Cloning of the Pumpkin Ascorbate Oxidase Gene and Analysis of a Cis-Acting Region Involved in Induction by Auxin

A genomic clone encoding ascorbate oxidase was isolated frompumpkin (Cucurbita sp.)- This gene is consisted of four exonsand three introns. Analyses of the promoter fusion to ß-glucuronidasereporter gene by transient expression assay in pumpkin fruittissues suggested the existence of a cis-acting region responsiblefor auxin regulation. (Received November 28, 1996; Accepted March 8, 1997)