Suppression of γ-Aminobutyric Acid (GABA) Transaminases Induces Prominent GABA Accumulation, Dwarfism and Infertility in the Tomato (Solanum lycopersicum L.)

Tomatoes accumulate γ-aminobutyric acid (GABA) at high levels in the immature fruits. GABA is rapidly converted to succinate during fruit ripening through the activities of GABA transaminase (GABA-T) and succinate semialdehyde dehydrogenase (SSADH). Although three genes encoding GABA-T and both pyruvate- and α-ketoglutarate-dependent GABA-T activities have been detected in tomato fruits, the mechanism underlying the GABA-T-mediated conversion of GABA has not been fully understood. In this work, we conducted loss-of-function analyses utilizing RNA interference (RNAi) transgenic plants with suppressed pyruvate- and glyoxylate-dependent GABA-T gene expression to clarify which GABA-T isoforms are essential for its function. The RNAi plants with suppressed SlGABA-T gene expression, particularly SlGABA-T1, showed severe dwarfism and infertility. SlGABA-T1 expression was inversely associated with GABA levels in the fruit at the red ripe stage. The GABA contents in 35S::SlGABA-T1RNAi lines were 1.3–2.0 times and 6.8–9.2 times higher in mature green and red ripe fruits, respectively, than the contents in wild-type fruits. In addition, SlGABA-T1 expression was strongly suppressed in the GABA-accumulating lines. These results indicate that pyruvate- and glyoxylate-dependent GABA-T is the essential isoform for GABA metabolism in tomato plants and that GABA-T1 primarily contributes to GABA reduction in the ripening fruits

Effects of Oak Wilt Disease on Fungal Community Composition and Wood Decomposition in Dead Quercus serrata Trunks

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Cross-talk between TLR3 and TNF-α or IFN-γ Signaling in Induction of CXCL8/IL-8 and CXCL10/IP-10 Expression in Airway Epithelial Cells

CXCL8/IL-8 is a chemoattractant for neutrophils and mast cells, and regulates inflammatory cell recruitment in allergy, infection, and other neutrophil-related diseases. Interferon (IFN) -γ-inducible protein 10 (CXCL10/IP-10) is a chemokine that attracts mononuclear cells, Th1 cells, and natural killer cells. We investigated the levels of CXCL8/IL-8 and CXCL10/IP-10 expression by airway epithelial cells after exposure to the inflammatory cytokines tumor necrosis factor (TNF) -α and IFN-γ, and to poly I:C, a synthetic analog of double-stranded RNA that is a ligand of Toll-like receptor 3 (TLR3). Poly I:C, TNF-α, IFN-γ, and combinations of poly I:C with TNF-α or IFN-γ were used to stimulate the airway epithelial cell line BEAS-2B. Following stimulation, we determined CXCL8/IL-8 and CXCL10/IP-10 mRNA levels by real-time PCR and protein levels by ELISA. Poly I:C treatment upregulated mRNA and protein expression for both CXCL8/IL-8 and CXCL10/IP-10. The addition of TNF-α, but not IFN-γ, to poly I:C further increased the expression of CXCL8/IL-8 mRNA and protein. The addition of either TNF-α or IFN-γ to the poly I:C treatment further increased CXCL10/IP-10 mRNA and protein expression. Cross-talk between TLR3 signaling and inflammatory cytokines regulates the expression of CXCL8/IL-8 and CXCL10/IP-10 in airway epithelial cells. From our results, TNF-α and IFN-γ produce different effects on TLR3 signaling

Tomato Glutamate Decarboxylase Genes SlGAD2 and SlGAD3 Play Key Roles in Regulating γ-Aminobutyric Acid Levels in Tomato (Solanum lycopersicum)

Tomato (Solanum lycopersicum) can accumulate relatively high levels of γ-aminobutyric acid (GABA) during fruit development. However, the molecular mechanism underlying GABA accumulation and its physiological function in tomato fruits remain elusive. We previously identified three tomato genes (SlGAD1, SlGAD2 and SlGAD3) encoding glutamate decarboxylase (GAD), likely the key enzyme for GABA biosynthesis in tomato fruits. In this study, we generated transgenic tomato plants in which each SlGAD was suppressed and those in which all three SlGADs were simultaneously suppressed. A significant decrease in GABA levels, i.e. 50–81% compared with wild-type (WT) levels, was observed in mature green (MG) fruits of the SlGAD2-suppressed lines, while a more drastic reduction (up to <10% of WT levels) was observed in the SlGAD3- and triple SlGAD-suppressed lines. These findings suggest that both SlGAD2 and SlGAD3 expression are crucial for GABA biosynthesis in tomato fruits. The importance of SlGAD3 expression was also confirmed by generating transgenic tomato plants that over-expressed SlGAD3. The MG and red fruits of the over-expressing transgenic lines contained higher levels of GABA (2.7- to 5.2-fold) than those of the WT. We also determined that strong down-regulation of the SlGADs had little effect on overall plant growth, fruit development or primary fruit metabolism under normal growth conditions

Tumor Necrosis Factor-alpha and Transforming Growth Factor-beta Synergistically Upregulate Endothelin-1 Expression in Human Bronchial Epithelial Cells BEAS-2B

Endothelin-1 is a peptide with many functions including bronchoconstriction and the stimulation of fibroblasts, and myofibroblasts, and airway smooth muscle cell proliferation. These functions are related to airway remodeling and endothelin-1 is known to be upregulated in the epithelium of patients with severe asthma. We thus sought to elucidate the mechanisms underlying endothelin-1 expression in bronchial epithelial cells in vitro. The human bronchial epithelial cell line BEAS-2B was grown in culture and then treated with tumor necrosis factor-alpha (TNF-α), interleukin-4 (IL-4), interleukin-13 (IL-13), and transforming growth factor-beta (TGF-β). Expression of endothelin-1 mRNA and protein was quantified by real-time polymerase chain reaction and enzyme-linked immunosorbent assay, respectively. We also repressed expression of the key transcription factor in the pathogenesis of severe asthma, nuclear factor-kappa B (NF-κB), using small interfering RNA (siRNA). TNF-α and TGF-β significantly increased the release of endothelin-1 protein into the culture medium of BEAS-2B cells at 24 h after treatment compared to untreated cells; however, the Th2 cytokines, IL-4 and IL-13, had no effect. Endothelin-1 mRNA expression was also upregulated by TNF-α and TGF-β with a peak time point at 4 h after stimulation. Finally, the combination of TNF-α and TGF-β synergistically increased both endothelin-1 protein secretion and mRNA expression, and this upregulation was significantly suppressed in cells transfected with siRNA to repress NF-κB expression. TNF-α and TGF-β synergistically upregulate the expression of endothelin-1 in human bronchial epithelial cells, possibly via the activity of NF-κB. Our findings thus suggest NF-κBa as a potential therapeutic target for the regulation of airway remodeling

Anti-IgE therapy for allergic bronchopulmonary aspergillosis

Allergic bronchopulmonary aspergillosis (ABPA) is a severe type of asthma. Some cases are resistant to treatment, even with regular use of antiasthmatic drugs and antifungal agents. The diagnosis of ABPA was made in a 40-year-old patient with ABPA according to the Rosenberg-Patterson criteria. Symptoms were not controlled despite regular use of antiasthmatic drugs, daily systemic steroids, and antifungal agents. Omalizumab, administered in an attempt to stabilize these uncontrolled symptoms, was effective with no adverse events. Our experience suggests omalizumab is a potential candidate drug for controlling steroid-dependent ABPA

Clarithromycin-resistant Mycobacterium Shinjukuense Lung Disease: Case Report and Literature Review

Mycobacterium shinjukuense is a species of non-tuberculous mycobacteria newly reported in 2010. Herein, we report on an 85-year-old woman with M. shinjukuense lung disease. Radiographic examinations showed consolidation with bronchiectasis, which responded transiently to clarithromycin monotherapy. After 1 year of monotherapy, the diagnosis was established and drug susceptibility testing revealed elevated minimum inhibitory concentration for clarithromycin. The patient was then treated successfully with a combination of antituberculosis drugs. The transient response to clarithromycin suggested that the M. shinjukuense had acquired resistance to clarithromycin. Appropriate treatment for M. shinjukuense lung disease has not yet been established; therefore, it is important to accumulate information from case reports

Untapped work capacity among old persons and their potential contributions to the “silver dividend” in Japan

In contemporary Japan, the utilization of healthy elderly persons in economic production is one of the most urgent policy matters. In this paper, we have measured the untapped work capacity of old persons, using the microdata gathered in the Japanese Study of Aging and Retirement (JSTAR), a longitudinal survey carried out on subjects aged 50–75. Our computed results show that the volume of untapped work capacity of the Japanese elderly aged 60–79 is vast, amounting to more than 11 million workers at present. We have also applied the computed results to the National Transfer Accounts (NTA) framework, and quantified the magnitude of the use of the untapped work capacity upon potential economic growth. The accumulated effect of the economic support ratio upon potential economic growth is substantial in the long term, generating a sizable amount of the so called “silver dividend”. We have also examined the issue of whether or not the use of untapped work capacity provided by old persons could affect the well-being of workers in other age groups. The regression results support the view that the substitutability between the selected age groups of the elderly and the young is negligible, so that the utilization of potential work capacity of elderly persons is unlikely to pose any serious threat to the employment opportunities of their young counterparts in Japan

Expression of ICAM-1 on human bronchial epithelial cells after influenza virus infection

Damage of bronchial epithelium is a feature of airway viral infection and airway inflammatory disease, such as bronchial asthma. Adhesion molecules, which are expressed on bronchial epithelium, play an important role in the pathogenesis of epithelial damage and airway inflammation. We analysed ICAM-1 and VCAM-1 expression on human bronchial epithelial cell line, NCI-H292, after influenza virus A infection. ICAM-1 was expressed on control cells constitutively. Influenza virus A infection caused a three-fold increase in ICAM-1 expression on NCI-H292 cells. Supernatant of virus-infected cells was analysed for the concentration of IL-1β and TNF-α but these cytokines were not detected. VCAM-1 was not expressed on control cells and did not change after cytokine stimulation or virus infection. These findings suggest that influenza virus infection may induce ICAM-1 expression on bronchial epithelium without intervention of leukocytes, and ICAM-1 expressed on epithelium plays a major part in the pathophysiology of airway inflammatory disease caused by viral infection

A long-term impact of forest disturbance on spruce seedling regeneration on coarse woody debris

Storm disturbance have huge impacts on subalpine forest ecosystems. However, long-term effects of such disturbance on regeneration of subalpine forests are poorly understood. Picea jezoensis var. hondoensis has limited and discontinuous distributions in subalpine central Japan. Thus, evaluating long-term effects of forest disturbance and their mechanisms have great conservation significance. Picea needs coarse woody debris (CWD) such as logs and stumps for their seedling colonization, and thus their establishment is greatly affected by CWD condition which largely depending on decay activity of decomposer fungal community. Recent studies in Europe found that frequency of occurrence of brown rot fungi, a certain functional group of fungi which decay wood holocellulose without decaying lignin, tends to be increase after forest dieback1), and that CWD decayed by brown rot fungi negatively affects spruce seedling density2). Because the decay process of CWD is known to be several decades long, we hypothesized that the effect of forest disturbance on fungal community and wood decay of CWD, and its negative effect on spruce seedling establishment would be long lasting. To test this hypothesis, we compared fungal communities within CWD and spruce seedling density among forest sites of the three different categories (control old-growth forest, damaged forest with the logs left, and damaged forest with the logs removed) in an old-growth subalpine coniferous forest in Mt. Yastugatake, central Japan. This forest had got a wide-range disturbance by a typhoon in 1959. We surveyed totally 95 logs in 9 sites (5, 2, and 2 sites for the forest categories held in above, respectively). Fungal communities within CWDs were documented using Illumina sequencing. Seedling and epiphytic bryophyte communities were recorded and were analyzed with CWD properties such as wood decay type (white rot, brown rot, and soft rot), pH, moisture, and bryophyte coverage. Illumina sequencing did not show obvious difference in fungal communities among the forest categories. Also, frequencies of the occurrence of wood decay type were not significantly different among the categories. None of these variables had significant association with spruce seedling density. However, experience of the disturbance certainly reduced current spruce seedling density. These results suggested that the forest disturbance do have a long lasting effect on spruces seedling regeneration on CWD, but the effect might not be attributable to their impacts on CWD fungal community and wood decay. 1)Vogel S, Alvarez B, Bassler C Müller J, Thorn S (2017) The Red-belted Bracket (Fomitopsis pinicola) colonizes spruce trees early after bark beetle attack and persists. Fungal Ecology 27:282-288. 2)Bače R, Svoboda M, Pouska V, Janda P, Červenka, J (2012) Natural regeneration in Central-European subalpine spruce forests: Which logs are suitable for seedling recruitment? Forest Ecology and Management 266:254-262.peerReviewe