Molecular cloning and sequence analysis of methionine adenosyltransferase from the economic seaweed Undaria pinnatifida

The methionine adenosyltransferase gene (MAT) had been isolated from an economic seaweed Undaria pinnatifida by PCR using degenerate primers. The cDNA was 1,491 bp in length with an open reading frame of 1,194 nucleotides, encoding a deduced protein of 397 amino acids. The protein had a predicted molecular weight of 43.2 kDa, and the isoelectric point was 5.244. The sequence contains a 92 bp 5′-untranslated region (UTR) and a 205 bp 3′-UTR. The methionine adenosyltransferase (MAT) sequence of U. pinnatifida (UpMAT) shared 68–92 % identities with the previous published MAT sequences of other species. Phylogenetic analysis indicated that the phylogenetic relationship of UpMAT with some other seaweeds was closer than with those of higher plants. Under different stress conditions, the relative mRNA expression levels of the MAT of U. pinnatifida (UpMAT) were measured by real-time quantitative PCR, and the results demonstrated that the UpMAT might help to protect the alga against various abiotic stresses.     

The Arabidopsis Anaphase-Promoting Complex/Cyclosome Subunit 1 is Critical for Both Female Gametogenesis and Embryogenesis

Anaphase-promoting complex/cyclosome (APC/C), a multisubunit E3 ligase, plays a critical role in cell cycle control, but the functional characterization of each subunit has not yet been completed. To investigate the function of APC1 in Arabidopsis, we analyzed four mutant alleles of APC1, and found that mutation in APC1 resulted in significantly reduced plant fertility, accumulation of cyclin B, and disrupted auxin distribution in embryos. The three mutant alleles apc1-1, apc1-2 and apc1-3 shared variable defects in female gametogenesis including degradation, abnormal nuclear number, and disrupted polarity of nuclei in the embryo sac as well as in embryogenesis, in which embryos were arrested at multiple stages. All of these defects are similar to those previously identified in apc4. The mutant apc1-4, in which the T-DNA was inserted after the transmembrane domain at the C-terminus, showed much more severe phenotypes; that is, most of the ovules were arrested at the one-nucleate female gametophyte stage (stage FG1). In the apc1 apc4 double mutants, the fertility was further reduced by one-third in apc1-1/+ apc4-1/+, and in some cases no ovules even survived in siliques of apc1-4/+ apc4-1/+. Our data thus suggest that APC1, an essential component of APC/C, plays a synergistic role with APC4 both in female gametogenesis and in embryogenesis.     

A novel strategy to inhibit the reproduction and translation of hepatitis C virus

Hepatitis C virus (HCV), a positive single-stranded RNA virus, is a major cause of liver disease in humans. Herein we report a novel strategy to inhibit the reproduction and translation of HCV using a short RNA, named an Additional RNA, to activate the endonuclease activity of Argonaute 2 (Ago2). In the presence of the Additional RNA, the HCV genome RNA has the requisite 12 nucleotides of base-pairing with microRNA-122. This activates the endonuclease activity of Ago2, resulting in cleavage and release of the HCV genome RNA from Ago2 and microRNA-122. The free HCV genome RNA would be susceptible to intracellular degradation, effectively inhibiting its reproduction and translation. This study presents a new method to inhibit HCV that may hold great potential for HCV treatment in the future.     

Pulmonary Tuberculosis Incidence and Risk Factors in Rural Areas of China: A Cohort Study

The incidence of tuberculosis (TB) and its risk factors in China remains unclear. This study examined TB incidence and relative risk factors in rural areas of China. Participants (n = 177,529) were recruited in Xiangtan County (in the central area of China) and in Danyang County (in the eastern area of China) in 2009 and a followed-up study was conducted for one year. The incidence density of pulmonary TB and smear-positive TB were 91.6 (95% CI: 78.7, 106.0) per 100,000 person-year and 36.7 (95% CI: 33.1, 52.4) per 100,000 person-year respectively in Xiangtan, and 47.3 (95% CI: 38.2, 57.5) per 100,000 person-year and 22.7 (95% CI: 16.5, 30.8) per 100,000 person-year in Danyang. The medical history of TB was associated with TB, with the relative risk (RR) of 7.00 (95% CI: 2.76, 17.18) in Xiangtan and that of 31.08 (95% CI: 13.22, 73.10) in Danyang. The association between TB and per capita living space over median was found in Xiangtan, with the RR of 1.86 (95% CI: 1.15, 3.01). No association was found between TB and the insurance status, the contact history with TB, the history of diabetes, smoking, or per capita annual income. The host genetic susceptibility, and social factors such as education and income could be considered in future studies.     

Large-Scale Screens of miRNA-mRNA Interactions Unveiled That the 3′UTR of a Gene Is Targeted by Multiple miRNAs

Animal microRNA (miRNA) target prediction is still a challenge, although many prediction programs have been exploited. MiRNAs exert their function through partially binding the messenger RNAs (mRNAs; likely at 3′ untranslated regions [3′UTRs]), which makes it possible to detect the miRNA-mRNA interactions in vitro by co-transfection of miRNA and a luciferase reporter gene containing the target mRNA fragment into mammalian cells under a dual-luciferase assay system. Here, we constructed a human miRNA expression library and used a dual-luciferase assay system to perform large-scale screens of interactions between miRNAs and the 3′UTRs of seven genes, which included more than 3,000 interactions with triplicate experiments for each interaction. The screening results showed that the 3′UTR of one gene can be targeted by multiple miRNAs. Among the prediction algorithms, a Bayesian phylogenetic miRNA target identification algorithm and a support vector machine (SVM) presented a relatively better performance (27% for EIMMo and 24.7% for miRDB) against the average precision (17.3%) of the nine prediction programs used here. Additionally, we noticed that a relatively high conservation level was shown at the miRNA 3′ end targeted regions, as well as the 5′ end (seed region) binding sites.     

Cancer Incidence and Mortality in Patients with Type 2 Diabetes Treated with Human Insulin: A Cohort Study in Shanghai

Aim

The aim was to investigate the association between human insulin and cancer incidence and mortality in Chinese patients with type 2 diabetes.

Methods

We recruited 8,774 insulin-naïve diabetes patients from the Shanghai Diabetes Registry (SDR). The follow-up rate was 85.4%. All subjects were divided into the insulin use cohort (n = 3,639) and the non-insulin use cohort (n = 5,135). The primary outcome was the first diagnosis of any cancer. The secondary outcome was all-cause mortality. Cox proportional hazards model was used to estimate the relative risk (RR) of cancer and mortality.

Results

We observed 98 cancer events in the insulin use cohort and 170 in the non-insulin use cohort. Cancer incidence rates were 78.6 and 74.3 per 10,000 patients per year in the insulin users and the non-insulin users, respectively. No significant difference in cancer risk was observed between the two cohorts (adjusted RR = 1.20, 95% CI 0.89–1.62, P = 0.228). Regarding site-specific cancers, only the risk of liver cancer was significantly higher in the insulin users compared to that in the non-insulin users (adjusted RR = 2.84, 95% CI 1.12–7.17, P = 0.028). The risks of overall mortality (adjusted RR = 1.89, 95% CI 1.47–2.43, P<0.0001) and death from cancer (adjusted RR = 2.16, 95% CI 1.39–3.35, P = 0.001) were all significantly higher in the insulin users than in the non-insulin users.

Conclusion

There was no excess risk of overall cancer in patients with type 2 diabetes who were treated with human insulin. However, a significantly higher risk of liver cancer was found in these patients. Moreover, insulin users showed higher risks of overall and cancer mortality. Considering that individuals treated with insulin were more likely to be advanced diabetic patients, caution should be used in interpreting these results.     

Clinical significance of serum transforming growth factor-β1 in lung cancer

Background: Transforming growth factor-β1 (TGF-β1) plays a critical role in human cancer development. Present study aimed to explore the clinical significance of serum TGF-β1 levels in patients with lung cancer and analyze the relationship between TGF-β1 and existing tumor markers for lung cancer. Methods: Serum was collected from 118 patients with lung cancer and 40 healthy volunteers. Serum TGF-β1 levels were measured by enzyme-linked immunosorbent assay (ELISA), and the association with various clinical characteristics was analyzed. The diagnostic value of TGF-β1 was assessed alone and in combination with existing tumor markers for lung cancer. Results: Serum TGF-β1 levels were significantly higher in patients with lung cancer compared to healthy volunteers [0.6 × 10⁵ (0.4 × 10⁵, 0.9 × 10⁵) pg/ml vs 0.5 × 10⁵ (0.3 × 10⁵, 0.7 × 10⁵) pg/ml, P = 0.040]. Although there was a positive correlation between serum TGF-β1 levels and advanced stages, the significant difference was not found between early stages and advanced stages (P = 0.116). The ability of serum TGF-β1 to discriminate lung cancer at a cutoff value of 79,168 pg/ml exhibited sensitivity of 30.6% and specificity of 97.5%. Serum TGF-β1 levels were correlated to cytokeratin fragment 21-1 (CYFRA21-1; R = 0.308, P = 0.020) and neuron-specific enolase (NSE; R = 0.558, P = 0.003). The diagnostic accuracy rates for the existing lung-tumor markers, as SCC, CYFRA21-1, and NSE, were increased from 20.0%, 34.6%, and 45.9% to 48.9%, 51.7%, and 54.5%, respectively by the inclusion of serum TGF-β1 levels. Conclusion: Quantification of serum TGF-β1 levels by ELISA may provide a novel complementary tool for the clinical diagnosis of lung cancer.     

Increased Expression of IL-22 Is Associated with Disease Activity in Behcet’s Disease

Objective

Interleukin (IL)-22 has been reported to be involved in the development of autoimmune diseases. This study aimed to analyze the expression and potential role of IL-22 in the pathogenesis of Behcet’s disease (BD).

Methods

The levels of IL-22 in patient sera or supernatants of cultured peripheral blood mononuclear cells (PBMCs) and CD4⁺T cells were detected by enzyme-linked immunosorbent assay (ELISA). Flow cytometry was used to evaluate the frequency of IL-22–producing CD4⁺ T cells. IL-22 mRNA from erythema nodosum skin lesions was examined using real time quantitative RT-PCR.

Results

BD patients with active uveitis showed a significantly higher expression of IL-22 in the supernatants of stimulated PBMCs and CD4⁺T cells compared with BD patients without active uveitis and normal controls. An increased frequency of IL-22-producing CD4⁺T cells was also found in BD patients with active uveitis. IL-22 mRNA expression was elevated in erythema nodosum skin lesions. In BD patients, a high IL-22 level in the supernatant of stimulated PBMCs correlated with the presence of retinal vasculitis and erythema nodosum.

Conclusions

IL-22 was associated with disease activity in BD and correlated with the presence of small vessel inflammation, suggesting that it may be involved in its pathogenesis.     

Autophagy promotes ferroptosis by degradation of ferritin

Macroautophagy/autophagy is an evolutionarily conserved degradation pathway that maintains homeostasis. Ferroptosis, a novel form of regulated cell death, is characterized by a production of reactive oxygen species from accumulated iron and lipid peroxidation. However, the relationship between autophagy and ferroptosis at the genetic level remains unclear. Here, we demonstrated that autophagy contributes to ferroptosis by degradation of ferritin in fibroblasts and cancer cells. Knockout or knockdown of Atg5 (autophagy-related 5) and Atg7 limited erastin-induced ferroptosis with decreased intracellular ferrous iron levels, and lipid peroxidation. Remarkably, NCOA4 (nuclear receptor coactivator 4) was a selective cargo receptor for the selective autophagic turnover of ferritin (namely ferritinophagy) in ferroptosis. Consistently, genetic inhibition of NCOA4 inhibited ferritin degradation and suppressed ferroptosis. In contrast, overexpression of NCOA4 increased ferritin degradation and promoted ferroptosis. These findings provide novel insight into the interplay between autophagy and regulated cell death.