Involvement of Protein Tyrosine Phosphatases BcPtpA and BcPtpB in Regulation of Vegetative Development,Virulence and Multi-Stress Tolerance in Botrytis cinerea

Tyrosine phosphorylation and dephosphorylation have emerged as fundamentally important mechanisms of signal transduction and regulation in eukaryotic cells, governing many processes, but little has been known about their functions in filamentous fungi. In this study, we deleted two putative protein tyrosine phosphatase (PTP) genes (BcPTPA and BcPTPB) in Botrytis cinerea, encoding the orthologs of Saccharomyces cerevisiae Ptp2 and Ptp3, respectively. Although BcPtpA and BcPtpB have opposite functions in conidiation, they are essential for sclerotial formation in B. cinerea. BcPTPA and BcPTPB deletion mutants ΔBcPtpA-10 and ΔBcPtpB-4 showed significantly increased sensitivity to osmotic and oxidative stresses, and to cell wall damaging agents. Inoculation tests showed that both mutants exhibited dramatically decreased virulence on tomato leaves, apples and grapes. In S. cerevisiae, it has been shown that Ptp2 and Ptp3 negatively regulate the high-osmolarity glycerol (HOG) pathway and the cell wall integrity (CWI) pathway. Although both BcPtpA and BcPtpB were able to inactive Hog1 and Mpk1 in S. cerevisiae, in contrast to S. cerevisiae, they positively regulate phosphorylation of BcSak1 (the homologue of Hog1) and BcBmp3 (the homologue of Mpk1) in B. cinerea under stress conditions. These results demonstrated that functions of PTPs in B. cinerea are different from those in S. cerevisiae, and BcPtpA and BcPtpB play important roles in regulation of vegetative development, virulence and in adaptation to oxidative, osmotic and cell-wall damage stresses in B. cinerea.     

Activation of AMPK by Bitter Melon Triterpenoids Involves CaMKKβ

We recently showed that bitter melon-derived triterpenoids (BMTs) activate AMPK and increase GLUT4 translocation to the plasma membrane in vitro, and improve glucose disposal in insulin resistant models in vivo. Here we interrogated the mechanism by which these novel compounds activate AMPK, a leading anti-diabetic drug target. BMTs did not activate AMPK directly in an allosteric manner as AMP or the Abbott compound (A-769662) does, nor did they activate AMPK by inhibiting cellular respiration like many commonly used anti-diabetic medications. BMTs increased AMPK activity in both L6 myotubes and LKB1-deficient HeLa cells by 20–35%. Incubation with the CaMKKβ inhibitor, STO-609, completely attenuated this effect suggesting a key role for CaMKKβ in this activation. Incubation of L6 myotubes with the calcium chelator EGTA-AM did not alter this activation suggesting that the BMT-dependent activation was Ca²⁺-independent. We therefore propose that CaMKKβ is a key upstream kinase for BMT-induced activation of AMPK.     

INTRASPECIFIC VARIATION AND THE ROLE OF SUPERPARASITISM IN DIAPAUSE INDUCTION OF TRICHOGRAMMA EVANESCENS WEST-WOOD*

Abstract The diapause of two populations of Trichogramma evanescens Westwood (T. evanescens A and T. evanescens B), collected from different Iranian insect pests, was studied. T. evunescens A in the eggs of Corcyra cephalonica was easily induced to stable diapause with constant 8°C, 15 % and fluctuating temperature (11–23) C -11C. T. evanescens B could not be induced to diapause with the same temperatural regime and host. The experiments showed that the host of maternal generations and the ability of avoiding super-parasitism may play an important role in intraspecific variation of T. evunescens in diapause induction.     

Genome‐wide scans to detect positive selection in Large White and Tongcheng pigs

Due to the direction, intensity, duration and consistency of genetic selection, especially recent artificial selection, the production performance of domestic pigs has been greatly changed. Therefore, we reasoned that there must be footprints or selection signatures that had been left during domestication. In this study, with porcine 60K BeadChip genotyping data from both commercial Large White and local Chinese Tongcheng pigs, we calculated the extended haplotype homozygosity values of the two breeds using the long‐range haplotype method to detect selection signatures. We found 34 candidate regions, including 61 known genes, from Large White pigs and 25 regions comprising 57 known genes from Tongcheng pigs. Many selection signatures were found on SSC1, SSC4, SSC7 and SSC14 regions in both populations. According to quantitative trait loci and network pathway analyses, most of the regions and genes were linked to growth, reproduction and immune responses. In addition, the average genetic differentiation coefficient F_ST was 0.254, which means that there had already been a significant differentiation between the breeds. The findings from this study can contribute to further research on molecular mechanisms of pig evolution and domestication and also provide valuable references for improvement of their breeding and cultivation.     

Interactions of chloride and amiloride with the renal Na+/H/ antiporter

Amiloride is a reversible inhibitor of the Na+/H+ antiporter which acts at the external aspect of the transport system. The kinetics of inhibition of the Na+/H+ antiporter with amiloride have been controversial, with the usual finding of simple competitive inhibition, but with other reports of mixed and noncompetitive inhibition of the transporter by amiloride. The present experiments demonstrate that the chloride content of the external transport buffer affects the kinetics of amiloride inhibition. Either simple competitive or mixed inhibition by amiloride was observed in the same vesicle preparations depending on the presence of chloride or gluconate in the buffer. The effect of chloride on the inhibitory effect of amiloride was dependent on the concentration of chloride and amiloride. Similar effects were observed with more potent analogues of amiloride. These findings suggest that the external aspect of the antiporter has a site or sites at which the inhibitory effects of amiloride on the Na+/H+ antiporter can be modified by chloride, even though chloride has only slight effects on the kinetics of the Na+/H+ antiporter in the absence of amiloride.     

Clearance of rat C-reactive protein in vivo and by perfused liver.

The clearance in vivo of rat C-reactive protein (CRP) was studied: (i) in the whole animal and (ii) by using a rat liver perfusion system. Rat CRP is a glycosylated serum protein containing a complex-type biantennary carbohydrate structure on each of its five subunits. The half-life of rat asialo CRP was approximately 5 min. More than 75% of the radioactivity associated with rat asialo CRP and asialo alpha 1-acid glycoprotein (AGP) was recovered in the liver. A small amount of radioactivity (0.8%) associated with rat CRP and rat asialo CRP was found in the lungs. Competitive inhibition of the clearance of 125I-labelled rat asialo CRP from the circulation by asialo AGP was dose dependent, and resulted in a corresponding decrease in the recovery of radioactivity associated with rat asialo CRP in the liver. This indicated that asialo AGP and rat asialo CRP were cleared by the hepatic asialoglycoprotein receptor. This observation was confirmed when the clearance of rat asialo CRP was studied using a rat liver perfusion system. Using this system, the clearance of rat asialo CRP and asialo AGP from the perfusate was inhibited by N-acetylgalactosamine, but not by phosphorylcholine, a ligand through which most of the CRP reactions are mediated. This study provides an example of a circulating serum glycoprotein containing a biantennary carbohydrate structure that is cleared by the asialoglycoprotein receptor.     

BJ-HCC-20, a potential novel cancer-testis antigen.

In an effort to identify novel Cancer-Testis genes, we analyzed the sequence in the q26-28 region of human X chromosome by several on-line tools. The candidate sequences were then confirmed by experiments. We have obtained a novel Cancer-Testis gene, BJ-HCC-20. In vivo, it was found to have two isoforms. In samples of liver, colon, gastric and lung cancer tested, the expression frequency of BJ-HCC-20 is 25%, 17%, 21% and 15%, respectively. Full-length cDNAs of both BJ-HCC-20 isoforms were isolated and their gene structures and promoter regions were characterized. BJ-HCC-20 might have implications in theoretical and practical tumor biology.