A novel pepper membrane-located receptor-like protein gene CaMRP1 is required for disease susceptibility, methyl jasmonate insensitivity and salt tolerance

Plant receptor proteins are involved in the signaling networks required for defense against pathogens. The novel pepper pathogen-induced gene CaMRP1 was isolated from pepper leaves infected with Xanthomonas campestris pv. vesicatoria (Xcv). This gene is predicted to encode a membrane-located receptor-like protein that has an N-terminal signal peptide and a C-terminal transmembrane helix. A CaMRP1-GFP fusion protein localized primarily to the plasma membrane of plant cells. Strong and early induction of CaMRP1 expression occurred following exposure of pepper plants to Xcv, Colletotricum coccodes, methyl jasmonate (MeJA) and wounding stress. Virus-induced gene silencing (VIGS) of CaMRP1 in pepper conferred enhanced basal resistance to Xcv infection, accompanied by induction of genes encoding basic PR1 (CaBPR1), defensin (CaDEF1) and SAR8.2 (CaSAR82A). In contrast, CaMRP1 overexpression (OX) in transgenic Arabidopsis plants resulted in increased disease susceptibility to Hyaloperonospora parasitica infection. Arabidopsis plants overexpressing CaMRP1 exhibited insensitivity to MeJA by causing reduced expression of MeJA-responsive genes. Overexpression also resulted in tolerance to NaCl and during salt stress, the expression of several abscisic acid-responsive genes was induced. Together, these results suggest that pepper CaMRP1 may belong to a new subfamily of membrane-located receptor-like proteins that regulate disease susceptibility, MeJA-insensitivity and salt tolerance.     

水稻中受体激酶的系统树分析

植物受体激酶(RLKs)在植物细胞内的反应中发挥着重要作用。为了比较拟南芥(Arabidopsis thaliana L.)和水稻(Oryza sativa L.)中受体激酶的进化关系,作者通过对北京华大基因研究中心(BGI)的籼稻蛋白质数据库进行BLASTP搜索,找到267个受体激酶类似基因,根据它们的胞外结构域可以将这些基因分为不同的类型。与拟南芥中受体激酶的系统树比较分析表明,不同类型的受体激酶具有不同的序列保守性,说明在植物进化过程中,不同类型的受体激酶具有不同的进化关系。水稻受体激酶与拟南芥受体激酶BRI1的多序列匹配结果也表明二者可能具有不同的磷酸化位点。     

Impact of drug discovery on stem cell biology

Despite the rapid technical progress in pharmaceutical industry in the past decade, it is still a great challenge to find new drugs and the situation seems more and more serious. However, the history of pharmaceutical industry clearly indicated that the significance of drug discovery went far beyond providing new drugs. For instance, drugs or candidates could be used as selective probes to reveal novel cellular mechanisms, which is a fundamental tenet of chemical biology. More interestingly, accumulating evidence indicates that drugs and candidates can find important use in stem cell biology. Not only approved drugs but also undeveloped pharmacological agents could serve as efficient agents to regulate stem cell fate. Moreover, the target and activity knowledge accumulated during the drug discovery process will help select the stem cell fate modulators in a rational manner. As the progress in stem cell biology will bring positive influence to drug discovery, it can be expected that the current drug discovery efforts will finally bear great fruits in the future.     

Targeting cholangiocarcinoma

Cholangiocarcinoma (CCA) represents a diverse group of epithelial cancers associated with the biliary tract, and can best be stratified anatomically into intrahepatic (iCCA), perihilar (pCCA) and distal (dCCA) subsets. Molecular profiling has identified genetic aberrations associated with these anatomic subsets. For example, IDH catalytic site mutations and constitutively active FGFR2 fusion genes are predominantly identified in iCCA, whereas KRAS mutations and PRKACB fusions genes are identified in pCCA and dCCA. Clinical trials targeting these specific driver mutations are in progress. However, The Tumor Genome Atlas (TCGA) marker analysis of CCA also highlights the tremendous molecular heterogeneity of this cancer rendering comprehensive employment of targeted therapies challenging. CCA also display a rich tumor microenvironment which may be easier to target. For example, targeting cancer associated fibroblasts for apoptosis with BH3-mimetics and/or and reversing T-cell exhaustion with immune check point inhibitors may help aid in the treatment of this otherwise devastating malignancy. Combinatorial therapy attacking the tumor microenvironment plus targeted therapy may help advance treatment for CCA. This article is part of a Special Issue entitled: Cholangiocytes in Health and Disease edited by Jesus Banales, Marco Marzioni, Nicholas LaRusso and Peter Jansen.     

Gatekeeper tyrosine phosphorylation is autoinhibitory for Symbiosis Receptor Kinase

Plant receptor-like kinases (RLKs) are distinguished by having a tyrosine in the ‘gatekeeper’ position. Previously we reported Symbiosis Receptor Kinase from Arachis hypogaea (AhSYMRK) to autophosphorylate on the gatekeeper tyrosine (Y670), though this phosphorylation was not necessary for the kinase activity. Here we report that recombinant catalytic domain of AhSYMRK with a phosphomimic substitution in the gatekeeper position (Y670E) is catalytically almost inactive and is conformationally quite distinct from the corresponding native enzyme. Additionally, we show that gatekeeper-phosphorylated AhSYMRK polypeptides are inactive and depletion of this inactive form leads to activation of intramolecular autophosphorylation of AhSYMRK. Together, our results suggest gatekeeper tyrosine autophosphorylation to be autoinhibitory for AhSYMRK.     

Phosphorylation and activation of protein tyrosine phosphatase (PTP) 1B by insulin receptor

We have previously reported a direct in vivo interaction between the activated insulin receptor and protein-tyrosine phosphatase-1B (PTP1B), which leads to an increase in PTP1B tyrosine phosphorylation. In order to determine if PTP1B is a substrate for the insulin receptor tyrosine kinase, the phosphorylation of the Cys 215 Ser, catalytically inactive mutant PTP1B (CS-PTP1B) was measured in the presence of partially purified and activated insulin receptor. In vitro, the insulin receptor tyrosine kinase catalyzed the tyrosine phosphorylation of PTP1B. 53% of the total cellular PTP1B became tyrosine phosphorylated in response to insulin in vivo. Tyrosine phosphorylation of PTP1B by the insulin receptor was absolutely dependent upon insulin-stimulated receptor autophosphorylation and required an intact kinase domain, containing insulin receptor tyrosines 1146, 1150 and 1151. Tyrosine phosphorylation of wild type PTP1B by the insulin receptor kinase increased phosphatase activity of the protein. Intermolecular transdephosphorylation was demonstrated both in vitro and in vivo, by dephosphorylation of phosphorylated CS-PTP1B by the active wild type enzyme either in a cell-free system or via expression of the wild type PTP1B into Hirc-M cell line, which constitutively overexpress the human insulin receptor and CS-PTP1B. These results suggest that PTP1B is a target protein for the insulin receptor tyrosine kinase and PTP1B can regulate its own phosphatase activity by maintaining the balance between its phosphorylated (the active form) and dephosphorylated (the inactive form) state.     

A proteomic investigation into the human cervical cancer cell line HeLa treated with dicitratoytterbium (III) complex

Lanthanides have been reported to induce apoptosis in cancer cell lines. Human cervical cancer cell line HeLa was found to be more sensitive to dicitratolanthanum (III) complex ([LaCit₂]³⁻) than other cancer cell lines. However, the effect and mechanism of dicitratoytterbium (III) complex ([YbCit₂]³⁻) on HeLa cells is unknown. Using biochemical and comparative proteomic analyses, [YbCit₂]³⁻ was found to inhibit HeLa cell growth and induce apoptosis. Similar to the effects of [LaCit₂]³⁻, proteomics results from [YbCit₂]³⁻-treated cells revealed profound changes in proteins relating to mitochondria and oxidative stress, suggesting that mitochondrial dysfunction plays a key role in [YbCit₂]³⁻-induced apoptosis. This was confirmed by the decreased mitochondrial transmembrane potential and the increased generation of reactive oxygen species in [YbCit₂]³⁻-treated cells. Western blot analysis showed that [YbCit₂]³⁻-induced apoptosis was accompanied by the activation of caspase-9 and specific proteolytic cleavage of PARP, leading to an increase in the pro-apoptotic protein Bax and a decrease in the anti-apoptotic protein Bcl-2. These results suggest a mitochondrial pathway of cell apoptosis in [YbCit₂]³⁻-treated cells, which will help us understand the molecular mechanisms of lanthanide-induced apoptosis in tumor cells.     

Identification and characterization of DUSP27, a novel dual-specific protein phosphatase

A novel human dual-specific protein phosphatase (DSP), designated DUSP27, is here described. The DUSP27 gene contains three exons, rather than the predicted 4-14 exons, and encodes a 220 amino acid protein. DUSP27 is structurally similar to other small DSPs, like VHR and DUSP13. The location of DUSP27 on chromosome 10q22, 50 kb upstream of DUSP13, suggests that these two genes arose by gene duplication. DUSP27 is an active enzyme, and its kinetic parameters and were determined. DUSP27 is a cytosolic enzyme, expressed in skeletal muscle, liver and adipose tissue, suggesting its possible role in energy metabolism.     

Impact of Ca2+ on structure of soybean CDPKβ and accessibility of the Tyr-24 autophosphorylation site

Several plant CDPKs were recently shown to be dual specificity kinases rather than Ser/Thr kinases as traditionally classified by sequence analysis. In the present study we confirm the autophosphorylation of recombinant soybean His₆-GmCDPKβ at the Tyr-24 site using sequence- and modification- specific antibodies. Homology modeling of soybean CDPKβ based on recent structures determined for several apicomplexan CDPKs suggested that phosphotyrosine-24 may be inaccessible to phosphatases. However, we report that dephosphorylation of CDPKβ by the protein tyrosine phosphatase 1B, PTP1B, was not restricted in the presence of calcium. Thus, despite conformational changes likely associated with calcium binding to the CDPKs, phosphotyrosine sites remain fully accessible to dephosphorylation suggesting the possibility of conformational breathing and flexing.