Five hepatopancreatic and one epidermal chitinases from a pandalid shrimp (Pandalopsis japonica): cloning and effects of eyestalk ablation on gene expression

Six cDNAs encoding chitinase proteins in Pandalopsis japonica were isolated by using polymerase chain reaction (PCR) cloning methods and bioinformatic analysis of expressed sequence tags (ESTs). The cDNAs, designated Pj-Cht1, 2, 3A, 3B, 3C, and 4, encoded proteins ranging from 388 to 607 amino acid residues in length (43.61-67.62 kDa) and displayed a common structural organization: an N-terminal catalytic domain, a Thr/Pro-rich linker region, and either 0 (Pj-Cht2, 3A), 1 (Pj-Cht1, 3B, and 3C), or 2 (Pj-Cht4) C-terminal chitin-binding domain(s) (CBD). Pj-Cht1 and 2 lacked the 5′ end of the open reading frame (ORF); the other Pj-Chts contained the complete ORF. All known decapod crustacean chitinases were segregated into at least four groups based on phylogenetic analysis and domain organization. Group 1 chitinases, represented by Pj-Cht1, were most closely related to insect group I chitinases and may function in the digestion of the peritrophic membrane. Group 2 chitinases including Pj-Cht2 show different domain organizations and pI value from other chitinases and appear to function in degradation of the old exoskeleton during the premolt period. Group 3 chitinases, represented by Pj-Cht3A, 3B, and 3C, may function in digestion of chitin-containing food and defense against pathogens. Group 4 chitinases, represented by Pj-Cht4, have two CBDs and their functions are unknown. Five Pj-Chts (Pj-Cht1, 3A, 3B, 3C, and 4) are expressed in the hepatopancreas and intestine, whereas Pj-Cht2 is expressed in epidermis and SG/XO complex suggesting crustacean chitinases can be classified into two groups (hepatopancreatic and epidermal) based on the expression profile. Eyestalk ablation (ESA) down-regulated the hepatopancreatic chitinase expression (Pj-Cht1, 3A, and 3C); Pj-Cht3B expression was not significantly affected by ESA. By contrast, mRNA levels of Pj-Cht2 were significantly upregulated in 7 days post-ESA. Pj-Cht4 mRNA levels were too low for measurement with quantitative polymerase chain reaction. ESA had no significant effect on chitinase expression in the intestine. These data indicate that Pj-Cht1, 3A, 3B, 3C, and 4 are hepatopancreatic chitinases that may function in the digestion of ingested chitin and the modification of peritrophic membrane in the intestine. By contrast, epidermal chitinase, Pj-Cht2 may play a role in chitin metabolism during molt cycle as shown in other crustacean group 2 chitinases.     

Functional genomic approaches using the nematode Caenorhabditis elegans as a model system

Since the completion of the genome project of the nematode C. elegans in 1998, functional genomic approaches have been applied to elucidate the gene and protein networks in this model organism. The recent completion of the whole genome of C. briggsae, a close sister species of C. elegans, now makes it possible to employ the comparative genomic approaches for identifying regulatory mechanisms that are conserved in these species and to make more precise annotation of the predicted genes. RNA interference (RNAi) screenings in C. elegans have been performed to screen the whole genome for the genes whose mutations give rise to specific phenotypes of interest. RNAi screens can also be used to identify genes that act genetically together with a gene of interest. Microarray experiments have been very useful in identifying genes that exhibit co-regulated expression profiles in given genetic or environmental conditions. Proteomic approaches also can be applied to the nematode, just as in other species whose genomes are known. With all these functional genomic tools, genetics will still remain an important tool for gene function studies in the post genome era. New breakthroughs in C. elegans biology, such as establishing a feasible gene knockout method, immortalized cell lines, or identifying viruses that can be used as vectors for introducing exogenous gene constructs into the worms, will augment the usage of this small organism for genome-wide biology.     

Mathematical model for simultaneous nitrification and denitrification (SND) in membrane bioreactor (MBR) under Low Dissolved Oxygen (DO) concentrations

Activated Sludge Model no. 1 (ASM1) was modified and applied to Simultaneous Nitrification and Denitrification (SND) in oxygen-limited MBR. In order to calibrate the model correctly, the parametric sensitivity was performed using AQUASIM 2.0 to find the most important coefficients. The most sensitive coefficients in the model of oxygen-limited MBR were related to the growth of heterotrophic biomass. While the total autotrophic biomass concentration (X_BA) was decreased by decreasing DO concentration, there was an increase in the nitrite-oxidizing biomass concentration by a small amount. This model also showed that over 97% of permeate Soluble Chemical Oxygen Demand (S_COD) was the Soluble Inert (S_I). The model showed the change in the ammonia-oxidizing and nitrite-oxidizing biomass was decreased by decreasing DO concentration. However, there was an increase in the nitrite-oxidizing biomass concentration by a small amount due to the biomass retained in the bioreactor with membrane. It is contradictory to the reported observations for conventional activated sludge process.     

Polo-like kinase 1 (Plk1) is expressed by cutaneous T-cell lymphomas (CTCLs), and its downregulation promotes cell cycle arrest and apoptosis

Polo-like kinases are serine/threonine kinases crucial for mitosis and DNA integrity. Plk1, the most well studied member of this family, is upregulated in several cancers, as well as in dividing cells with peak expression during G2/M phase. Recently, employing lesional skin from patients with cutaneous T-cell lymphoma (CTCL), we showed that Plk1 was increased mainly in advanced lesions. In this study, employing western blot and quantitative RT-PCR analyses, we demonstrated that Plk1 was overexpressed in multiple CTCL cell lines (HH, Hut78, MyLa, SeAx and SZ4). Further, a genetic knockdown (by short hairpin RNA) or enzyme activity inhibition (via a small molecule inhibitor, GW843682X) was found to result in a decrease in cell growth, viability and proliferation. Plk1 inhibition in CTCL cells also resulted in: (1) increased G2/M phase cell cycle arrest, (2) alteration in key mitotic proteins, (3) apoptosis and (4) multiple mitotic errors. Given our findings, clinical trials of Plk1 inhibitors in CTCL may be a promising area for further translational investigation. We speculate that overexpression of Plk1 may prove to be relevant to the progression and prognosis of CTCL through its direct impact on the regulation of tumor cell proliferation and indirect influence on the acquisition of somatic mutations by proliferating tumor cells.     

Expression of a bifunctional fusion of the Escherichia coli genes for trehalose-6-phosphate synthase and trehalose-6-phosphate phosphatase in transgenic rice plants increases trehalose accumulation and abiotic stress tolerance without stunting growth

Trehalose plays an important role in stress tolerance in plants. Trehalose-producing, transgenic rice (Oryza sativa) plants were generated by the introduction of a gene encoding a bifunctional fusion (TPSP) of the trehalose-6-phosphate (T-6-P) synthase (TPS) and T-6-P phosphatase (TPP) of Escherichia coli, under the control of the maize (Zea mays) ubiquitin promoter (Ubi1). The high catalytic efficiency (Seo et al., 2000) of the fusion enzyme and the single-gene engineering strategy make this an attractive candidate for high-level production of trehalose; it has the added advantage of reducing the accumulation of potentially deleterious T-6-P. The trehalose levels in leaf and seed extracts from Ubi1::TPSP plants were increased up to 1.076 mg g fresh weight(-1). This level was 200-fold higher than that of transgenic tobacco (Nicotiana tabacum) plants transformed independently with either TPS or TPP expression cassettes. The carbohydrate profiles were significantly altered in the seeds, but not in the leaves, of Ubi1::TPSP plants. It has been reported that transgenic plants with E. coli TPS and/or TPP were severely stunted and root morphology was altered. Interestingly, our Ubi1::TPSP plants showed no growth inhibition or visible phenotypic alterations despite the high-level production of trehalose. Moreover, trehalose accumulation in Ubi1::TPSP plants resulted in increased tolerance to drought, salt, and cold, as shown by chlorophyll fluorescence and growth inhibition analyses. Thus, our results suggest that trehalose acts as a global protectant against abiotic stress, and that rice is more tolerant to trehalose synthesis than dicots.     

Adenomyomatous hamartoma of lung mimicking benign mucinous tumor in fine needle aspiration biopsy: a case report

BACKGROUND: Typical cytologic features of pulmonary hamartoma (PH) are usually smears of hyaline cartilage, fibrous tissue, smooth muscle, adipocytic components and respiratory epithelium. Cytologic features of adenomyomatous hamartoma, a special variant of PH, are not documented in the literature and are confused with epithelial neoplasm in the case of sparse stromal cellularity. CASE: A 59-year-old man presented with a solitary pulmonary nodule by chest radiograph at his routine health examination. Fine needle aspiration biopsy (FNAB) revealed numerous mucinous epithelial cells presenting predominantly in cohesive cellular sheets that suggested benign mucinous epithelial lesion. The patient underwent surgery for the tumor, and it was histologically proven to be an adenomyomatous hamartoma. CONCLUSION: An unusual type of PH could lead to misdiagnosis by FNAB in the case of few stromal components. This case demonstrates the wide spectrum of PH in FNAB and led us to consider PH as a differential diagnosis despite lack of chondromyxoid stromal components.     

Effect of different training mode on Interleukin-6 (IL-6) and C-reactive protein (CRP) in type 2 diabetes mellitus (T2DM) patients

[Purpose]

This study was carried out to investigate the effects of different training modes on IL-6 and CRP in patients with type 2 diabetes mellitus (T2DM).

[Methods]

The subjects consisted of 16 middle-aged women with type 2 diabetes mellitus (T2DM), all of whom had no other complications. The 16 subjects were randomly assigned to two experimental groups: the circuit training group (CTG, n = 8) and aerobic training group (ATG, n = 8). Based on measured THR (target heart rate) for maximum oxygen consumption rate, the circuit training group (CTG) exercised at 60% intensity, 60 min/day, 5 sets, 3 days/week for 12 weeks. Based on measured THR (target heart rate) for maximum oxygen consumption rate, the aerobic training group (ATG) exercised at 60% intensity (which was increased gradually in weeks 4, 7, and 10) 60 min/day, 3 days/week for 12 weeks.

[Results]

The results are as follows. Significant decreases in the post training values of weight, % Fat, BMI, IL-6 and CRP (p < .05) were observed in the CTG compared to pre-training. However, there were no differences in the physical characteristic and blood inflammatory factors between the groups (ATG & CTG).

[Conclusion]

In conclusion, the results of this study suggest that circuit training (CT) may be considered as an effective training mode for helping to decrease the blood inflammatory factors (IL-6 and CRP) in patients with type 2 diabetes mellitus (T2DM).