Identification of QTLs Conferring Resistance to Bacterial Diseases in Rice.

Bacterial panicle blight, bacterial leaf streak, and bacterial brown stripe are common bacterial diseases in rice that represent global threats to stable rice yields. In this study, we used the rice variety HZ, Nekken and their 120 RIL population as experimental materials. Phenotypes of the parents and RILs were quantitatively analyzed after inoculation with Burkholderia glumae, Xanthomonas oryzae pv. oryzicola, and Acidovorax avenae subsp. avenae. Genetic SNP maps were also constructed and used for QTL mapping of the quantitative traits. We located 40 QTL loci on 12 chromosomes. The analysis of disease resistance-related candidate genes in the QTL regions with high LOD value on chromosomes 1, 3, 4, and 12 revealed differential expression before and after treatment, suggesting that the identified genes mediated the variable disease resistance profiles of Huazhan and Nekken2. These results provide an important foundation for cloning bacterial-resistant QTLs of panicle blight, leaf streak, and brown stripe in rice.

PHOX2B as a Reliable Marker for Neuroblastoma in Tissue and Cytology Specimens.

To investigate the diagnostic utility of immunohistochemistry for paired-like homeobox 2B (PHOX2B) expression in neuroblastomas (NBs) and tumors that mimic them, tissue samples (n = 229) from 157 cases of NB, 210 central nervous system tumors, and 170 extracranial non-NB solid tumors (n = 170) were immunostained for PHOX2B. Additionally, PHOX2B expression in 67 body fluid cytology specimens was analyzed. In tissue specimens, PHOX2B expression was positive in NBs, pheochromocytomas, and paragangliomas but negative in all of the other tumors evaluated. PHOX2B was detected by immunohistochemistry in 5 NB cytology specimens; all of the others were negative. These results suggest that PHOX2B may be a sensitive and specific immunohistochemical marker for the pathological diagnosis and differential diagnosis of NB in both tissue and cytology specimens.

C/EBP-α induces autophagy by binding to Beclin1 through its own acetylation modification in activated hepatic stellate cells.

The activation of hepatic stellate cells (HSCs) plays a key role in the occurrence of liver fibrosis，and promoting the apoptosis of activated HSCs or reducing the number of activated HSCs can reverse the development of liver fibrosis. In our previous studies, we have demonstrated that the CCAAT/enhancer binding protein α (C/EBP-α) played an important role in promoting the apoptosis of activated HSCs, thereby exerting an anti-liver fibrosis effect. Unlike apoptosis, autophagy, as a caspase-independent programmed cell death, can promptly remove the abnormal accumulation of substances or damaged organelles in cells and play a key role in regulating the homeostasis of intracellular environment. However, it is still unclear whether C/EBP-α participates in the occurrence of autophagy in HSCs. Therefore, in this study, we firstly used the methods of Western blot and immunofluorescence to characterize the consequence of C/EBP-α overexpression on the expression of proteins LC3B, P62, ATG5 and Beclin1 which were related to autophagy in HSCs. Subsequently, we performed Western blot and site-directed mutagenesis methods to clarify the type and related mechanism of autophagy which was induced by C/EBP-α. Here we show that C/EBP-α promotes the occurrence of autophagy in HSCs and the autophagy induced by C/EBP-α belongs to mitophagy. The stability of C/EBP-α protein regulates the level of autophagy in HSCs. In addition, acetylation of C/EBP-α also regulates the occurrence of autophagy in HSCs. Acetylation of lysine at positions K298, K302 and K326 of C/EBP-α promotes its binding to Beclin1. In conclusion, our study uncovers the role of C/EBP-α in regulating autophagy in HSCs, thereby providing a new strategy for clinical treatment of liver fibrosis.

Poorly differentiated SMARCB1/INI1-negative chordomas.

AIMS: To analyze the clinicopathological characteristics of poorly differentiated chordomas (PDCs) with SMARCB1/INI1 loss in children. MATERIALS: Four cases of PDCs were included in the study. METHODS: Immunohistochemistry was performed with respect to brachyury, Glut-1, keratin 18, keratin 19, INI1, vimentin, S-100, CK, EMA, GFAP, etc. Fluorescence in situ hybridization (FISH) was performed for SMARCB1/INI1 from 3 patients. RESULTS: Histologically, contrary to typical histologic features for conventional chordomas, 4 tumors were composed of ovoid or atypical fusiform cells. Sporadic physaliphorous cells were evident. Tumor cells had large vacuoles in the cytoplasm that were even remarkable on the imprint cytology slide. By immunohistochemistry, each case revealed loss of SMARCB1/INI1 expression and nuclear expression of brachyury. Glut-1, keratin 18, keratin 19, CK, EMA, and vimentin were positive in these PDCs. Except for 1 patient who had not yet completed FISH, the other 3 cases demonstrated the loss of SMARCB1/INI1 gene by fluorescence in situ hybridization. CONCLUSION: Poorly differentiated SMARCB1/INI1-negative chordoma is a unique subset of chordoma representing a clinically, histopathologically, and molecularly distinct entity with rapid progression and poor prognosis which should not be confused with conventional chordomas. Sporadic physaliphorous cells (tumor cells with large vacuoles in the cytoplasm) provided important diagnostic clues of PDCs. Combination use of characteristic markers of notochord cells (brachyury, Glut-1, keratin 18, and keratin 19) along with INI1 were effective diagnostic tools.

Nuclear prelamin a recognition factor and iron dysregulation in multiple sclerosis.

Dysregulation of iron metabolism and aberrant iron deposition has been associated with multiple sclerosis. However, the factors that contribute to this pathological state remain to be understood. In this study, human multiple sclerosis and mice brain samples were analyzed through mass spectrometry as well as histological and immunoblot techniques, which demonstrated that iron deposition is associated with increased levels of nuclear prelamin A recognition factor (NARF). NARF is a protein associated with the mitochondria which has also been linked to mitochondrial defects in multiple sclerosis. We report NARF to be associated in multiple sclerosis pathology and aberrant iron deposition.

Predictive biomarkers of colorectal cancer.

Colorectal cancer is one of the top leading causes of cancer mortality worldwide, especially in China. However, most of the current treatments are invasive and can only be applied to very few cancers. The earlier a malignant tumor is diagnosed, the higher the patient's survival rate. In this study, we proposed a computational framework to identify highly-reliable and easierly-detectable biomarkers capable of secreting into blood, urine and saliva by integrating transcriptomics and proteomics data at the system biology level. First, a large number of transcriptome data were processed to identify candidate biomarkers for colorectal cancer. Second, three classified models are constructed to predict biomarkers for colorectal cancer capable of secreting into blood, urine and saliva, which are effective disease diagnosis media to facilitate clinical screening. Then biological functions and molecular mechanisms of the candidate biomarkers of colorectal cancer are inferred utilizing multi-source biological knowledge and literature mining. Furthermore, the classification power of different combinations of candidate biomarkers is verified by machine learning models. In addition, the targeted drugs of the predicted biomarkers are further analyzed to provide assistance for clinical treatment of colorectal cancer. In this paper, our proposed computational model not only provides the effective candidate biomarkers ESM1, CTHRC1, AZGP1 for colorectal cancer capable of secreting into blood, urine and saliva, but also helps to understand the molecular mechanism of colorectal cancer. This computational framework can span the huge gap between transcriptome and proteomics, which can easily be applied to the biomarker research for other types of tumor.

The Role of Deimination in Regenerative Reprogramming of Neurons.

Neurons from the adult central nervous system (CNS) demonstrate limited mRNA transport and localized protein synthesis versus developing neurons, correlating with lower regenerative capacity. We found that deimination (posttranslational conversion of protein-bound arginine into citrulline) undergoes upregulation during early neuronal development while declining to a low basal level in adults. This modification is associated with neuronal arborization from amphibians to mammals. The mRNA-binding proteins (ANP32a, REF), deiminated in neurons, have been implicated in local protein synthesis. Overexpression of the deiminating cytosolic enzyme peptidyl arginine deiminase 2 in nervous systems results in increased neuronal transport and neurite outgrowth. We further demonstrate that enriching deiminated proteins rescues transport deficiencies both in primary neurons and mouse optic nerve even in the presence of pharmacological transport blockers. We conclude that deimination promotes neuronal outgrowth via enhanced transport and local protein synthesis and represents a new avenue for neuronal regeneration in the adult CNS.

Trichostatin A inhibits the activation of Hepatic stellate cells by Increasing C/EBP-α Acetylation in vivo and in vitro.

Reversal of activated hepatic stellate cells (HSCs) to a quiescent state and apoptosis of activated HSCs are key elements in the reversion of hepatic fibrosis. CCAAT/enhancer binding protein α (C/EBP-α) has been shown to inhibit HSC activation and promote its apoptosis. This study aims to investigate how C/EBP-α acetylation affects the fate of activated HSCs. Effects of a histone deacetylation inhibitor trichostatin A (TSA) on HSC activation were evaluated in a mouse model of liver fibrosis caused by carbon tetrachloride (CCl4) intoxication. TSA was found to ameliorate CCl4-induced hepatic fibrosis and improve liver function through increasing the protein level and enhancing C/EBP-α acetylation in the mouse liver. C/EBP-α acetylation was determined in HSC lines in the presence or absence of TSA, and the lysine residue K276 was identified as a main acetylation site in C/EBP-α protein. C/EBP-α acetylation increased its stability and protein level, and inhibited HSC activation. The present study demonstrated that C/EBP-α acetylation increases the protein level by inhibiting its ubiquitination-mediated degradation, and may be involved in the fate of activated HSCs. Use of TSA may confer an option in minimizing hepatic fibrosis by suppressing HSC activation, a key process in the initiation and progression of hepatic fibrosis.

Design and synthesis of novel dasatinib derivatives as inhibitors of leukemia stem cells.

We used the concept of bioisosteres to design and synthesize a novel series of dasatinib derivatives for the treatment of leukemia. Unfortunately, most of the dasatinib derivatives did not show appreciable inhibition against leukemia cell lines K562 and HL60. However, acrylamide compound 2c had comparable inhibitory activity with dasatinib against K562 cells (IC50 = 0.039 nM vs. 0.069 nM). And amide compound 2a and acrylamide compound 2c also had comparable inhibitory activity with dasatinib against the leukemia cell line HL60 (IC50 = 0.25 nM and 0.26 nM vs. 0.11 nM). Against the leukemia progenitor cell line KG1a, triazole compounds 15a and 15d-15f and oxadiazole compounds 24a-24d were more potent than dasatinib. In particular, the hydroxyl compounds 15a and 24a were about 64 and 180 fold more potent than dasatinib against KG1a cells (IC50 = 0.14 µM and 0.05 µM vs. 8.98 µM). Compounds 15a and 24a also inhibited colony formation in MCF-7 cells and inhibited cell migration in the cell wound scratch assay in B16BL6 cells. Moreover, hydroxyl compounds 15a and 24a had low toxicity in vivo.

Selective enrichment of N-linked glycopeptides and glycans by using a dextran-modified hydrophilic material.

Glycosylation analysis of proteins from biological sources utilizing mass spectrometry based approaches is challenging due to the relatively low abundance of glycopeptides, the structural diversity of glycans, and the coexisting matrices. In this study, a customized dextran-bonded silica-based stationary phase was introduced for selective enrichment of glycopeptides and glycans from complex biological samples. This material has exhibited superior selectivity and broader glycosylation site coverage over commercial Sepharose in glycoproteomic evaluation. Additionally, the glycomic analysis of fetuin, α1 -acid glycoprotein, and human serum N-glycome also indicated the relatively higher sensitivity, selectivity, and glycoform coverage of dextran-bonded silica than that of Sepharose and porous graphitized carbon. Therefore, the dextran-bonded silica is expected to make contributions in the fields of glycoproteomics and glycomics.

TSA increases C/EBP­α expression by increasing its lysine acetylation in hepatic stellate cells.

CCAAT enhancer binding protein­α (C/EBP­α) is a transcription factor expressed only in certain tissues, including the liver. It has been previously demonstrated that C/EBP­α may induce apoptosis in hepatic stellate cells (HSCs), raising the question of whether acetylation of C/EBP­α is associated with HSCs, and the potential associated mechanism. A total of three histone deacetylase inhibitors (HDACIs), including trichostatin A (TSA), suberoylanilide hydroxamic acid and nicotinamide, were selected to determine whether acetylation affects C/EBP­α expression. A Cell Counting Kit­8 assay was used to determine the rate of proliferation inhibition following treatment with varying doses of the three HDACIs in HSC­T6 and BRL­3A cells. Western blot analysis was used to examine Caspase­3, ­8, ­9, and ­12 levels in HSC­T6 cells treated with adenoviral­C/EBP­α and/or TSA. Following treatment with TSA, a combination of reverse transcription­quantitative polymerase chain reaction and western blot analyses was used to determine the inherent C/EBP­α mRNA and protein levels in HSC­T6 cells at 0, 1, 2, 4, 8, 12, 24, 36 and 48 h. Nuclear and cytoplasmic proteins were extracted to examine C/EBP­α distribution. Co­immunoprecipitation analysis was used to examine the lysine acetylation of C/EBP­α. It was observed that TSA inhibited the proliferation of HSC­T6 cells to a greater extent compared with BRL­3A cells, following treatment with the three HDACIs. TSA induced apoptosis in HSC­T6 cells and enhanced the expression of C/EBP­α. Following treatment of HSC­T6 cells with TSA, inherent C/EBP­α expression increased in a time­dependent manner, and its lysine acetylation simultaneously increased. Therefore, the results of the present study suggested that TSA may increase C/EBP­α expression by increasing its lysine acetylation in HSCs.

Rab14 Suppression Mediated by MiR-320a Inhibits Cell Proliferation, Migration and Invasion in Breast Cancer.

We found that microRNA-320a (miR-320a) was an attractive prognostic biomarker in breast cancer (BC) previously, whereas its regulatory mechanism in BC was not well understood. Our aim was to identify miR-320a target gene, examine the clinical relationship between miR-320a and its target, and further explore the functions of its target in BC. In this study, miR-320a downstream target gene was determined in HEK-293T cells by dual luciferase reporter assay. Then western blotting and immunohistochemistry were used to assess miR-320a target gene expression in fresh frozen (n=19, breast cancer and matched non-malignant adjacent tissue samples) and formalin-fixed paraffin-embedded (FFPE) (n=130, invasive BC tissues, the same panel detected for miR-320a expression previously) breast tissues, respectively. The results suggested that miR-320a could significantly suppressed Rab14 3'-untranslated region luciferase-reporter activity, and thus Rab14 was first identified as miR-320a target in BC. In 19 matched breast tissues, 12 (63%) breast cancer tissues showed high expression of Rab14 compared with the corresponding normal tissues. Rab14 immunoreactivity was mainly detected in the cytoplasm, 77/130 (59.2%) showed high expression. Furthermore, Rab14 expression was found to be inversely correlated with miR-320a expression in fresh-frozen breast tissues as well as in FFPE invasive breast cancer samples. In addition, Rab14 expression levels were positively related to tumor size (P = 0.034), lymph node metastasis (P < 0.001), distant metastasis (P = 0.001), histological grade (P = 0.035) and clinical tumor lymph-node metastasis stage (P = 0.001). Patients with higher Rab14 expression showed shorter overall survival time. Moreover, silencing of Rab14 could suppress proliferation, migration and invasion in breast cancer cell lines. Collectively, our results indicate that miR-320a could target Rab14 and that they could interact biologically in BC.

TWIST1 and BMI1 in Cancer Metastasis and Chemoresistance.

Purpose Increasing evidences revealed that cancer cells with the characteristics of epithelial-mesenchymal transition (EMT) or cancer stem cells (CSC) have high ability of progression, invasion, metastasis and chemoresistance. TWIST1 and BMI1 are crucial transcription factors required for EMT and CSC. Both TWIST1 and BMI1 are up-regulated in various cancers and have a positive correlation with poor prognosis. Although recent results showed that the two molecules function in promoting cancer metastasis and chemoresistance respectively, the correlation of TWIST1 and BMI1 is not well understood. Methods In this review, we summarize recent advance in cancer research focus on TWIST1 and BMI1 in cancer metastasis and chemoresistance, and emphasize the possible link between EMT and CSC. Results Further investigation of TWIST1 and BMI1 cooperately promote CSC proliferation due to EMT-associated effect will help to understand the mechanism of tumor cells metastasis and chemoresistance. Conclusions TWIST1 and BMI1 in cancer cells will be effective targets for treating chemoresistant metastatic lesions.

MicroRNA-320a inhibits breast cancer metastasis by targeting metadherin.

Dysregulated microRNAs play important pathological roles in carcinogenesis that are yet to be fully elucidated. This study was performed to investigate the biological functions of microRNA-320a (miR-320a) in breast cancer and the underlying mechanisms. Function analyses for cell proliferation, cell cycle, and cell invasion/migration, were conducted after miR-320a silencing and overexpression. The specific target genes of miR-320a were predicted by TargetScan algorithm and then determined by dual luciferase reporter assay and rescue experiment. The relationship between miR-320a and its target genes was explored in human breast cancer tissues. We found that miR-320a overexpression could inhibit breast cancer invasion and migration abilities in vitro, while miR-320a silencing could enhance that. In addition, miR-320a could suppress activity of 3'-untranslated region luciferase of metadherin (MTDH), a potent oncogene. The rescue experiment revealed that MTDH was a functional target of miR-320a. Moreover, we found that MTDH was negatively correlated with miR-320a expression, and it was related to clinical outcomes of breast cancer. Further xenograft experiment also showed that miR-320a could inhibit breast cancer metastasis in vivo. Our findings clearly demonstrate that miR-320a suppresses breast cancer metastasis by directly inhibiting MTDH expression. The present study provides a new insight into anti-oncogenic roles of miR-320a and suggests that miR-320a/MTDH pathway is a putative therapeutic target in breast cancer.