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To understand the potential effects of cancer cells on surrounding normal mammary epithelial cells, we performed direct co-culture of non-tumorigenic mammary epithelial MCF10A cells and various breast cancer cells. Firstly, we observed dynamic cell–cell interactions between the MCF10A cells and breast cancer cells including lamellipodia or nanotube-like contacts and transfer of extracellular vesicles. Co-cultured MCF10A cells exhibited features of epithelial-mesenchymal transition, and showed increased capacity of cell proliferation, migration, colony formation, and 3-dimensional sphere formation. Direct co-culture showed most distinct phenotype changes in MCF10A cells followed by conditioned media treatment and indirect co-culture. Transcriptome analysis and phosphor-protein array suggested that several cancer-related pathways are significantly dysregulated in MCF10A cells after the direct co-culture with breast cancer cells. S100A8 and S100A9 showed distinct up-regulation in the co-cultured MCF10A cells and their microenvironmental upregulation was also observed in the orthotropic xenograft of syngeneic mouse mammary tumors. When S100A8/A9 overexpression was induced in MCF10A cells, the cells showed phenotypic features of directly co-cultured MCF10A cells in terms of in vitro cell behaviors and signaling activities suggesting a S100A8/A9-mediated transition program in non-tumorigenic epithelial cells. This study suggests the possibility of dynamic cell–cell interactions between non-tumorigenic mammary epithelial cells and breast cancer cells that could lead to a substantial transition in molecular and functional characteristics of mammary epithelial cells. [ABSTRACT FROM AUTHOR]

A process has been developed for post fabrication manipulation of silica nanowire lengths with reproducible mean length target ability by manual grinding in liquid media. The process is based on the relationship between nanowire Reynolds number and the laminar or turbulent motion of nanowires in a media. Mean lengths of nanowires prepared by this process are predicted to be inversely proportional to the density over viscosity of the media used. Experimental results giving the mean length measurements are in very good agreement with the predicted dependence on medium density and viscosity. [ABSTRACT FROM AUTHOR]

While transfer-RNAs (tRNAs) are known to transport amino acids to ribosome, new functions are being unveiled from tRNAs and their fragments beyond protein synthesis. Here we show that phosphorylation of 90-kDa RPS6K (ribosomal proteins S6 kinase) was enhanced by tRNALeu overexpression under amino acids starvation condition. The phosphorylation of 90-kDa RPS6K was decreased by siRNA specific to tRNALeu and was independent to mTOR (mammalian target of rapamycin) signaling. Among the 90-kDa RPS6K family, RSK1 (ribosomal S6 kinase 1) and MSK2 (mitogen-and stress-activated protein kinase 2) were the major kinases phosphorylated by tRNALeu overexpression. Through SILAC (stable isotope labeling by/with amino acids in cell culture) and combined mass spectrometry analysis, we identified EBP1 (ErbB3-binding protein 1) as the tRNALeu-binding protein. We suspected that the overexpression of free tRNALeu would reinforce ErbB2/ErbB3 signaling pathway by disturbing the interaction between ErbB3 and EBP1, resulting in RSK1/MSK2 phosphorylation, improving cell proliferation and resistance to death. Analysis of samples from patients with breast cancer also indicated an association between tRNALeu overexpression and the ErbB2-positive population. Our results suggested a possible link between tRNALeu overexpression and RSK1/MSK2 activation and ErbB2/ErbB3 signaling. [ABSTRACT FROM AUTHOR]

Aminoacyl-tRNA synthetases (ARSs) acylate transfer (t)RNAs with amino acids. Charging tRNAs with the right amino acids is the first step in translation; therefore, the accurate and error-free functioning of ARSs is an essential prerequisite fortranslational fidelity. A recent study found that methionine (Met) can be incorporated into non-Met residues of proteins through methionylation of non-cognate tRNAs under conditions of oxidative stress. However, it was not understood how this mis-methionylation is achieved. Here, we report that methionyl-tRNA synthetase (MRS) is phosphorylated at Ser209 and Ser825 by extracellular signal-related kinase (ERK1/2) under conditions of stress caused by reactive oxygen species (ROS), and that this phosphorylated MRS shows increased affinity for non-cognate tRNAs with lower affinity for tRNAMet, leading to an increase in Met residues in cellular proteins. The expression of a mutant MRS containing the substitutions S209D and S825D, mimicking dual phosphorylation, reduced ROS levels and cell death. This controlled inaccuracy of MRS seems to serve as a defense mechanism against ROS-mediated damage at the cost of translational fidelity. [ABSTRACT FROM AUTHOR]

Lysyl-tRNA synthetase (KRS), a protein synthesis enzyme in the cytosol, relocates to the plasma membrane after a laminin signal and stabilizes a 67-kDa laminin receptor (67LR) that is implicated in cancer metastasis; however, its potential as an antimetastatic therapeutic target has not been explored. We found that the small compound BC-K-YH16899, which binds KRS, impinged on the interaction of KRS with 67LR and suppressed metastasis in three different mouse models. The compound inhibited the KRS-67LR interaction in two ways. First, it directly blocked the association between KRS and 67LR. Second, it suppressed the dynamic movement of the N-terminal extension of KRS and reduced membrane localization of KRS. However, it did not affect the catalytic activity of KRS. Our results suggest that specific modulation of a cancer-related KRS-67LR interaction may offer a way to control metastasis while avoiding the toxicities associated with inhibition of the normal functions of KRS. [ABSTRACT FROM AUTHOR]

Vancomycin resistant enterococci (VRE) isolates from humans (23 isolates) and poultry (20 isolates) were characterized by antibiotic susceptibility, vancomycin resistance transferability, pulsed-field gel electrophoresis (PFGE), and structural analysis of Tn 1546-like elements. VRE isolates from humans and poultry showed different resistance patterns, transferability, and transfer rate. In addition to these phenotypic differences between humans and poultry VRE, PFGE and the structure of Tn 1546-like elements were also distinct. Most poultry isolates (16/20) were identical to the prototype vanA transposon, Tn 1546, while most human isolates (21/23) had multiple integrations of insertion sequence. The transmission of VRE and vancomycin resistance determinant between humans and poultry could not be demonstrated in this study. [ABSTRACT FROM AUTHOR]

Objectives: To identify the staphylococcal cassette chromosome mec (SCCmec) types of methicillin-resistant Staphylococcus aureus (MRSA) isolated from bovine milk, and examine the genetic relatedness between MRSA from bovine milk and MRSA from human isolates.Methods: Antimicrobial susceptibility tests were performed on MRSA isolated from bovine milk. PCR and sequencing analysis were performed to determine the SCCmec type of MRSA, and to confirm their toxin carriage. Genetic relatedness among the bovine isolates and between bovine and human isolates was detected with PFGE and multilocus sequence typing (MLST).Results: Fourteen MRSA and a silent mecA-carrying methicillin-susceptible S. aureus (smMSSA) were isolated from the milk of cows with an isolation ratio of 0.18%. SCCmec of 14 MRSA strains were designated as new subtype IVg, and one smMSSA strain was not classified. All 14 MRSA strains shared Panton-Valentine leucocidin (PVL) and staphylococcal enterotoxin D (SED), SEI and SEJ; the smMSSA strain had only PVL. All MRSA and smMSSA isolates showed no multidrug resistance and had community-acquired MRSA (CA-MRSA) characteristics. PFGE revealed that all isolates except the smMSSA belonged to the same genetic lineage, and MLST analysis showed that they had no genetic relatedness with CA-MRSA which had caused human infection in Korea.Conclusions: MRSA isolated from bovine milk harboured a unique SCCmec subtype, and they may not be correlated with the emergence of CA-MRSA in human infection in Korea. [ABSTRACT FROM AUTHOR]

Aminoacyl-tRNA synthetase-interacting multifunctional protein 2 (AIMP2) is a non-enzymatic component required for the multi-tRNA synthetase complex. While exon 2 skipping alternatively spliced variant of AIMP2 (AIMP2-DX2) compromises AIMP2 activity and is associated with carcinogenesis, its clinical potential awaits further validation. Here, we found that AIMP2-DX2/AIMP2 expression ratio is strongly correlated with major cancer signaling pathways and poor prognosis, particularly in acute myeloid leukemia (AML). Analysis of a clinical patient cohort revealed that AIMP2-DX2 positive AML patients show decreased overall survival and progression-free survival. We also developed targeted RNA-sequencing and single-molecule RNA-FISH tools to quantitatively analyze AIMP2-DX2/AIMP2 ratios at the single-cell level. By subclassifying hematologic cancer cells based on their AIMP2-DX2/AIMP2 ratios, we found that downregulating AIMP2-DX2 sensitizes cells to anticancer drugs only for a subgroup of cells while it has adverse effects on others. Collectively, our study establishes AIMP2-DX2 as a potential biomarker and a therapeutic target for hematologic cancer. Ku, Kim et al develop a method to analyse the ratio of the alternatively spliced variant of AIMP2 to full length AIMP via single-molecule RNA-FISH. They can subclassify hematologic cancer based on AIMP2-DX2/AIMP2 ratio and find that cells with high AIMP2-DX2 ratio can be sensitized to chemotherapy drugs by depleting AIMP2-DX2. [ABSTRACT FROM AUTHOR]

AIMP2-DX2, an exon 2-deleted splice variant of AIMP2 (aminoacyl-tRNA synthetase-interacting multifunctional protein 2), is highly expressed in lung cancer and involved in tumor progression in vivo. Oncogenic function of AIMP2-DX2 and its correlation with poor prognosis of cancer patients have been well established; however, the application of this potentially important biomarker to cancer research and diagnosis has been hampered by a lack of antibodies specific for the splice variant, possibly due to the poor immunogenicity and/or stability of AIMP2-DX2. In this study a monoclonal antibody, H5, that specifically recognizes AIMP2-DX2 and its isoforms was generated via rabbit immunization and phage display techniques, using a short peptide corresponding to the exon 1/3 junction sequence as an antigen. Furthermore, based on mutagenesis, limited cleavage, and mass spectrometry studies, it is also suggested that the endogenous isoform of AIMP2-DX2 recognized by H5 is produced by proteolytic cleavage of 33 amino acids from N-terminus and is capable of inducing cell proliferation similarly to the uncleaved protein. H5 monoclonal antibody is applicable to enzyme-linked immunosorbent assay, immunoblot, immunofluorescence, and immunohistochemistry, and expected to be a valuable tool for detecting AIMP2-DX2 with high sensitivity and specificity for research and diagnostic purposes. [ABSTRACT FROM AUTHOR]