Elucidation of the critical epitope of an anti-EGFR monoclonal antibody EMab-134

The epidermal growth factor receptor (EGFR) is a type-1 transmembrane receptor tyrosine kinase, which activates the downstream signaling cascades in many tumors, such as oral and lung cancers. We previously developed EMab-134, a novel anti-EGFR monoclonal antibody (mAb), which reacts with endogenous EGFR-expressing cancer cell lines and normal cells independent of glycosylation in Western blotting, flow cytometry, and immunohistochemical analysis. EMab-134 showed very high sensitivity (94.7%) to oral squamous cell carcinomas in immunohistochemical analysis. In this study, we performed enzyme-linked immunosorbent assay (ELISA), flow cytometry, and immunohistochemical analysis to determine the epitope of EMab-134. A blocking peptide (375–394 amino acids of EGFR) neutralized the EMab-134 reaction against oral cancer cells in flow cytometry and immunohistochemistry. The minimum epitope of EMab-134 was found to be the _377-RGDSFTHTPP_?386 sequence. Our findings can be applied for the production of more functional anti-EGFR mAbs that in turn can be used for antitumor treatments.     

Area-ratio Fraunhofer line depth (aFLD) method approach to estimate solar-induced chlorophyll fluorescence in low spectral resolution spectra in a cool-temperate deciduous broadleaf forest

Journal of Plant Research - Solar-induced chlorophyll fluorescence (SIF) emissions were estimated by the "area-ratio Fraunhofer line depth (aFLD) method", a new retrieval methodology in...     

Cloning and Sequencing of a 23-kb Region of the Bacillus subtilis Genome between the iol and hut Operons

Within the framework of an international project for the sequencingof the entire Bacillus subtilis genome, a 23-kb chromosomalsegment, which covers the region between the iol and hut operons,has been cloned and sequenced, creating a 99-kb contig fromthe gnt operon to the wapA locus. This region (23351 bp) contains25 complete open reading frames (ORFs; genes) including deoR,dra, nupC and pdp and two partial ones. The region (5140 bp)containing these four genes, being also sequenced by H. H. Saxildet al., was sequenced by subjecting a long polymerase chainreaction product to random sequencing using phage M13mp19. However,we could detect no conflict, between two independently determinedsequences, which could be attributed to our sequencing method.A homology search for the 24 newly identified gene productsrevealed significant homology to known proteins in 14 of them.It was notable that three proteins, encoded by the successivegenes (yxeMNO), exhibited meaningful homology to the E. coliGlnHPQ products constituting a periplasmic ATP-dependent transportsystem for glutamine.     

Oral administration of Aloe vera gel powder prevents UVB-induced decrease in skin elasticity via suppression of overexpression of MMPs in hairless mice

This study reports the effects of oral Aloe vera gel powder (AVGP) containing Aloe sterols on skin elasticity and the extracellular matrix in ultraviolet B (UVB)-irradiated hairless mice. Ten-week-old hairless mice were fed diets containing 0.3% AVGP for 8 weeks and irradiated UVB for 6 weeks. Mice treated with AVGP showed significant prevention of the UVB-induced decrease in skin elasticity. To investigate the mechanism underlying this suppression of skin elasticity loss, we measured the expression of matrix metalloproteinase (MMP)-2, -9, and -13. AVGP prevented both the UVB-induced increases in MMPs expressions. Moreover, we investigated hyaluronic acid (HA) content of mice dorsal skin and gene expression of HA synthase-2 (Has2). In the results, AVGP oral administration prevented UVB-induced decreasing in skin HA content and Has2 expression and attenuates the UVB-induced decrease in serum adiponectin, which promotes Has2 expression. These results suggested that AVGP has the ability to prevent the skin photoaging.     

Suppressed recombination on mouse chromosome 15 defined regions of chromosomal inversions associated with koala (koa) and hairy ears (eh) mutations.

Koala (Koa) and hairy ears (Eh) mutations of mice are associated with chromosomal inversions in the distal half of chromosome 15. Since these two mutant mice show some common phenotypic features including extra hair on pinna and craniofacial dysmorphogenesis and have similar inverted regions, we determined the inverted regions of these two chromosomal inversions to examine whether a common gene is responsible for the phenotypes of these two mutants. The inverted regions were identified as the recombination-suppressed regions by linkage analysis. The length of the recombination-suppressed regions of Koa and Eh were approximately 52 and 47 Mb, respectively, and these inverted regions were not the same. These results indicate that the phenotypes of Koa and Eh mutant mice are likely to be caused by different genes.     

Dimorphism of the seed-dispersing organ in a pantropical coastal plant,Scaevola taccada: heterogeneous population structures across islands

Scaevola taccada is characterized by dimorphic fruits, with one type having cork and pulp (C-morph) and the other type having only pulp (NC-morph). Although within-individual dimorphism has not been observed, both morphs can occur at the same sites. The cork floats on seawater, and the pulp is eaten by birds. Thus, the morphs may have different seed dispersal abilities, via sea currents and birds, respectively. This study aimed to determine the functional characteristics of the dimorphic fruits. First, the potential seed dispersal ability of sea currents and birds was compared between the two fruits morphs by conducting a floating test and by measuring the proportions of different parts of the fruits, respectively. Next, the frequencies of the two morphs across different substrates (beaches, rocks, and cliffs) in the southern islands around Japan were analyzed. Most C-morph fruits remained floating for more than 180 days in a seawater tank, whereas all NC-morph fruits sank after approximately 1 week. The NC-morph fruits had a more pulp volume and less indigestible material than the C-morph fruits. Although both types of plants were found on many islands and all substrates, the C-morph was dominant on beaches, whereas the NC-morph was most frequent on cliffs. The frequencies of the morphs on different substrates might be influenced by the differences in their seed dispersal abilities. The two morphs may differ in fitness on different substrates. These findings improve our understanding of plant adaptations for dispersal.     

Erratum to: Loss of heterophylly in aquatic plants: not ABA-mediated stress but exogenous ABA treatment induces stomatal leaves in Potamogeton perfoliatus

Journal of Plant Research -     

Tamoxifen feeding method is suitable for efficient conditional knockout

The Cre-driver system is used to generate conditional knockout mice. Tamoxifen inducible Cre-driver mice can be used for spatiotemporal knockout by administration of the drug. A major tamoxifen administration is performed by intraperitoneal administration or oral administration. However, these forced administrations may be damaging to mice. Herein, we have demonstrated an improved method of administering tamoxifen with powdered food to mice. A mouse line expressing the tamoxifen-inducible Cre gene was used ubiquitously in this experiment to evaluate the efficiency of Cre recombination in the whole body. Our method also achieved efficient recombination without causing injury to mice. The X-gal staining intensity of the feeding method was equivalent to that of the intraperitoneal administration method. Furthermore, this method can be used for recombination before birth, or during the fetal period. We recommend researchers to employ this feeding method to administer tamoxifen to minimize the risk of injury to mice.     

Subconductance Gating and Voltage Sensitivity of Sarcoplasmic Reticulum K+ Channels: A Modeling Approach

Sarcoplasmic reticulum (SR) K⁺ channels are voltage-regulated channels that are thought to be actively gating when the membrane potential across the SR is close to zero as is expected physiologically. A characteristic of SR K⁺ channels is that they gate to subconductance open states but the relevance of the subconductance events and their contribution to the overall current flowing through the channels at physiological membrane potentials is not known. We have investigated the relationship between subconductance and full conductance openings and developed kinetic models to describe the voltage sensitivity of channel gating. Because there may be two subtypes of SR K⁺ channels (TRIC-A and TRIC-B) present in most tissues, to conduct our study on a homogeneous population of SR K⁺ channels, we incorporated SR vesicles derived from Tric-a knockout mice into artificial membranes to examine the remaining SR K⁺ channel (TRIC-B) function. The channels displayed very low open probability (Po) at negative potentials (≤0 mV) and opened predominantly to subconductance open states. Positive holding potentials primarily increased the frequency of subconductance state openings and thereby increased the number of subsequent transitions into the full open state, although a slowing of transitions back to the sublevels was also important. We investigated whether the subconductance gating could arise as an artifact of incomplete resolution of rapid transitions between full open and closed states; however, we were not able to produce a model that could fit the data as well as one that included multiple distinct current amplitudes. Our results suggest that the apparent subconductance openings will provide most of the K⁺ flux when the SR membrane potential is close to zero. The relative contribution played by openings to the full open state would increase if negative charge developed within the SR thus increasing the capacity of the channel to compensate for ionic imbalances.