Characterization of aerobic granular sludge developed under variable and low organic loading rate

Understanding the formation of aerobic granules sludge (AGS) under the variations of organic loading rate (OLR) could give a different insight on AGS stability, which had become the bottleneck for practical application in sewage treatment. This study demonstrates the formation of AGS that had previously been stored for eight months at 5ºC in sequencing batch reactor (SBR) with sewage as substrate. Despite being redeveloped under variable OLR of 0.26 to 0.81 kg CODs/m3 d and low superficial air velocity (SAV) of 1.33 cm/s, the loose structure of AGS during storage can be recovered within 46 days of formation process. Variations in OLR intrude the formation process particularly during low OLR, resulting in longer period to achieve mature AGS or full granulation of biomass in reactor. The next-generation sequencing (NGS) analysis indicated that the shift in microbial community from Rhodocyclaceae to Comamonadaceae class for denitrification process was accommodated with the changes in the AGS size from 326 μm to more than 600 μm

The roles of ribosomal proteins in nasopharyngeal cancer: culprits, sentinels or both

Ribosomal protein genes encode products that are essential for cellular protein biosynthesis and are major components of ribosomes. Canonically, they are involved in the complex system of ribosome biogenesis pivotal to the catalysis of protein translation. Amid this tightly organised process, some ribosomal proteins have unique spatial and temporal physiological activity giving rise to their extra-ribosomal functions. Many of these extra-ribosomal roles pertain to cellular growth and differentiation, thus implicating the involvement of some ribosomal proteins in organogenesis. Consequently, dysregulated functions of these ribosomal proteins could be linked to oncogenesis or neoplastic transformation of human cells. Their suspected roles in carcinogenesis have been reported but not specifically explained for malignancy of the nasopharynx. This is despite the fact that literature since one and half decade ago have documented the association of ribosomal proteins to nasopharyngeal cancer. In this review, we explain the association and contribution of dysregulated expression among a subset of ribosomal proteins to nasopharyngeal oncogenesis. The relationship of these ribosomal proteins with the cancer are explained. We provide information to indicate that the dysfunctional extra-ribosomal activities of specific ribosomal proteins are tightly involved with the molecular pathogenesis of nasopharyngeal cancer albeit mechanisms yet to be precisely defined. The complete knowledge of this will impact future applications in the effective management of nasopharyngeal cancer

Inhibition of lysosomal vacuolar proton pump down-regulates cellular acidification and enhances E. coli bactofection efficiency

Endosomal escape is considered a crucial barrier that needs to be overcome by integrin-mediated E. coli for gene delivery into mammalian cells. Bafilomycin, a potent inhibitor of the H+ proton pump commonly employed to lower endosomal pH, was evaluated as part of the E. coli protocol during delivery. We found an increase in green fluorescent protein expression up 6.9, 3.2, 5.0, 2.8, and 4.5 fold in HeLa, HEK-293, A549, HT1080, and MCF-7 respectively, compared to untreated cells. Our result showed for the first time that Inhibition of lysosomal VATPase enhances E. coli efficiency

Multigenic prognosis assessment model for nasopharyngeal carcinoma via a modified meta-analysis approach

Objectives Currently, clinically relevant multigene-based prognostic assessment models for nasopharyngeal carcinoma (NPC) are limited. This paper reports a novel NPC prognosis assessment model based on multiple established NPC-associated biomarkers. Methods We used a modified meta-analysis approach to retrieve eligible studies and analyse the data. Different prognostic biomarkers and hazard ratios (HRs) with 95 % confidence intervals (CIs) of overall survival (OS) data were extracted and tabulated from eligible studies. We then used the formula based on Parmar et al. to determine OS (expressed as HR with 95 % CI). Prognosis assessment risk scores assigned to the logarithm of HR were the basis for interpreting the multigene prognosis assessment model. Finally, we explained the biological significance of this model using a multigenic NPC oncogenesis network system. Results We constructed a multigenic NPC prognosis assessment model consisting of 10 prognostic biomarkers to determine the OS rate in NPC patients. Based on the biomarkers’ expression patterns, the model could determine 1,023 possible OS rates of NPC patients. The risk score derived determines the prognosis status of the NPC patients. The higher the total risk assessment score, the poorer the prognosis. An NPC-associated network involving all ten biomarkers was also derived. Conclusions We provided a novel multigenic NPC prognosis assessment model comprising ten prognostic biomarkers on OS rate in NPC patients. A conceptual molecular-based pathophysiological network of NPC oncogenesis supported the biological relevance of this model

Human Ribosomal Proteins RPeL27, RPeL43 and RPeL41 are Up-regulated in Nasopharyngeal Carcinoma Cell Lines

Apart from their canonical role in ribosome biogenesis, there is increasing evidence of ribosomal protein genes’ involvement in various cancers. A previous study by us revealed significant differential expression of three ribosomal protein genes (RPeL27, RPeL41 and RPeL43) between cell lines derived from tumor and normal nasopharyngeal epithelium. However, the results therein were based on a semi-quantitative assay, thus preliminary in nature. Herein, we provide findings of a deeper analysis of these three genes in the context to nasopharyngeal carcinoma (NPC) tumorigenesis. Their expression patterns were analyzed in a more quantitative manner at transcript level. Their protein expression levels were also investigated. We showed results that are contrary to previous report. Rather than down-regulation, these genes were significantly overexpressed in NPC cell lines compared to normal control at both transcript and protein levels. Nevertheless, their association with NPC has been established. Immunoprecipitation pulldown assays indicate the plausible interaction of either RPeL27 or RPeL43 with POTEE/TUBA1A and ACTB/ACTBL2 complexes. In addition, RPeL43 is shown to bind with MRAS and EIF2S1 proteins in a NPC cell line (HK1). Our findings support RPeL27, RPeL41 and RPeL43 as potential markers of NPC, and provide insights into the interaction targets of RPeL27 and RPeL43 proteins

RFP-based method for real-time tracking of invasive bacteria in a heterogeneous population of cells

Quantification of bacterial invasion into eukaryotic cells is a prerequisite to unfold the molecular mechanisms of this vector’s function to obtain insights for improving its efficiency. Invasion is traditionally quantified by antibiotic protection assays that require dilution plating and counting of colony-forming units rescued from infected cells. However, to differentiate between attached and internalized bacteria vector, this assay requires supplementation by a time-consuming and tedious immunofluorescence staining, making it laborious and reduces its reliability and reproducibility. Here we describe a new red fluorescent protein (RFP)-based highthroughput and inexpensive method for tracking bacterial adherence and internalization through flow cytometry to provide a convenient and real-time quantification of bacterial invasiveness in a heterogeneous population of cells. We invaded MCF-7, A549, and HEK-293 cells with the E. coli vector and measured RFP using imaging flow cytometry. We found high cellular infection of up to 70.47% in MCF-7 compared to 27.4% and 26.2% in A549 and HEK-293 cells, respectively. The quantitative evaluation of internalized E. coli is rapid and celldependent, and it distinctively differentiates between attached and cytosolic bacteria while showing the degree of cellular invasiveness. This imaging flow cytometry approach can be applied broadly to study host bacteria interaction

Differential expression of a subset of ribosomal protein genes in cell lines derived from human nasopharyngeal epithelium

Extraribosomal functions of human ribosomal proteins (RPs) include the regulation of cellular growth and differentiation, and are inferred from studies that linked congenital disorders and cancer to the deregulated expression of RP genes. We have previously shown the upregulation and downregulation of RP genes in tumors of colorectal and nasopharyngeal carcinomas (NPCs), respectively. Herein, we show that a subset of RP genes for the large ribosomal subunit is differentially expressed among cell lines derived from the human nasopharyngeal epithelium. Three such genes (RPL27, RPL37a and RPL41) were found to be significantly downregulated in all cell lines derived from NPC tissues compared with a nonmalignant nasopharyngeal epithelial cell line. The expression of RPL37a and RPL41 genes in human nasopharyngeal tissues has not been reported previously. Our findings support earlier suspicions on the existence of NPC-associated RP genes, and indicate their importance in human nasopharyngeal organogenesis

Short homologies efficiently generate detectable homologous recombination events

When recombineering bacterial artificial chromosomes (BACs), it is common practice to design the ends of the donor molecule with 50 bp of homology specifying its insertion site. We demonstrate that desired recombinants can be produced using intermolecular homologies as short as 15 bp. Although the use of shorter donor end regions decreases total recombinants by several fold, the frequency of recombinants with correctly inserted donor molecules was high enough for easy detection by simple polymerase chain reaction (PCR) screening. This observation may have important implications for the design of oligonucleotides for recombineering, including significant cost savings, especially for high-throughput projects that use large quantities of primers

Predicted interaction of human Ribosomal Protein S15 with Fragile X Mental Retardation Protein

In addition to the central role of ribosome biogenesis, the human ribosomal protein S15 (RPS15) has extra-ribosomal roles that include its association with a congenital disease and a few types of cancer. However, current knowledge of its functions in the context of extra-ribosomal activities remains fragmented. An approach to gain insights into the interaction between RPS15 and possible protein partners is via Bioinformatics strategies. Based on the sequence-to-structure-to-function paradigm, structural data of a protein can be computationally analysed to derive logical interacting partners. This method can include three-dimensional model construction, structural neighbour prediction, and molecular docking analysis. By using this approach, we have constructed RPS15 3D-structural models that have allowed the prediction of 23 structural neighbours. Of these, two that are from human origin were further analysed and only one have logical prospect of binary protein-protein interactions. Further analysis of this structural neighbour revealed 7 candidate docking partners. From these, our molecular docking analysis demonstrated two most logical dock models of interactions between RPS15 with two different domains of the Fragile X Mental Retardation Protein 1 (FMRP1) protein. Hence, we have provided in silico evidence of de novo protein-protein interaction between RPS15 and the Fragile X Mental Retardation Protein 1 (FMRP1)

A Self-Replicating Linear DNA

TelN and tos are a unique DNA linearization unit isolated from bacteriophage N15. While being transferable, the TelN cleaving-rejoining activities remained stable to function on tos in both bacterial and mammalian environments. However, TelN contribution in linear plasmid replication in mammalian cells remains unknown. Herein, we investigated the association of TelN in linear tos-containing DNA (tos-DNA) replication in mammalian cells. Additionally, the mammalian origin of replication (ori) that is well-known to initiate the replication event of plasmid vectors was also studied. In doing so, we identified that both TelN and mammalian initiation sites were essential for the replication of linear tos-DNA, determined by using methylation sensitive DpnI/MboI digestion and polymerase chain reaction (PCR) amplification approaches. Furthermore, we engineered the linear tos-DNA to be able to retain in mammalian cells using S/MAR technology. The resulting S/MAR containing tos-DNA was robust for at least 15 days, with (1) continuous tos-DNA replication, (2) correct splicing of gene transcripts, and (3) stable exogenous gene expression that was statistically comparable to the endogenous gene expression level. Understanding the activities of TelN and tos in mammalian cells can potentially provide insights for adapting this simple DNA linearization unit in developing novel genetic engineering tools, especially to the eukaryotic telomere/telomerase study