Auxin-inducible protein depletion system in fission yeast

Background  

Inducible inactivation of a protein is a powerful approach for analysis of its function within cells. Fission yeast is a useful model for studying the fundamental mechanisms such as chromosome maintenance and cell cycle. However, previously published strategies for protein-depletion are successful only for some proteins in some specific conditions and still do not achieve efficient depletion to cause acute phenotypes such as immediate cell cycle arrest. The aim of this work was to construct a useful and powerful protein-depletion system in Shizosaccaromyces pombe.     

Improved eicosapentaenoic acid production in Pythium splendens RBB-5 based on metabolic regulation analysis

Applied Microbiology and Biotechnology - Eicosapentaenoic acid (EPA) is an essential polyunsaturated fatty acid for human beings. At present, the production of commercially available long-chain...     

Targeted overexpression of inducible 6-phosphofructo-2-kinase in adipose tissue increases fat deposition but protects against diet-induced insulin resistance and inflammatory responses

Increasing evidence demonstrates the dissociation of fat deposition, the inflammatory response, and insulin resistance in the development of obesity-related metabolic diseases. As a regulatory enzyme of glycolysis, inducible 6-phosphofructo-2-kinase (iPFK2, encoded by PFKFB3) protects against diet-induced adipose tissue inflammatory response and systemic insulin resistance independently of adiposity. Using aP2-PFKFB3 transgenic (Tg) mice, we explored the ability of targeted adipocyte PFKFB3/iPFK2 overexpression to modulate diet-induced inflammatory responses and insulin resistance arising from fat deposition in both adipose and liver tissues. Compared with wild-type littermates (controls) on a high fat diet (HFD), Tg mice exhibited increased adiposity, decreased adipose inflammatory response, and improved insulin sensitivity. In a parallel pattern, HFD-fed Tg mice showed increased hepatic steatosis, decreased liver inflammatory response, and improved liver insulin sensitivity compared with controls. In both adipose and liver tissues, increased fat deposition was associated with lipid profile alterations characterized by an increase in palmitoleate. Additionally, plasma lipid profiles also displayed an increase in palmitoleate in HFD-Tg mice compared with controls. In cultured 3T3-L1 adipocytes, overexpression of PFKFB3/iPFK2 recapitulated metabolic and inflammatory changes observed in adipose tissue of Tg mice. Upon treatment with conditioned medium from iPFK2-overexpressing adipocytes, mouse primary hepatocytes displayed metabolic and inflammatory responses that were similar to those observed in livers of Tg mice. Together, these data demonstrate a unique role for PFKFB3/iPFK2 in adipocytes with regard to diet-induced inflammatory responses in both adipose and liver tissues.     

Novel potassium N-[(2S,3R,4R,5R)-2,3,4,5,6-pentahydroxylhex-1-yl]-L-amino acid dichloroplatinates(II) with high anti-tumor activity and low side reaction

To discover whether novel anti-tumor platinum agents are capable of selectively accumulating in tumor tissue, three novel potassium N-[(2S,3R,4R,5R)-2,3,4,5,6-pentahydroxylhex-1-yl]-L-amino acid dichloroplatinates(II) were prepared. At a dose of 1.67 μmol kg(-1) the in vivo anti-tumor potencies of two of the compounds were higher than that of oxaliplatin. The mortality analysis indicated that these compounds resulted in a 100% survival rate, whereas oxaliplatin lead to an 80% survival rate. The organ damage examination indicated that these compounds induced less damage than oxaliplatin. The platinum accumulation in the organs, blood and bone was significantly lower than that of oxaliplatin treated mice, while the platinum accumulation in the tumor tissue was significantly higher than that of the oxaliplatin treated mice.     

Inhibition of CLIC4 enhances autophagy and triggers mitochondrial and ER stress-induced apoptosis in human glioma U251 cells under starvation

CLIC4/mtCLIC, a chloride intracellular channel protein, localizes to mitochondria, endoplasmic reticulum (ER), nucleus and cytoplasm, and participates in the apoptotic response to stress. Apoptosis and autophagy, the main types of the programmed cell death, seem interconnected under certain stress conditions. However, the role of CLIC4 in autophagy regulation has yet to be determined. In this study, we demonstrate upregulation and nuclear translocation of the CLIC4 protein following starvation in U251 cells. CLIC4 siRNA transfection enhanced autophagy with increased LC3-II protein and puncta accumulation in U251 cells under starvation conditions. In that condition, the interaction of the 14-3-3 epsilon isoform with CLIC4 was abolished and resulted in Beclin 1 overactivation, which further activated autophagy. Moreover, inhibiting the expression of CLIC4 triggered both mitochondrial apoptosis involved in Bax/Bcl-2 and cytochrome c release under starvation and endoplasmic reticulum stress-induced apoptosis with CHOP and caspase-4 upregulation. These results demonstrate that CLIC4 nuclear translocation is an integral part of the cellular response to starvation. Inhibiting the expression of CLIC4 enhances autophagy and contributes to mitochondrial and ER stress-induced apoptosis under starvation.     

Stem Cell Transplantation Increases Antioxidant Effects in Diabetic Mice

Intra bone marrow-bone marrow transplantation (IBM- BMT) + thymus transplantation (TT) has been shown to reduce the incidence of graft versus host disease (GVHD) and restore donor-derived T cell function. In addition, an increase in insulin sensitivity occurred in db/db mice after IBM-BMT+TT treatment. Heme oxygenase (HO)-1 is a stress inducible enzyme which exert antioxidant, antiapoptotic, and immune-modulating properties. We examined whether IBM-BMT+TT could modulate the expression of HO-1 in the kidneys of db/db mice. Six-week-old db/db mice with blood glucose levels higher than 250 mg/dl were treated with IBM-BMT+TT. Six weeks later, the db/db mice showed decreased body weight, blood glucose levels and insulin, and increased plasma adiponectin levels. The upregulation of HO-1 was associated with significantly (p<0.05) increased levels of peNOS and pAKT, but decreased levels of iNOS in the kidneys of db/db mice. Plasma creatinine levels also decreased (p<0.05), and the expression of type IV collagen was improved. Thus IBM-BMT+TT unregulated the expression of HO-1, peNOS and pAKT, while decreasing iNOS levels in the kidney of db/db mice. This was associated with an improvement in renal function.     

Chondrocalcinosis is common in the absence of knee involvement

Introduction

We aimed to describe the distribution of radiographic chondrocalcinosis (CC) and to examine whether metacarpophalangeal joint (MCPJ) calcification and CC at other joints occurs in the absence of knee involvement.

Methods

This was a cross-sectional study embedded in the Genetics of Osteoarthritis and Lifestyle study (GOAL). All participants (n = 3,170) had radiographs of the knees, hands, and pelvis. These were scored for radiographic changes of osteoarthritis (OA), for CC at knees, hips, symphysis pubis, and wrists, and for MCPJ calcification. The prevalence of MCPJ calcification and CC overall, at each joint, and in the presence or absence of knee involvement, was calculated.

Results

The knee was the commonest site of CC, followed by wrists, hips, and symphysis pubis. CC was more likely to be bilateral at knees and wrists but unilateral at hips. MCPJ calcification was usually bilateral, and less common than CC at knees, hips, wrists, and symphysis pubis. Unlike that previously reported, CC commonly occurred without any knee involvement; 44.4% of wrist CC, 45.9% of hip CC, 45.5% of symphysis pubis CC, and 31.3% of MCPJ calcification occurred in patients without knee CC. Those with meniscal or hyaline articular cartilage CC had comparable ages (P = 0.21), and neither preferentially associated with fibrocartilage CC at distant joints.

Conclusions

CC visualized on a plain radiograph commonly occurs at other joints in the absence of radiographic knee CC. Therefore, knee radiographs alone are an insufficient screening test for CC. This has significant implications for clinical practice, for epidemiologic and genetic studies of CC, and for the definition of OA patients with coexistent crystal deposition.