Lignin-based barrier restricts pathogens to the infection site and confers resistance in plants.

Pathogenic bacteria invade plant tissues and proliferate in the extracellular space. Plants have evolved the immune system to recognize and limit the growth of pathogens. Despite substantial progress in the study of plant immunity, the mechanism by which plants limit pathogen growth remains unclear. Here, we show that lignin accumulates in Arabidopsis leaves in response to incompatible interactions with bacterial pathogens in a manner dependent on Casparian strip membrane domain protein (CASP)-like proteins (CASPLs). CASPs are known to be the organizers of the lignin-based Casparian strip, which functions as a diffusion barrier in roots. The spread of invading avirulent pathogens is prevented by spatial restriction, which is disturbed by defects in lignin deposition. Moreover, the motility of pathogenic bacteria is negatively affected by lignin accumulation. These results suggest that the lignin-deposited structure functions as a physical barrier similar to the Casparian strip, trapping pathogens and thereby terminating their growth.

Morphological and Optical Engineering for High-Performance Polymer Solar Cells.

We demonstrate morphological and optical engineering by using processing additives and optical spacers for polymer solar cells. Among various processing additives, introduction of diphenyl ether (DPE) into the active layer results in the smoothest surface roughness with uniform and well-distributed donor/acceptor domains, and the device with DPE shows the highest device efficiency of 10.22% due to enhanced charge collection efficiency and minimized recombination loss. Additional ZnO optical spacers on the active layer controls the distribution of the electric field in the whole device and enhances the light absorption within the active layer, thereby improving device efficiency up to 10.81%.

The Rab GTPase RabG3b positively regulates autophagy and immunity-associated hypersensitive cell death in Arabidopsis.

A central component of the plant defense response to pathogens is the hypersensitive response (HR), a form of programmed cell death (PCD). Rapid and localized induction of HR PCD ensures that pathogen invasion is prevented. Autophagy has been implicated in the regulation of HR cell death, but the functional relationship between autophagy and HR PCD and the regulation of these processes during the plant immune response remain controversial. Here, we show that a small GTP-binding protein, RabG3b, plays a positive role in autophagy and promotes HR cell death in response to avirulent bacterial pathogens in Arabidopsis (Arabidopsis thaliana). Transgenic plants overexpressing a constitutively active RabG3b (RabG3bCA) displayed accelerated, unrestricted HR PCD within 1 d of infection, in contrast to the autophagy-defective atg5-1 mutant, which gradually developed chlorotic cell death through uninfected sites over several days. Microscopic analyses showed the accumulation of autophagic structures during HR cell death in RabG3bCA cells. Our results suggest that RabG3b contributes to HR cell death via the activation of autophagy, which plays a positive role in plant immunity-triggered HR PCD.

Evaluation of a transgenic poplar as a potential biomass crop for biofuel production.

A transgenic poplar, in which the RabG3bCA gene from Arabidopsis was overexpressed, was analyzed for its biomass composition and enzymatic digestibility after chemical pretreatment. In comparison with a wild-type poplar (WT), the transgenic poplar (OX8) showed 9.8% higher glucan content. The levels of other biomass components did not differ greatly between WT and OX8. When WT and OX8 samples were pretreated by sulfuric acid (1%, w/v at 190 °C), sodium hydroxide (1%, w/v at 190 °C), or ammonia (14%, w/w at 80 °C), the washed pretreated solids of OX8 exhibited a higher enzymatic digestibility than those of WT in each chemical pretreatment. The sodium hydroxide pretreatment was the most effective among the three pretreatment processes, showing 58.7% and 69.4% of theoretical glucose yield from the saccharification of pretreated OX8 and WT, respectively. The transgenic poplar, growing faster and taller, was found to contain more glucan and have a higher enzymatic digestibility than WT.

Overexpression of constitutively active Arabidopsis RabG3b promotes xylem development in transgenic poplars.

An Arabidopsis small GTPase, RabG3b, was previously characterized as a component of autophagy and as a positive regulator for xylem development in Arabidopsis. In this work, we assessed whether RabG3b modulates xylem-associated traits in poplar in a similar way as in Arabidopsis. We generated transgenic poplars (Populus alba × Populus tremula var. glandulosa) overexpressing a constitutively active form of RabG3b (RabG3bCA) and performed a range of morphological, histochemical and molecular analyses to examine xylogenesis. RabG3bCA transgenic poplars showed increased stem growth due to enhanced xylem development. Autophagic structures were observed in differentiating xyelm cells undergoing programmed cell death (PCD) in wild-type poplar, and were more abundant in RabG3bCA transgenic poplar plants and cultured cells. Xylogenic activation was also accompanied by the expression of secondary wall-, PCD- and autophagy-related genes. Collectively, our results suggest that Arabidopsis RabG3b functions to regulate xylem growth through the activation of autophagy during wood formation in Populus, as does the same in Arabidopsis.

Role of Arabidopsis RabG3b and autophagy in tracheary element differentiation.

The vascular system of plants consists of two conducting tissues, xylem and phloem, which differentiate from procambium cells. Xylem serves as a transporting system for water and signaling molecules and is formed by sequential developmental processes, including cell division/expansion, secondary cell wall deposition, vacuole collapse and programmed cell death (PCD). PCD during xylem differentiation is accomplished by degradation of cytoplasmic constituents, and it is required for the formation of hollow vessels, known as tracheary elements (TEs). Our recent study revealed that the small GTPase RabG3b acts as a regulator of TE differentiation through its autophagic activation. By using an Arabidopsis in vitro cell culture system, we showed that autophagy is activated during TE differentiation. Overexpression of a constitutively active RabG3b (RabG3bCA) significantly enhances both autophagy and TE differentiation, which are consistently suppressed in transgenic plants overexpressing a dominant negative form (RabG3bDN) or RabG3b RNAi (RabG3bRNAi), a brassinosteroid-insensitive mutant bri1-301 and an autophagy mutant atg5-1. On the basis of our results, we propose that RabG3b functions as a component of autophagy and regulates TE differentiation by activating the process of PCD.

The Rab GTPase RabG3b functions in autophagy and contributes to tracheary element differentiation in Arabidopsis.

The tracheary elements (TEs) of the xylem serve as the water-conducting vessels of the plant vascular system. To achieve this, TEs undergo secondary cell wall thickening and cell death, during which the cell contents are completely removed. Cell death of TEs is a typical example of developmental programmed cell death that has been suggested to be autophagic. However, little evidence of autophagy in TE differentiation has been provided. The present study demonstrates that the small GTP binding protein RabG3b plays a role in TE differentiation through its function in autophagy. Differentiating wild type TE cells were found to undergo autophagy in an Arabidopsis culture system. Both autophagy and TE formation were significantly stimulated by overexpression of a constitutively active mutant (RabG3bCA), and were inhibited in transgenic plants overexpressing a dominant negative mutant (RabG3bDN) or RabG3b RNAi (RabG3bRNAi), a brassinosteroid insensitive mutant bri1-301, and an autophagy mutant atg5-1. Taken together, our results suggest that autophagy occurs during TE differentiation, and that RabG3b, as a component of autophagy, regulates TE differentiation.

Two Arabidopsis 3-ketoacyl CoA synthase genes, KCS20 and KCS2/DAISY, are functionally redundant in cuticular wax and root suberin biosynthesis, but differentially controlled by osmotic stress.

Very-long-chain fatty acids (VLCFAs) are essential precursors of cuticular waxes and aliphatic suberins in roots. The first committed step in VLCFA biosynthesis is condensation of C(2) units to an acyl CoA by 3-ketoacyl CoA synthase (KCS). In this study, two KCS genes, KCS20 and KCS2/DAISY, that showed higher expression in stem epidermal peels than in stems were isolated. The relative expression of KCS20 and KCS2/DAISY transcripts was compared among various Arabidopsis organs or tissues and under various stress conditions, including osmotic stress. Although the cuticular waxes were not significantly altered in the kcs20 and kcs2/daisy-1 single mutants, the kcs20 kcs2/daisy-1 double mutant had a glossy green appearance due to a significant reduction of the amount of epicuticular wax crystals on the stems and siliques. Complete loss of KCS20 and KCS2/DAISY decreased the total wax content in stems and leaves by 20% and 15%, respectively, and an increase of 10-34% was observed in transgenic leaves that over-expressed KCS20 or KCS2/DAISY. The stem wax phenotype of the double mutant was rescued by expression of KSC20. In addition, the kcs20 kcs2/daisy-1 roots exhibited growth retardation and abnormal lamellation of the suberin layer in the endodermis. When compared with the single mutants, the roots of kcs20 kcs2/daisy-1 double mutantss exhibited significant reduction of C(22) and C(24) VLCFA derivatives but accumulation of C(20) VLCFA derivatives in aliphatic suberin. Taken together, these findings indicate that KCS20 and KCS2/DAISY are functionally redundant in the two-carbon elongation to C(22) VLCFA that is required for cuticular wax and root suberin biosynthesis. However, their expression is differentially controlled under osmotic stress conditions.

Disruption of glycosylphosphatidylinositol-anchored lipid transfer protein gene altered cuticular lipid composition, increased plastoglobules, and enhanced susceptibility to infection by the fungal pathogen Alternaria brassicicola.

All aerial parts of vascular plants are covered with cuticular waxes, which are synthesized by extensive export of intracellular lipids from epidermal cells to the surface. Although it has been suggested that plant lipid transfer proteins (LTPs) are involved in cuticular lipid transport, the in planta evidence is still not clear. In this study, a glycosylphosphatidylinositol-anchored LTP (LTPG1) showing higher expression in epidermal peels of stems than in stems was identified from an Arabidopsis (Arabidopsis thaliana) genome-wide microarray analysis. The expression of LTPG1 was observed in various tissues, including the epidermis, stem cortex, vascular bundles, mesophyll cells, root tips, pollen, and early-developing seeds. LTPG1 was found to be localized in the plasma membrane. Disruption of the LTPG1 gene caused alterations of cuticular lipid composition, but no significant changes on total wax and cutin monomer loads were seen. The largest reduction (10 mass %) in the ltpg1 mutant was observed in the C29 alkane, which is the major component of cuticular waxes in the stems and siliques. The reduced content was overcome by increases of the C29 secondary alcohols and C29 ketone wax loads. The ultrastructure analysis of ltpg1 showed a more diffuse cuticular layer structure, protrusions of the cytoplasm into the vacuole in the epidermis, and an increase of plastoglobules in the stem cortex and leaf mesophyll cells. Furthermore, the ltpg1 mutant was more susceptible to infection by the fungus Alternaria brassicicola than the wild type. Taken together, these results indicated that LTPG1 contributed either directly or indirectly to cuticular lipid accumulation.

Oxaliplatin/5-FU without leucovorin chemotherapy in metastatic colorectal cancer.

PURPOSE: Fluorouracil (5-FU) and leucovorin combination therapy have shown synergistic or additive effect against advanced colorectal cancer, but the frequency of mucositis and diarrhea is increased. Most previous studies have used high dose leucovorin (300 approximately 500 mg/m(2)). However, some studies of oxaliplatin and 5-FU with low-dose or high-dose leucovorin in Korea have shown similar response rates. Therefore, we studied the necessity of leucovorin and evaluated the objective tumor response rates and toxicities of a regimen of oxaliplatin and 5-FU without leucovorin every 2 weeks in metastatic colorectal cancer patients. MATERIALS AND METHODS: Twenty-four patients with metastatic colorectal cancer were enrolled between January 2002 and March 2003. Patients received 85 mg/m(2) of oxaliplatin on day 1, a bolus 5-FU 400 mg/m(2) on day 1 and a continuous 5-FU infusion at 600 mg/m(2)/ 22 hours days 1 and 2, every 2 weeks. RESULTS: Of the 24 patients treated, 17 patients received previous 5FU with leucovorin and/or other chemotherapy. Three patients could not be evaluated. Five partial responses were observed with overall response rate of 21% (n=24). Of the previous chemotherapy group (n=17), 4 partial responses were observed with response rate of 24%. Median overall survival was 18 months (range 4 approximately 32 months) and median progression free survival was 4 months (range 2 approximately 6 months). This regimen was well tolerated and only 1 grade 3 anemia was observed. CONCLUSION: Oxaliplatin/5-FU combination therapy without leucovorin achieved a relatively high response rate even in patients resistant to the previous 5-FU chemotherapy, and toxicity was minimal.

Expression of caspase-3 and c-myc in non-small cell lung cancer.

PURPOSE: Caspase-3 is a cysteine protease that plays an important role in the process of apoptotic cell death, but little has been studied clinically on caspase-3 in lung cancer. Increased c-myc expression can result in mitosis or apoptosis, and its contribution to the pathogenesis and prognosis of lung cancer has gained interest. In the present study, the expressions of caspase-3 and c-myc, along with their possible correlations with prognostic variables, were analyzed in resected non-small cell lung carcinomas (NSCLC). MATERIALS AND METHODS: Archival tumor tissues from 147 previously untreated NSCLC patients were examined by immunohistochemistry for the expressions of caspase-3 and c-myc proteins. Clinical information was obtained through the computerized retrospective database from the tumor registry. RESULTS: The expressions of caspase-3 and c-myc were detected in 60 (88/147) and 16% (24/147) of tumors, respectively. No association was found between caspase-3 and c-myc expressions. A multivariate analysis demonstrated the N status and pathologic stage to be significantly correlated with poor survival (p-value=.018 and .002, respectively), but positive expression of caspase-3 was associated with a good prognosis (p=.03). CONCLUSION: Our data suggest the involvement of caspase-3 in the tumorigenesis of NSCLC. It is also noteworthy that caspase-3 expression might be a favorable prognostic indicator in these tumors.

Expression of c-kit and p53 in non-small cell lung cancers.

PURPOSE: Increasing experimental evidence indicates that abnormal expression of c-kit may be implicated in the pathogenesis of a variety of solid tumors. It has been reported that over 70% of small cell lung cancer (SCLC) contain the c-kit receptor. In the present study, a c-kit analysis has been extended to non-small cell lung cancer (NSCLC). The expressions of p53, vascular endothelial growth factor (VEGF) and cd34, in addition to c-kit, were evaluated to investigate the correlations between these proteins and to determine their potential relationships with the clinicopathological data. MATERIALS AND METHODS: Paraffin-embedded tumor sections, obtained from 147 patients with NSCLC, were immunohistochemically investigated using anti-c-kit, anti-p53, anti-VEGF and anti-cd34 antibodies. RESULTS: c-kit was expressed in 40 (27%) of the tumors examined: 27% of the adenocarcinomas, 27% of the squamous cell carcinomas and 29% of the undifferentiated carcinomas. p53 and VEG F immunoreactivities were present in 107 (73%) and 110 (75%) carcinomas, respectively. Anti-cd34 was negative in all samples. No associations were established among these proteins. The c-kit, however, showed a strong correlation with the T factor: T1 (n=11), 0%; T2 (n=49), 16% and T3 (n=87), 37% (p=.006). CONCLUSION: It is suggested that in NSCLC c-kit is expressed relatively frequently and may become a therapeutic target for the patients with inoperable or recurrent c-kit positive tumors. The alterations in p53 probably constitute an early event, whereas the activated c-kit may contribute to tumor progression.

Effect of Vinorelbine, Ifosfamide and Cisplatin Combination Chemotherapy in Stage III-IV Non-Small-Cell Lung Cancer.

PURPOSE: To evaluate the response rates, toxicitiesy, and survival rates, to vinorelbine (Navelbine(R)), cisplatin and ifosfamide combination chemotherapy, of the patients with inoperable NSCLC (stage III and IV), who received vinorelbine (Navelbine(R)), cisplatin, ifosfamide combinationthe mentioned chemotherapy every 4 weeks. MATERIALS AND METHODS: This study included 26 patients with inoperable NSCLC (stage III and IV), who attended St. Vincent's Hospital Bbetween April 1999 and December 2001, 26 patients were included at St.Vincent's Hospital. The chemotherapy regimen consisted of vinorelbine (25 mg/m2 on days 1 and 8), ifosfamide (1,500 mg/m2 on days 1- and 2 with mesna), and cisplatin (30 mg/m2 on days 1- to 3). The cycles were administered every 4 weeks. A 25% reduction in the doses reduction was applied into subsequent courses if there werewas grade 3~4 neutropenia. RESULTS: The median age was 63 (range, 44~73) years and the male : to female ratio was 19 : 7. One patient had stage IIIa, 6 had stage IIIb and 19 had stage IV. Twenty two patients had an ECOG performance status of 0 or 1, andwith 4 hadhave one of 2. Eighteen of the patients had adenocarcinoma, 7 had squamous cell carcinomas, and 1 had an undifferentiated NSCLC. Two patients were innot able to be evaluatedble due to follow-up loss. Among Of the 24 patients able to be evaluatedble patients, 1 patient had a complete response and 9 patients hada partial responses, and thewith an overall response rate wasof 41.7%. During a total of 104 cycles, grade 3 neutropenia occurred in 29%, grade 4 neutropenia in 12%, grade 3~4 thrombocytopenia in 4%, grade 3 anemia in 11%, and grade 3~4 mucositis in 2%. The mean time to progression was 6.4 months (range 1~13) and the median overall survival was 10 months (range 1.5~32). CONCLUSION: The combination of vinorelbine, ifosfamide and cisplatin, in the dose and schedule employed in this study, shows an response rate of 41.7%, but, because grade 3- or 4 neutropenia occurred in 41%, a careful investigation is needed.