CIPK9: a calcium sensor-interacting protein kinase required for low-potassium tolerance in Arabidopsis

Potassium is one of the major macro-nutrients essential for a number of cellular processes in plants. Low potassium level in the soil represents a limiting factor for crop production. Recent studies have identified potassium transporters that are involved in potassium acquisition, and some of them are critical for potassium nutrition under low potassium conditions. However, little is understood on the molecular components involved in low potassium signaling and responses. We report here the identification ofa calcineurin B-like protein-interacting protein kinase （CIPK9） as a critical regulator of low potassium response in ,Arabidopsis. The CIPK9 gene was responsive to abiotic stress conditions, and its transcript was inducible in both roots and shoots by potassium deprivation. Disruption of CIPK9 function rendered the mutant plants hypersensitive to low potassium media. Further analysis indicated that K^＋ uptake and content were not affected in the mutant plants, implying CIPK9 in the regulation of potassium utilization or sensing processes.

Signal transduction mechanism of TRB3 in rats with non-alcoholic fatty liver disease

AIM： To evaluate the possible role of Tribble 3 （TRB3） in a rat model of non-alcoholic fatty liver disease （NAFLD） and its signal transduction mechanism.METHODS： Thirty Sprague-Dawley rats were randomized into three groups： normal control group, non-alcoholic fatty liver group A （fed on a high-fat diet for 8 wk） and group B （fed on a high-fat diet for 16 wk）. To determine the degree of hepatic steatosis in rats of each group, livers were stained with hematoxylin and eosin, and evaluated; real-time fluorescent quantitative reverse transcriptase-polymerase chain reaction was performed to measure the expression levels of TRI33 mRNA, and Western blotting analysis was done to determine the expression levels of protein kinase B （Akt） and phosphorylated protein kinase B （p-Akt-Thr308, p-Akt-Ser473）.RESULTS： Hepatic steatosis was evident in both NAFLD groups： mild to moderate hepatic steatosis occurred in group A, mainly as mild steatosis.Moderate to severe hepatic steatosis occurred in group B, mainly as severe steatosis. The expression level of TRB3 mRNA in group B was significantly higher than in the control group （122.28 ± 95.37 vs 3.06 ± 2.33,P = 0.002） and group A （122.28 ± 95.37 vs 5.77 ± 4.20,P = 0.001）. There was no significant difference in the expression levels of Akt （1.03 ± 0.53 vs 1.12 ± 0.77,P = 0.729） and p-Akt-Thr308 （0.82 ± 0.45 vs 0.92 ± 0.38, P = 0.592） between group A and the control group. The expression level of Akt and p-Akt-Thr308 in group B was significantly lower than in group A （Akt 0.41 ± 0.16 vs 1.12 ± 0.77, P = 0.008; p-Akt-Thr308 0.47 ± 0.19 vs 0.82 ± 0.45, P = 0.036） and the control group （Akt 0.41 ± 0.16 vs 1.03 ± 0.53, P = 0.018;p-Akt-Thr308 0.47 ± 0.19 vs 0.92 ± 0.38, P = 0.010）.The expression level of p-Akt-Ser473 in group A was significantly higher than in group B （1.48 ± 0.50 vs 0.81± 0.39, P = 0.041） as well as the control group （1.48 ± 0.50 vs 0.45 ± 0.26, P = 0.003）.CONCLUSION： TRB3 blocks insulin signaling by inhibiting Akt activation, which contributes to insulin resistance. It may be an important factor in the occurrence and development of NAFLD.

Potential role of chitinase 3-like-1 in inflammation-associated carcinogenic changes of epithelial cells

The family of mammalian chitinases includes members both with and without glycohydrolase enzymatic activity against chitin, a polymer of N-acetylglucosamine. Chitin is the structural component of fungi, crustaceans, insects and parasitic nematodes, but is completely absent in mammals. Exposure to antigens containing chitin- or chitin-like structures sometimes induces strong T helper type-I responses in mammals, which may be associated with the induction of mammalian chitinases. Chitinase 3-like-1 （CHI3L1）, a member of the mammalian chitinase family, is induced specifically during the course of inflammation in such disorders as inflammatory bowel disease, hepatitis and asthma. In addition, CHI3L1 is expressed and secreted by several types of solid tumors including glioblastoma, colon cancer, breast cancer and malignant melanoma. Although the exact function of CHI3L1 in inflammation and cancer is still largely unknown, CHI3L1 plays a pivotal role in exacerbating the inflammatory processes and in promoting angiogenesis and remodeling of the extracellular matrix. CHI3L1 may be highly involved in the chronic engagement of inflammation which potentiates development of epithelial tumorigenesis presumably by activating the mitogen-activated protein kinase and the protein kinase B signaling pathways. Anti-CHI3L1 antibodies or pan-chitinase inhibitors may have the potential to suppress CHI3Ll-mediated chronic inflammation and the subsequent carcinogenic change in epithelial cells.

Baicalin attenuates high fat diet-induced insulin resistance and ectopic fat storage in skeletal muscle,through modulating the protein kinase B/Glycogen synthase kinase 3 beta pathway

Insulin resistance is the pathophysiological basis of many diseases.Overcoming early insulin resistance highly significant in prevention diabetes,non-alcoholic fatty liver,and atherosclerosis.The present study aimed at evaluating the therapeutic effects of baicalin on insulin resistance and skeletal muscle ectopic fat storage in high fat diet-induced mice,and exploring the potential molecular mechanisms.Insulin resistance in mice was induced with a high fat diet for 16 weeks.Animals were then treated with three different doses of baicalin(100,200,and 400 mg·kg^(-1)·d^(-1)for 14 weeks.Fasting blood glucose,fasting serum insulin,glucose tolerance test(GTT),insulin tolerance test(ITT),and skeletal muscle lipid deposition were measured.Additionally,the AMP-activated protein kinase/acetyl-CoA carboxylase and protein kinase B/Glycogen synthase kinase 3 beta pathways in skeletal muscle were further evaluated.Baicalin significantly reduced the levels of fasting blood glucose and fasting serum insulin and attenuated high fat diet induced glucose tolerance and insulin tolerance.Moreover,insulin resistance was significantly reversed.Pathological analysis revealed baicalin dose-dependently decreased the degree of the ectopic fat storage in skeletal muscle.The properties of baicalin were mediated,at least in part,by inhibition of the AMPK/ACC pathway,a key regulator of de novo lipogenesis and activation of the Akt/GSK-3β pathway,a key regulator of Glycogen synthesis.These data suggest that baicalin,at dose up to 400 mg·kg^(-1)·d^(-1),is safe and able to attenuate insulin resistance and skeletal muscle ectopic fat storage,through modulating the skeletal muscle AMPK/ACC pathway and Akt/GSK-3β pathway.

PI3K expression and PIK3CA mutations are related to colorectal cancer metastases

AIM: To assess the significance of phosphatidylinositol 3-kinase (PI3K) in colorectal cancer (CRC) and toxicity of LY294002 in CRC cells with different metastatic abilities. METHODS: Sixty formalin-fixed and paraffin-embedded CRC tumor specimens were investigated. Adjacent normal colonic mucosa specimens from 10 of these cases were selected as controls. PI3K protein was detected by immunohistochemistry and PIK3CA mutations were investigated by gene sequencing analysis. A flowcytometry-based apoptosis detection kit was used to determine PI3K inhibitor-induced apoptosis in CRC cell lines SW480 and SW620. Expression of phosphorylated protein kinase B in CRC cell lines was detected by Western blotting. RESULTS: There was a significant difference in the proportion of primary lesions (30%, 18/60) vs metastatic lesions (46.7%, 28/60) that were positive for PI3K (P < 0.05). Mutations were detected in exon 9 (13.3%) and exon 20 (8.3%). Out of 60 cases, seven mutations were identified: two hotspot mutations, C.1633G>A resulting in E545A, and C.3140A>G resulting in H1047R; two novel missense mutations C.1624G>A and C.3079G>A; and three synonymous mutations (C.1641G>A, C.1581C>T and C.3027T>A). Exposure of SW480 cells to PI3K inhibitor for 48 h resulted in a significant increase of apoptotic cells in a dose-dependent manner [3.2% apoptotic cells in 0 μmol/L, 4.3% in 5 μmol/L, 6.3% in 10 μmol/L (P < 0.05), and 6.7% in 20 μmol/L (P < 0.05)]. Moreover, PI3K inhibitor induced a similar significant increase of apoptotic cells in the SW620 cell line for 48 h [3.3% apoptotic cells in 0 μmol/L, 13.3% in 5 μmol/L (P < 0.01), 19.2% in 10 μmol/L (P < 0.01), and 21.3% in 20 μmol/L (P < 0.01)]. CONCLUSION: High PI3K expression is associated with CRC metastasis. PI3K inhibitor induced apoptosis in CRC cells and displayed strong cytotoxicity for highly metastatic cells. PI3K inhibition may be an effective treatment for CRC.

Evodiamine induces extrinsic and intrinsic apoptosis of ovarian cancer cells via the mitogen-activated protein kinase/phosphatidylinositol-3-kinase/protein kinase B signaling pathways

OBJECTIVE: To explore the effects of evodiamine on ovarian cancer cells and the mechanisms underlying such effects.METHODS: Human ovarian cancer cells HO-8910 PM were treated with evodiamine at 0, 1.25,2.5, and 5 μM for 1-4 d. 3-(4,5-Dimethiylthiazol-2-yl)-2,5-diphenyltetrazolium bromide(MTT) assay was used to detect the growth inhibition rate of evodiamine-treated HO-8910 PM cells. The cell cycle was observed via propidium iodide(PI) staining. Apoptosis induction was assessed via Annexin V-fluorescein isothiocyanate/propidium iodide(Annexin V-FITC/PI) double staining assay. To verify the mechanism of apoptosis, caspase-dependent apoptotic pathway-related protein was detected by Western blot analysis. The expression levels of mitogen-activated protein kinase(MAPK)and/or phosphatidylinositol-3-kinase(PI3K)/pro-tein kinase B(Akt) pathway-related proteins were also investigated.RESULTS: Evodiamine significantly inhibited the proliferation of HO-8910 PM cells in a dose- and time-dependent manner. Evodiamine induced G2/M arrest with an increase of cyclin B1 level, and promoted cell apoptosis with a decrease of B cell lymphoma/lewkmia-2(Bcl-2) and an increase of Bcl-2-associated X protein(Bax) level. In addition,evodiamine treatment led to the activation of caspase-8, caspase-9, and caspase-3 and the cleavage of poly(ADP-ribose)-polymerase(PARP). Evodiamine targeted the MAPK and/or PI3K/Akt pathways by reducing the expression and activity of PI3 K, Akt, and extracellular signal-regulated kinase mitogen-activated protein kinase(ERK1/2 MAPK)and the activity of p38 MAPK.CONCLUSION: Evodiamine can inhibit the growth of ovarian cancer cells by G2/M arrest and intrinsic and extrinsic apoptosis. In addition, evodiamine-induced PI3K/Akt, ERK1/2 MAPK, and p38 MAPK signaling may be involved in cell death.

内脂素激活PI3K-Akt和MAPK-ERK1/2信号通路抑制MIN6细胞凋亡

探讨内脂素对胰岛β细胞株MIN6细胞信号通路和棕榈酸诱导细胞凋亡的影响,并探讨其分子机制.人重组内脂素呈剂量和时间依赖性促进MIN6细胞细胞外信号调节激酶（ERK）1/2和蛋白激酶B（Akt）的磷酸化,抑制棕榈酸诱导的MIN6细胞凋亡（P〈0.05或P〈0.01）.激活磷脂酰肌醇3激酶（PBK）-Akt和丝裂原活化蛋白激酶（MAPK）-ERK1/2信号通路是内脂素抑制MIN6细胞凋亡的分子机制之一.