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Atrophic autoimmune thyroiditis (AAT) is a type of autoimmune hypothyroidism without goiter. TSH receptor-blocking antibodies (TSBAb) are involved in its etiology in adults. Reportedly, this disease is extremely rare in children. In this study, we aimed to investigate the prevalence of TSBAb during AAT onset in children using a commercially available cell-based bioassay TSAb kit. We conducted a multicenter retrospective observational study. We collected data of patients with AAT who were < 15 yr old, enrolled in a collaborative research group, and diagnosed since July 2003. AAT was defined as acquired autoimmune hypothyroidism without thyroid enlargement. Eighteen patients (including 15 females) whose TSH receptor antibody (TRAb) or TSBAb levels were measured within a year from the initial visit were included. The median age at diagnosis was 9.3 years, and the estimated time between onset and diagnosis was 2.6 yr. The positive rate for either TSBAb or TRAb was 38.8% (95% confidence interval: 18.3–59.5%). There were no significant differences in age, the estimated time between onset and diagnosis, and FT4 levels at diagnosis between the TSBAb-positive and -negative groups. Unlike previous reports, we showed that the prevalence of TSBAb-positivity in childhood-onset AATs is not rare, as in adults.

At the initial stage of carcinogenesis, newly emerging transformed cells are often eliminated from epithelial layers via cell competition with the surrounding normal cells. For instance, when surrounded by normal cells, oncoprotein RasV12-transformed cells are extruded into the apical lumen of epithelia. During cancer development, multiple oncogenic mutations accumulate within epithelial tissues. However, it remains elusive whether and how cell competition is also involved in this process. In this study, using a mammalian cell culture model system, we have investigated what happens upon the consecutive mutations of Ras and tumor suppressor protein Scribble. When Ras mutation occurs under the Scribble-knockdown background, apical extrusion of Scribble/Ras double-mutant cells is strongly diminished. In addition, at the boundary with Scribble/Ras cells, Scribble-knockdown cells frequently undergo apoptosis and are actively engulfed by the neighboring Scribble/Ras cells. The comparable apoptosis and engulfment phenotypes are also observed in Drosophila epithelial tissues between Scribble/Ras double-mutant and Scribble single-mutant cells. Furthermore, mitochondrial membrane potential is enhanced in Scribble/Ras cells, causing the increased mitochondrial reactive oxygen species (ROS). Suppression of mitochondrial membrane potential or ROS production diminishes apoptosis and engulfment of the surrounding Scribble-knockdown cells, indicating that mitochondrial metabolism plays a key role in the competitive interaction between double- and single-mutant cells. Moreover, mTOR (mechanistic target of rapamycin kinase) acts downstream of these processes. These results imply that sequential oncogenic mutations can profoundly influence cell competition, a transition from loser to winner. Further studies would open new avenues for cell competition-based cancer treatment, thereby blocking clonal expansion of more malignant populations within tumors.

Summary At the early stage of cancer development, oncogenic mutations often cause multilayered epithelial structures. However, the underlying molecular mechanism still remains enigmatic. By performing a series of screenings targeting plasma membrane proteins, we have found that collagen XVII (COL17A1) and CD44 accumulate in RasV12-, Src-, or ErbB2-transformed epithelial cells. In addition, the expression of COL17A1 and CD44 is also regulated by cell density and upon apical cell extrusion. We further demonstrate that the expression of COL17A1 and CD44 is profoundly upregulated at the upper layers of multilayered, transformed epithelia in vitro and in vivo. The accumulated COL17A1 and CD44 suppress mitochondrial membrane potential and reactive oxygen species (ROS) production. The diminished intracellular ROS level then promotes resistance against ferroptosis-mediated cell death upon cell extrusion, thereby positively regulating the formation of multilayered structures. To further understand the functional role of COL17A1, we performed comprehensive metabolome analysis and compared intracellular metabolites between RasV12 and COL17A1-knockout RasV12 cells. The data imply that COL17A1 regulates the metabolic pathway from the GABA shunt to mitochondrial complex I through succinate, thereby suppressing the ROS production. Moreover, we demonstrate that CD44 regulates membrane accumulation of COL17A1 in multilayered structures. These results suggest that CD44 and COL17A1 are crucial regulators for the clonal expansion of transformed cells within multilayered epithelia, thus being potential targets for early diagnosis and preventive treatment for precancerous lesions.

Recent studies have revealed that newly emerging transformed cells are often apically extruded from epithelial tissues. During this process, normal epithelial cells can recognize and actively eliminate transformed cells, a process called epithelial defence against cancer (EDAC). Here, we show that mitochondrial membrane potential is diminished in RasV12-transformed cells when they are surrounded by normal cells. In addition, glucose uptake is elevated, leading to higher lactate production. The mitochondrial dysfunction is driven by upregulation of pyruvate dehydrogenase kinase 4 (PDK4), which positively regulates elimination of RasV12-transformed cells. Furthermore, EDAC from the surrounding normal cells, involving filamin, drives the Warburg-effect-like metabolic alteration. Moreover, using a cell-competition mouse model, we demonstrate that PDK-mediated metabolic changes promote the elimination of RasV12-transformed cells from intestinal epithelia. These data indicate that non-cell-autonomous metabolic modulation is a crucial regulator for cell competition, shedding light on the unexplored events at the initial stage of carcinogenesis.

POU class 1 homeobox 1 (POU1F1) regulates pituitary cell-specific gene expression of somatotropes, lactotropes, and thyrotropes. In humans, two POU1F1 isoforms (long and short isoform), which are generated by the alternative use of the splice acceptor site for exon 2, have been identified. To date, more than 30 POU1F1 mutations in patients with combined pituitary hormone deficiency (CPHD) have been described. All POU1F1 variants reported to date affect both the short and long isoforms of the POU1F1 protein; therefore, it is unclear at present whether a decrease in the function of only one of these two isoforms is sufficient for disease onset in humans. Here, we described a sibling case of CPHD carrying a heterozygous mutation in intron 1 of POU1F1. In vitro experiments showed that this mutation resulted in exon 2-skipping of only in the short isoform of POU1F1, while the long isoform remained intact. This result strongly suggests the possibility, for the first time, that isolated mutations in the short isoform of POU1F1 could be sufficient for induction of POU1F1-related CPHD. This finding improves our understanding of the molecular mechanisms, and developmental course associated with mutations in POU1F1.

Perturbations of the central spindle by depletion of a microtubule nucleation regulator, augmin, revealed its unexpected contributions to the control of cleavage furrow ingression, as well as to cytokinesis completion. These are achieved through nonredundant targeting mechanisms of cytokinesis regulators., During anaphase, distinct populations of microtubules (MTs) form by either centrosome-dependent or augmin-dependent nucleation. It remains largely unknown whether these different MT populations contribute distinct functions to cytokinesis. Here we show that augmin-dependent MTs are required for the progression of both furrow ingression and abscission. Augmin depletion reduced the accumulation of anillin, a contractile ring regulator at the cell equator, yet centrosomal MTs were sufficient to mediate RhoA activation at the furrow. This defect in contractile ring organization, combined with incomplete spindle pole separation during anaphase, led to impaired furrow ingression. During the late stages of cytokinesis, astral MTs formed bundles in the intercellular bridge, but these failed to assemble a focused midbody structure and did not establish tight linkage to the plasma membrane, resulting in furrow regression. Thus augmin-dependent acentrosomal MTs and centrosomal MTs contribute to nonredundant targeting mechanisms of different cytokinesis factors, which are required for the formation of a functional contractile ring and midbody.

At the initial stage of carcinogenesis, transformation occurs in single cells within the epithelium. Recent studies have revealed that the newly emerging transformed cells are often apically eliminated from epithelial tissues. However, the underlying molecular mechanisms of this cancer preventive phenomenon still remain elusive. In this study, we first demonstrate that myosin-II accumulates in Src-transformed cells when they are surrounded by normal epithelial cells. Knock-down of the heavy chains of myosin-II substantially diminishes apical extrusion of Src cells, suggesting that accumulated myosin-II positively regulates the apical elimination of transformed cells. Furthermore, we have identified β-spectrin as a myosin-II-binding protein under the coculture of normal and Src-transformed epithelial cells. β-spectrin is also accumulated in Src cells that are surrounded by normal cells, and the β-spectrin accumulation is regulated by myosin-II. Moreover, knock-down of β-spectrin significantly suppresses apical extrusion of Src cells. Collectively, these results indicate that accumulation of the myosin-II-spectrin complex plays a positive role in apical extrusion of Src-transformed epithelial cells. Further elucidation of the molecular mechanisms of apical extrusion would lead to the establishment of a novel type of cancer preventive medicine.

A gradient of Aurora B activity determines the distribution of the microtubule depolymerase Kif2A at the central spindle and specifies the subsequent spindle structure necessary for proper cytokinesis., The central spindle is built during anaphase by coupling antiparallel microtubules (MTs) at a central overlap zone, which provides a signaling scaffold for the regulation of cytokinesis. The mechanisms underlying central spindle morphogenesis are still poorly understood. In this paper, we show that the MT depolymerase Kif2A controls the length and alignment of central spindle MTs through depolymerization at their minus ends. The distribution of Kif2A was limited to the distal ends of the central spindle through Aurora B–dependent phosphorylation and exclusion from the spindle midzone. Overactivation or inhibition of Kif2A affected interchromosomal MT length and disorganized the central spindle, resulting in uncoordinated cell division. Experimental data and model simulations suggest that the steady-state length of the central spindle and its symmetric position between segregating chromosomes are predominantly determined by the Aurora B activity gradient. On the basis of these results, we propose a robust self-organization mechanism for central spindle formation.

Electron tomography and 3D modeling identifies augmin-dependent connections between the wall of one microtubule and the minus end of a neighboring one in the spindle., The formation of a functional spindle requires microtubule (MT) nucleation from within the spindle, which depends on augmin. How augmin contributes to MT formation and organization is not known because augmin-dependent MTs have never been specifically visualized. In this paper, we identify augmin-dependent MTs and their connections to other MTs by electron tomography and 3D modeling. In metaphase spindles of human cells, the minus ends of MTs were located both around the centriole and in the body of the spindle. When augmin was knocked down, the latter population of MTs was significantly reduced. In control cells, we identified connections between the wall of one MT and the minus end of a neighboring MT. Interestingly, the connected MTs were nearly parallel, unlike other examples of end–wall connections between cytoskeletal polymers. Our observations support the concept of augmin-dependent MT nucleation at the walls of existing spindle MTs. Furthermore, they suggest a mechanism for maintaining polarized MT organization, even when noncentrosomal MT initiation is widespread.

We investigated thyroid function and urinary iodine concentration (UIC) in seven patients with severe motor intellectual disabilities. All seven received total enteral nutrition (TEN) for more than three years with a daily iodine intake of less than 20 µg. They were diagnosed as hypothyroidism due to iodine deficiency (HID) because of high TSH levels (7.6–82.3 µIU/ml), lower free T4 (FT4 0.4–1.5 ng/dl), negative anti-thyroid antibodies (anti-thyroglobulin antibody, anti-thyroidal peroxidase antibody) and extremely low UIC (

Noonan syndrome is an autosomal dominant disease characterized by dysmorphic features, webbed neck, cardiac anomalies, short stature and cryptorchidism. It shows phenotypic overlap with Costello syndrome and cardio-facio-cutaneous (CFC) syndrome. Noonan syndrome and related disorders are caused by germline mutations in genes encoding molecules in the RAS/MAPK pathway. Recently, a gain-of-function mutation in SHOC2, p.S2G, has been identified as causative for a type of Noonan-like syndrome characterized by the presence of loose anagen hair. In order to understand the contribution of SHOC2 mutations to the clinical manifestations of Noonan syndrome and related disorders, we analyzed SHOC2 in 92 patients with Noonan syndrome and related disorders who did not exhibit PTPN11, KRAS, HRAS, BRAF, MAP2K1/2, SOS1 or RAF1 mutations. We found the previously identified p.S2G mutation in eight of our patients. We developed a rapid detection system to identify the p.S2G mutation using melting curve analysis, which will be a useful tool to screen for the apparently common mutation. All the patients with the p.S2G mutation showed short stature, sparse hair and atopic skin. Six of the mutation-positive patients showed severe mental retardation and easily pluckable hair, and one showed leukocytosis. No SHOC2 mutations were identified in leukemia cells from 82 leukemia patients. These results suggest that clinical manifestations in SHOC2 mutation-positive patients partially overlap with those in patients with typical Noonan or CFC syndrome and show that easily pluckable/loose anagen hair is distinctive in SHOC2 mutation-positive patients. Journal of Human Genetics (2010) 55, 801-809; doi:10.1038/jhg.2010.116; published online 30 September 2010

The contractile ring, which is required for cytokinesis in animal and yeast cells, consists mainly of actin filaments. Here, we investigate the directionality of the filaments in fission yeast using myosin S1 decoration and electron microscopy. The contractile ring is composed of around 1,000 to 2,000 filaments each around 0.6 μm in length. During the early stages of cytokinesis, the ring consists of two semicircular populations of parallel filaments of opposite directionality. At later stages, before contraction, the ring filaments show mixed directionality. We consider that the ring is initially assembled from a single site in the division plane and that filaments subsequently rearrange before contraction initiates.

Amelogenin is a novel enamel matrix protein. Knockout mice showed enhanced osteoclast formation and resorption of tooth cementum. This study investigated the effects of amelogenin on osteoclastogenesis. In co-cultures with calvaria osteoblasts and purified bone marrow cells, amelogenin inhibited osteoclastogenesis dramatically. Furthermore, amelogenin inhibited the expression of receptor activator of nuclear factor kappaB ligand (RANKL), macrophage-colony stimulating factor (M-CSF) and fibronectin in osteoblasts, while RANKL expression was induced by fibronectin and inhibited by treatment with fibronectin small interfering RNA. These results suggest that the inhibitory effects of amelogenin on osteoclastogenesis lead to downregulation of RANKL, M-CSF and fibronectin production in osteoblasts.

A three-body abrasion test with a loose abrasive grain scattered on a variety of plastic counterface materials is conducted for Cercidiphyllum japonicum wood (katsura wood). The effect of the counterface material in rubbing with the katsura wood is investigated. The results show that a peak wear coefficient exists for the axial, tangential and radial sections of the katsura wood specimen when rubbed with a counterface material. The peak in the wear coefficient is also recognized in the plastic specimen experiments. The peaks in three-body abrasion experiments for both the katsura wood and plastic specimens are closely related to the variable of material yield stress. The peak on katsura wood specimen occurs when the yield stress of the counterface material is approximately twice as large as that of katsura wood, and the peak on the plastic specimen occurs when the yield stress of the counterface material is approximately the same as that of the specimen. The difference in the results between the katsura wood and plastic material could appear to be due to the change in embedding balance of the loose abrasive grain, which is likely affected by the porous wood structure.

The genetic variability of 125 respiratory syncytial virus (RSV) subgroup A isolates over 15 successive epidemics from 1980 to 1995 in an urban population of Japan was determined. Allocation of isolates into lineages was archived by reverse transcriptase-polymerase chain reaction amplification of selected regions of the nucleoprotein (NP) and attachment (G) protein gene followed by restriction fragment length polymorphism (RFLP) analysis. Three and seven distinct restriction patterns of the NP and G gene were observed, respectively. When the NP and G gene RFLP analyses were combined, ten different genetic lineages were identified in the 125 isolates. The strains with the same genotype were isolated in each epidemic and the dominant lineages were replaced by others after every one to three consecutive epidemics. Nucleotide and amino acid sequencing of the variable region of G gene of these predominant isolates revealed differences of 5–28% between strains. There was, however, no apparent accumulation of diversity with age to indicate progressive changes. The dominant strains were often closely related to those isolated in other parts of the world at a similar time. These observations suggest that dominant RSV strains are replaced frequently by others that have been co-circulating or have recently entered the community from a worldwide reservoir. The change of dominant strains may be influenced by the buildup of immunological resistance in the community to successive epidemics of the same strain. J. Med. Virol. 64:374–380, 2001. © 2001 Wiley-Liss, Inc.

An S4-allele-associated protein (S4-protein) was identified by both isoelectricfocusing (IEF) polyacrylamide-gel electrophoresis and N-terminal amino-acid sequences in self-incompatible Nijisseiki (self-in- compatibility genotype=S2S4) and self-compatible Osa-Nijisseiki (S2S4 SM, SM=stylar-part mutant) styles of the Japanese pear, Pyrus serotina Rehd. var. culta Rehd. Expression of the protein in the cultivars was compared during flower bud development and at post-transcriptional levels. In mature styles, S4-protein could be detected on IEF gel in both cultivars and the N-terminal amino acid sequences were identical, although Osa-Nijisseiki contained only one-third the amount contained in Nijisseiki. No difference was observed in S2-protien amounts. In Nijisseiki styles, S4-protein was already detectable 8 days before anthesis (DBA) and it was synthesized consistently until 2 days after anthesis (DAA); the amount increased 4.7-fold during these 10 days. In contrast, S4-protein in Osa-Nijisseiki was not detected earlier than 6 DBA; a small amount was found at 4 DBA, and it increased gradually as flowers developed. Thus, expression of Osa-Nijisseiki S4-protein is developmentally controlled in the same way as that of Nijisseiki S4-protein, but with a time lag of several days; the protein level at 2 DAA corresponded to that of Nijisseiki earlier than 4 DBA. S4-proteins from both Nijisseiki and Osa- Nijisseiki showed RNase activity and the activity was also developmentally controlled; it increased about fourfold during the interval from 8 DBA to 2 DAA in Nijisseiki, and 3.3-fold during the interval from 4 DBA to 2 DAA in Osa-Nijisseiki. Activity at 2 DAA, however, was twice as high in Nijisseiki. In vitro protein synthesis showed that poly(A)+ from mature Osa-Nijisseiki styles could form S4-like protein in a manner similar to that of Nijisseiki. These results suggest that the self-compatibility of Osa-Nijisseiki is due to a low level of S4-protein expression, a mechanism very similar to the low level of S-protein and weak incompatibility in immature styles of self-incompatible Nijisseiki. Part of this protein repression may be regulated during post-transcriptional events.

Neurotrophin 4 (NT-4) and its receptors regulate the differentiation of ameloblasts in tooth development. Gangliosides, sialic acids that contain glycosphingolipids (GSLs), are involved in a variety of membrane-associated cell physiological functions such as ligand-receptor signal transmission. However, the expression patterns and functions of GSLs during tooth development remain unclear. In this study, we identified strong expressions of GM3 and LacCer in dental epithelium, which give rise to differentiation into enamel-secreting ameloblasts. Exogenous GM3 and LacCer in dental epithelial cells induced the expression of ameloblastin ( Ambn), while it was also interesting that GM3 synergistically exerted enhancement of NT-4-mediated Ambn expression. In addition, consistently exogenous GM3 and LacCer in dental epithelial cells induced distinct activation of extracellular signal-regulated kinase 1/2 (ERK1/2), an event upstream of the expression of Ambn. Furthermore, depletion of GSLs from dental epithelial cells by D- threo-1-phenyl-2-decanoylamino-3-morpholino-1-propanol (D-PDMP) inhibited Ambn expression as well as phosphorylation of ERK1/2. In contrast, exogenous addition of GM3 or LacCer rescued the phosphorylation of ERK1/2 repressed by pre-treatment with D-PDMP. Taken together, these results suggest that GM3 and LacCer are essential for NT-4-mediated Ambn expression, and contribute to dental epithelial cell differentiation into ameloblasts.

The vascularization is an important factor along with the wound healing process, and there are some reports on extraction wounds. However, the ultrastructure of newly developed capillaries in the healing process is still argued. It is aimed of clearing the ultrastructural early healing process of endothelial cells in the experimental tooth extraction wound by electron microscopy.Immediately after the extraction, the diffused basal lamina intermittently surrounded endothelial cells. Three days after the extraction, the basal lamina was not continuous, but had no gap with endothelial cells. Four days after the extraction, the basal lamina almost reconstructed.On tannic acid injection into blood vessels, it was observed in the narrow lumen of cell masses which continued to capillaries. It is suggested that cell masses are derived from the endothelial cells, and are the precursor of capillaries.The cell masses were formed by two or three undifferentiated endothelial cells. The narrow lumen was observed in the cell mass and these cells project short folds into the lumen. These cells connected by tight junctions, but the basal lamina was not complete. On tannic acid injection, it does not perfuse into the stroma through the basal lamina. It is concluded that these types of connections between the endothelial cells, were one of the characteristic features of capillaries in the tooth extraction wound.

FBN2, FBN1, TGFBR1, and TGFBR2 were analyzed by direct sequencing in 15 probands with suspected congenital contractural arachnodactyly (CCA). A total of four novel FBN2 mutations were found in four probands (27%, 4/15), but remaining the 11 did not show any abnormality in either of the genes. This study indicated that FBN2 mutations were major abnormality in CCA, and TGFBR and FBN1 defects may not be responsible for the disorder. FBN2 mutations were only found at introns 30, 31, and 35 in this study. Thus analysis of a mutational hotspot from exons 22 to 36 (a middle part) of FBN2 should be prioritized in CCA as previously suggested.

Noonan syndrome (NS) is characterized by short stature, characteristic facial features, and heart defects. Recently, missense mutations of PTPN11, the gene encoding protein tyrosine phosphatase (PTP) SHP-2, were identified in patients with NS. Further, somatic mutations in PTPN11 were detected in childhood leukemia. Recent studies showed that the phosphatase activities of five mutations identified in NS and juvenile myelomonocytic leukemia (JMML) were increased. However, the functional properties of the other mutations remain unidentified. In this study, in order to clarify the differences between the mutations identified in NS and leukemia, we examined the phosphatase activity of 14 mutants of SHP-2. We identified nine mutations, including a novel F71I mutation, in 16 of 41 NS patients and two mutations, including a novel G503V mutation, in three of 29 patients with leukemia. Immune complex phosphatase assays of individual mutants transfected in COS7 cells showed that ten mutants identified in NS and four mutants in leukemia showed 1.4-fold to 12.7-fold increased activation compared with wild-type SHP-2. These results suggest that the pathogenesis of NS and leukemia is associated with enhanced phosphatase activity of mutant SHP-2. A comparison of the phosphatase activity in each mutant and a review of previously reported cases showed that high phosphatase activity observed in mutations at codons 61, 71, 72, and 76 was significantly associated with leukemogenesis.

In January 2001, 20 children among 40 residents under 2 years old at a nursery home in Sapporo, Japan had respiratory symptoms and were confirmed as having respiratory syncytial virus (RSV) infection by a conventional diagnostic kit. Nasopharyngeal aspirates were collected from four RSV-positive patients and total RNA was extracted directly from the specimens for the analysis of RSV grouping and genotyping. All four RSV strains had the same G protein gene sequence of subgroup B and were assigned to identical strains. Interestingly, the G protein gene had a duplication of 60 nucleotides at the C-terminal third of the G protein gene in which three nucleotides differed each other. The predicted polypeptide is lengthened by 20 amino acids. The clinical picture of these cases was not different from those of patients with other RSV strains. These novel mutations were thought to be introduced in vivo.

GD3 synthase is one of the key enzymes involved with ganglioside synthesis, and its activity regulates the main profile of ganglioside expression. We analyzed the expression of the GD3 synthase gene in laser-dissected teeth germs using RT-PCR. The GD3 synthase gene was found expressed in brain, thymus, and tooth germ tissues, however, not in liver or skin specimens. Further, it was highly expressed during the early stage of tooth germ development (embryonic day 14.5), especially in dental epithelia, which gradually reduced in the molar site until postnatal day 7, whereas it was not in dental mesenchyme tissues. In addition, dental epithelial cells transiently transfected with the GD3 synthase gene showed enhanced proliferation. These results indicate that the GD3 synthase gene may be involved in early tooth development, particularly in the proliferation of dental epithelium.

The yeast cell walls of the budding yeast Saccharomyces cerevisiae are well studied and the results show the existence of a framework composed of beta-1,3-glucan. It is reported that the cell wall of the fission yeast Schizosaccharomyces pombe has different components and our analysis by 13C-nuclear magnetic resonance (NMR) spectroscopy also showed there is alpha-1,3-glucan in its cell wall. To refine our understanding of the architecture of the yeast cell wall, we re-examined the cell wall glucans of S. pombe by NMR spectroscopy and prepared antibody against alpha-1,3-glucan, which is a characteristic component of this yeast. By the competitive enzyme-linked immunosorbent assay (ELISA) system, specificity of the antibody was restricted to alpha-1,3-glucan, which did not take a highly ordered structure. We analysed the localization of the cell wall glucans by immunoelectron microscopy. Transmission electron microscope (TEM) images showed that most of the alpha-1,3-glucan was along the cell membrane and appeared to enclose the cytoplasm, supporting previous reports that this glucan is synthesized on the cell membrane.

The genetic variability of 32 respiratory syncytial virus (RSV) sub-group B isolates from a single community in Japan during the 20 years from 1980 to 1999 was determined. Two variable regions of the attachment (G) protein gene were amplified by reverse transcriptase-polymerase chain reaction amplification and their products were sequenced directly. Phylogenetic analysis of nucleotide sequences revealed seven distinct branches in which strains isolated during seasons of close proximity were located: however, isolates from the same season were often in plural branches. There was a tendency for recent isolates to lie at the end of each branch and these linear evolutionary changes were typically represented in a branch containing nine strains isolated during 6 seasons from '80 to '86. Three kinds of usages of stop codons were confirmed and isolates located in each branch used the same stop codons. These observations suggest that there are multiple subgroup B lineages co-circulating and that each lineage strain may exhibit linear evolutionary genetic drifts in order to survive over successive epidemics within the same population, although it has a conserved uniform G protein length with the use of the same stop codon.

Longitudinal F-actin cables are thought to be important for transporting materials for polarized cell growth in fission yeast. We show that most F-actin in the cables is oriented such that the barbed end faces the nearest cell tip during interphase; however, this directionality is reversed during mitosis. These orientations of F-actin ensure proper transport of materials to growing sites during these cell-cycle stages.

A simple and rapid analytical method for detecting interactions between oligosaccharides of glycoproteins and different lectins was studied by surface plasmon resonance using a biosensor (BIAcore). The interactions of three lectins, Sambucus sieboldiana agglutinin (SSA), Ricinus communis agglutinin I (RCA I) and Concanavalin A (Con A) for fetuin and digested fetuins with glycosidases, asialo-, agalacto-, and aglucosamino-fetuin, were investigated as a model system. These fetuins were immobilized to the matrix of of the sensor chip and the lectins were injected into the sensor chip cartridge. The association and dissociation reactions could be monitored as resonance signals in real time. The interactions with lectins significantly changed as the oligosaccharides of fetuin were trimmed. The interactions between fetuin and SSA, asialofetuin and RCA I, and aglucosaminofetuin and Con A show the highest affinity properties, respectively. The association constants of these lectins were estimated to be 1.4 x 10(7), 1.9 x 10(8) and 5.3 x 10(7) (M-1), respectively. These results suggested that the interactions between lectins and glycoproteins could be well defined in real time and kinetically, and that the partial structure of oligosaccharides of glycoproteins can be estimated by determination of the interactions with various lectins after glycosidase digestions using the biosensor.

Effects of taurine on endogenous aspartic acid (Asp), glutamic acid (Glu) and gamma-aminobutyric acid (GABA) release has been investigated using synaptosomes prepared from rat cerebral cortex. Although basal release of these amino acids was not affected, taurine inhibited KCl (30 mM)-evoked overflow of Asp, Glu and GABA in a concentration-dependent manner with potencies (IC50) of 1 microM, 0.8 microM and 5 nM, respectively. Taurine (10 microM) maximally inhibited K(+)-evoked Asp, Glu and GABA overflow by 28, 37 and 65%, respectively. Phaclofen (10 microM, a GABAB receptor antagonist), but not bicuculline (10 microM, a GABAA receptor antagonist), counteracted the inhibition of GABA overflow, although the inhibition of Asp and Glu overflow was not attenuated. These data suggest that taurine may inhibit GABA release through the activation of presynaptic GABAB autoreceptors and, at high concentration, also act on Asp- and Glu-nerve terminals to regulate release of excitatory amino acids in rat cortex.

A simple system for continuous mass rearing of Rhagoletis pomonella (Walsh) was developed by using all sizes of ‘Golden Delicious’ apples as the larval rearing medium. From 25,000 to 50,000 pupae per week can be produced with about 20 hours of labor. A 10-fold net increase per generation was achieved easily. About 9 generations were completed in 1½ years.

Adult Laspeyresia pomonella (L.) were treated with Cobalt-50 (Yakima, Washington) and Cesium-137 (Vincennes, Indiana) at levels of 10, 15, 25, and 40 krad and evaluated by 5 criteria, male longevity, female longevity, mating, oviposition, and egg hatch. When a sterilizing dose of 25 krad was given to males only, the effects of cobalt on mating and egg hatch were significantly different from the effects of cesium; however, at a dose of 40 had, only the frequency of mating on the basis of the 5 criteria was significantly different. The mating Combinations (treated × untreated insects) and the doses significantly affected the moths compared with the untreated controls, regardless of source. Also, though some of the differences in the responses were real between the 2 sources and locations, the trends were similar, and both types of treatments effectively sterilized the adult codling moth.

The seasonal distribution of Synanthedon pictipes (Grote & Robinson) in 1969 on Washington Island was determined by (1) studying adult emergence from larvae on borer-infested cherry trees and (2) recording the number of male moths captured in field traps baited with caged virgin females. The estimates of population by the 2 methods were similar: a population of 16,008 borers was obtained from a larval survey and a population of 21,646 borers was obtained by using the percentage of native moths captured vs. the number of marked released males recaptured. These studies showed that 1 generation of moths emerged in 1969 from June to early September.

Larvae of Anastrepha suspensa (Loew) (Diptera: Tephritidae) were reared in Florida on the artificial larval diet developed for the Mexican fruit fly, A. ludens (Loew). Adults reproduced adequately when they were fed sugar, orange juice crystals, a hydrolyzed yeast protein, and water. At 27°C, duration of the stages in the life cycle of the fly was determined to be: egg 3 days; larva 8 days; pupa 12 days. Three generations of the fly were reared successfully.

In tests conducted in Hawaii from 1959 to 1964 both sexes of one or more of 3 species of tephritid flies were sterilized without toxic effects by treating food and water with tcpa, metepa, apholate, or tretamine, applying these materials topically to pupae or adults, or exposing adults to deposits of the chemosterilants. Methotrexate, aminopterin, colchicine, and 5-fluorouracil treatments sterilized females only. Tepa, apholate, and tretamine sterilized as effectively and efficiently as ionizing radiation. Tepa showed promise for field applications in combination with attractive protein hydrolyzates. In general, males were sterilized at lower concentrations than females; the melon fly, Dacus cucurbitae Coquillett, was the most susceptible; the oriental fruit fly, D. dorsalis Hendel, intermediate; and the Mediterranean fruit fly, Ceratitis capitata (Wiedemann), the least susceptible to test materials. Treatments were most effective against newly emerged flies, but deposition of hatchable eggs in old gravid fertile females was inhibited within 24–48 hr after treatment.

when a bait spray (a 1:4 mixture of 95% technical malathion and an acid hydrolysate of corn protein) was repealedly applied to small portions of navel orange trees in an orchard in Coatepec, Veracruz, Mexico, the number of adult Anastrepha ludens (Loew) trapped in the treated area was 85-98% less than the number trapped in the check area, even though reinfestation from surrounding untreated groves was probably constant. Also, the number of larvae-per-kilo was reduced as much as 84% in tree fruit and slightly more in fallen fruit. The fruit produced in the treated areas was of local market quality except at the spring peak of the population of the flies and probably could have been exported after fumigation. The increase in the numbers of whiteflies, probably cloudy-winged whiteflies, Dialeurodes citrifolii (Morgan), in the treated area may be attributable to the increased mortality of green lacewings, chrysopa sp.