OsCIP1, a secreted protein, binds to and stabilizes OsCR4 to promote aleurone layer development, seed germination and early seedling growth in rice.

The receptor kinase CRINKLY 4 (CR4) and its orthologs are known for their essential roles in cell differentiation and their shuttling between plasma membrane and cytoplasmic vesicles, a unique feature tied to their extracellular domain. However, the extracellular regulators of CR4 have been little known. Here we identified an OsCR4 Interacting Protein 1 (OsCIP1) (also named as OsLTPL36 in rice) by a yeast two-hybrid screen using the extracellular domain of OsCR4 (OsCR4E) as bait. OsCIP1/OsLTPL36 harbors a signal peptide and is localized to the outer surface of the plasma membrane. It interacted with the TNFR subdomain of OsCR4, causing an increase in OsCR4 recycling to the plasma membrane. oscip1, in which OsCR4 protein was decreased, exhibited thinner aleurone layer, late germination and delayed growth; while OsCIP1-overexpressing plants, in which OsCR4 protein was increased, displayed enhanced growth at the early seedling stage. OsCIP1 was cleaved between W61 and Q62, and the resulting C-terminal half exhibited a greater affinity for OsCR4E than did its precursor. Abolishing this cleavage site compromises OsCIP1's ability to promote seedling growth. Our results provide valuable clues for the regulation of CR4 activity and its functions in aleurone layer cell differentiation by a secreted small protein in rice.

Receptor-like kinase OsCR4 controls leaf morphogenesis and embryogenesis by fixing the distribution of auxin in rice.

Cell differentiation is a key event in organ development; it involves auxin gradient formation, cell signaling, and transcriptional regulation. Yet, how these processes are orchestrated during leaf morphogenesis is poorly understood. Here, we demonstrate an essential role for the receptor-like kinase OsCR4 in leaf development. oscr4 loss-of-function mutants displayed short shoots and roots, with tiny, crinkly, or even dead leaves. The delayed outgrowth of the first three leaves and seminal root in oscr4 was due to defects in plumule and radicle formation during embryogenesis. The deformed epidermal, mesophyll, and vascular tissues observed in oscr4 leaves arose at the postembryo stage; the corresponding expression pattern of proOsCR4:GUS in embryos and young leaves suggests that OsCR4 functions in these tissues. Signals from the auxin reporter DR5rev:VENUS were found to be altered in oscr4 embryos and disorganized in oscr4 leaves, in which indole-3-acetic acid accumulation was further revealed by immunofluorescence. OsWOX3A, which is auxin responsive and related to leaf development, was activated extensively and ectopically in oscr4 leaves, partially accounting for the observed lack of cell differentiation. Our data suggest that OsCR4 plays a fundamental role in leaf morphogenesis and embryogenesis by fixing the distribution of auxin.

Protein Phosphatase 2A B'α and B'ß Protect Centromeric Cohesion during Meiosis I.

During meiosis, the stepwise release of sister chromatid cohesion is crucial for the equal distribution of genetic material to daughter cells, enabling generation of fertile gametophytes. However, the molecular mechanism that protects centromeric cohesion from release at meiosis I is unclear in Arabidopsis (Arabidopsis thaliana). Here, we report that the protein phosphatase 2A regulatory subunits B'α and B'ß participate in the control of sister chromatid separation. The double mutant b'αß exhibited severe male and female sterility, caused by the lack of a nucleus or presence of an abnormal nucleus in mature microspores and embryo sacs. 4',6-Diamidino-2-phenylindole staining revealed unequal amounts of DNA in the mononuclear microspores. Transverse sections of the anthers revealed unevenly sized tetrads with or without a nucleus, suggesting a defect in meiocyte meiosis. An analysis of chromosome spreads showed that the sister chromatids separated prematurely at anaphase I in b'αß Immunoblotting showed that AtRECOMBINATION DEFECTIVE8 (AtREC8), a key member of the cohesin complex, was hyperphosphorylated in b'αß anthers and pistils during meiosis but hypophosphorylated in the wild type. Furthermore, yeast two-hybrid and bimolecular fluorescence complementation assays showed that B'α and B'ß interact specifically with AtREC8, AtSHUGOSHIN1 (AtSGO1), AtSGO2, and PATRONUS1. Given that B'α was reported to localize to the centromere in meiotic cells, we propose that protein phosphatase 2A B'α and B'ß are recruited by AtSGO1/2 and PATRONUS1 to dephosphorylate AtREC8 at the site of centromere cohesion to shield it from cleavage until anaphase II, contributing to the balanced separation of sister chromatids at meiosis.

Using receiver operating characteristic curves to evaluate the diagnostic value of the combination of multislice spiral CT and alpha-fetoprotein levels for small hepatocellular carcinoma in cirrhotic patients.

BACKGROUND: The various combination of multiphase enhancement multislice spiral CT (MSCT) makes the diagnosis of a small hepatocellular carcinoma (sHCC) on the background of liver cirrhosis possible. This study was to explore whether the combination of MSCT enhancement scan and alpha-fetoprotein (AFP) level could increase the diagnostic efficiency for sHCC. METHODS: This study included 35 sHCC patients and 52 cirrhotic patients without image evidence of HCC as a control group. The diagnoses were made by three radiologists employing a 5-point rating scale, with postoperative pathologic results as the gold standard. Receiver operating characteristic (ROC) curve analysis was performed to evaluate the diagnostic value of the three MSCT combination modes (arterial phase+portal-venous phase, arterial phase+delayed phase, arterial phase+portal-venous phase+delayed phase) and AFP levels for sHCC on the background of liver cirrhosis. RESULTS: The area under ROC curve (AUC), sensitivity, and specificity of the combination of arterial phase+portal-venous phase+delayed phase were 0.93, 93%, and 82%, respectively. The average AUC of the arterial phase+portal-venous phase+delayed phase combination was significantly greater than that of the arterial phase+portal-venous phase (AUC=0.84, P=0.01) and arterial phase+delayed phase (AUC=0.85, P=0.03). Arterial phase+portal-venous phase had a smaller AUC (0.84) than arterial phase+delayed phase (0.85), but the difference was insignificant (P=0.15). After combining MSCT enhancement scan with AFP, the AUC, sensitivity, and specificity were 0.95, 94%, and 83%, respectively, indicating a greatly increased diagnostic efficiency for sHCC. CONCLUSIONS: The combination of AFP and 3 phases MSCT enhancement scan could increase the diagnostic efficiency for sHCC on the background of liver cirrhosis. The application of ROC curve analysis has provided a new method and reference in HCC diagnosis.

Effects of silencing S100A8 and S100A9 with small interfering RNA on the migration of CNE1 nasopharyngeal carcinoma cells.

The calcium-binding S100 proteins are involved in functions such as cell growth, differentiation, migration, adhesion and signal transduction. S100A8 and S100A9 are highly expressed in a variety of tumor cells, and are implicated in tumor development and progression. However, the role of S100A8 and S100A9 in nasopharyngeal carcinoma (NPC) cell migration is unclear. The present study investigated the effect of S100A8 and S100A9 on migration using a NPC cell line, CNE1. The CNE1 cells were transfected with S100A8 or S100A9 small interfering RNA (siRNA). Reverse transcription-quantitative polymerase chain reaction (RT-qPCR) was used to detect S100A8 and S100A9 gene expression. Following the downregulation of S100A8 or S100A9, the effects on cell migration were determined using wound-healing assays. The expression of matrix metalloproteinase-7 (MMP7), a member of the MMP family that is associated with tumor cell invasion and migration, was also detected by RT-qPCR. S100A8 and S100A9 siRNAs effectively suppressed S100A8 and S100A9 gene expression, and substantially inhibited the migration of the CNE1 cells. In addition, MMP7 expression was reduced to varying extents in S100A8 and S100A9 siRNA-treated cells compared with controls. Thus, S100A8 and S100A9 promoted the migration of CNE1 NPC cells.

Structural characterisation and antioxidant activity evaluation of phenolic compounds from cold-pressed Perilla frutescens var. arguta seed flour.

A total of 11 phenolic compounds, as well as sucrose (12) and tryptophan (13), were isolated from cold-pressed Perilla frutescens var. arguta seed flour using column chromatography, and their chemical structures were identified as 3'-dehydroxyl-rosmarinic acid-3-o-glucoside (1), rosmarinic acid-3-o-glucoside (2), rosmarinic acid (3), rosmarinic acid methyl ester (4), luteolin (5), luteolin-5-o-glucoside (6), apigenin (7), caffeic acid (8), caffeic acid-3-o-glucoside (9), vanillic acid (10) and cimidahurinine (11) using NMR and time-of-flight mass spectrometry. Of these components, compound 1 is novel, and this is the first report of compounds 10 and 11 in perilla seeds. HPLC quantification combined with antioxidant activity evaluation revealed that rosmarinic acid and rosmarinic acid-3-o-glucoside were the dominant phenolic antioxidants with strong antioxidant activities.

[Study on the apoptosis and its mechanism of lovo human colorectal cancer cells induced by SFPS].

Human colorectal cancer (lovo) cells were chose to study the anti-tumor effects of SFPS and explore the significance of caspase 3 in the apoptosis of lovo cells induced by SFPS. Inhibition of the cell proliferation was measured by MTT assay. SFPS induced apoptosis of lovo cells was observed by electron microscopy, flow cytometry and DNA electrophoresis. Furthermore, the expressions of caspase 3 mRNA and pro-caspase 3 were tested by RT-PCR and Western blot. SFPS exhibited anti-proliferative activity in the dosage and time-dependent manner. After incubation for 24, 36, 48 and 72 h, the IC50 of SFPS on lovo cells was 375 mg/L, 355 mg/L, 178 mg/L and 60 mg/L, respectively. DNA ladders of lovo cells were showed on agarose gel electrophoresis and the fragments of DNA were integral of 180-200 bp 24 hours after SFPS treatment at the doses of 5 mg/L, 50 mg/L and 300 mg/L. However, mistiness DNA ladder or smear was found in lovo cells treated with 500 mg/L SFPS. When SFPS concentration was 1000 mg/l, DNA ladder disappeared and showed smear entirely. Morphological examination showed chromosomal condensation, karyotheca margination, cell shrinkage and the presence of apoptosis bodies with electron microscopy. With concentration dependent manner, the apoptosis and sub-G1 peaks were observed through flow cytometry, but the cell cycle didn't change obviously. Furthermore, the expression of pro-caspase 3 was decreased and the level of caspase 3 mRNA was increased in the time and dose-dependent manner. It suggests that SFPS may induce the apoptosis of lovo cells in vitro resulting in the inhibition of proliferation. And caspase 3 activation may participate in the processes of the apoptosis of lovo cells induced by SFPS.