Movement Protein Pns6 of Rice dwarf phytoreovirus Has Both ATPase and RNA Binding Activities

Cell-to-cell movement is essential for plant viruses to systemically infect host plants. Plant viruses encode movement proteins (MP) to facilitate such movement. Unlike the well-characterized MPs of DNA viruses and single-stranded RNA (ssRNA) viruses, knowledge of the functional mechanisms of MPs encoded by double-stranded RNA (dsRNA) viruses is very limited. In particular, many studied MPs of DNA and ssRNA viruses bind non-specifically ssRNAs, leading to models in which ribonucleoprotein complexes (RNPs) move from cell to cell. Thus, it will be of special interest to determine whether MPs of dsRNA viruses interact with genomic dsRNAs or their derivative sRNAs. To this end, we studied the biochemical functions of MP Pns6 of Rice dwarf phytoreovirus (RDV), a member of Phytoreovirus that contains a 12-segmented dsRNA genome. We report here that Pns6 binds both dsRNAs and ssRNAs. Intriguingly, Pns6 exhibits non-sequence specificity for dsRNA but shows preference for ssRNA sequences derived from the conserved genomic 5′- and 3′- terminal consensus sequences of RDV. Furthermore, Pns6 exhibits magnesium-dependent ATPase activities. Mutagenesis identified the RNA binding and ATPase activity sites of Pns6 at the N- and C-termini, respectively. Our results uncovered the novel property of a viral MP in differentially recognizing dsRNA and ssRNA and establish a biochemical basis to enable further studies on the mechanisms of dsRNA viral MP functions

Advances in tissue engineering through stem cell-based co-culture

Stem cells are the future in tissue engineering and regeneration. In a co-culture, stem cells not only provide a target cell source with multipotent differentiation capacity, but can also act as assisting cells that promote tissue homeostasis, metabolism, growth and repair. Their incorporation into co-culture systems seems to be important in the creation of complex tissues or organs. In this review, critical aspects of stem cell use in co-culture systems are discussed. Direct and indirect co-culture methodologies used in tissue engineering are described, along with various characteristics of cellular interactions in these systems. Direct cell–cell contact, cell–extracellular matrix interaction and signalling via soluble factors are presented. The advantages of stem cell co-culture strategies and their applications in tissue engineering and regenerative medicine are portrayed through specific examples for several tissues, including orthopaedic soft tissues, bone, heart, vasculature, lung, kidney, liver and nerve. A concise review of the progress and the lessons learned are provided, with a focus on recent developments and their implications. It is hoped that knowledge developed from one tissue can be translated to other tissues. Finally, we address challenges in tissue engineering and regenerative medicine that can potentially be overcome via employing strategies for stem cel

Deregulation of eIF4E: 4E-BP1 in Differentiated Human Papillomavirus-Containing Cells Leads to High Levels of Expression of the E7 Oncoprotein

Infections with high-risk human papillomaviruses (HPVs) are linked to more than 95% of cervical cancers. HPVs replicate exclusively in differentiated cells and the function of the HPV E7 oncoprotein is essential for viral replication. In this study, we investigated the mechanism that regulates E7 expression in differentiated cells. The level of E7 protein was strongly induced in HPV-containing Caski, HOK-16B, and BaP-T cells during growth in methylcellulose-containing medium, a condition that induces differentiation. Enhanced expression of E7 was observed between 4 and 8 h of culturing in methylcellulose and was maintained for up to 24 h. The increase was not due to altered stability of the E7 protein or an increase in the steady-state level of the E7 mRNA. Instead, the translation of the E7 mRNA was enhanced during differentiation. More than 70 to 80% of the E7 mRNA was found in the polysome fractions in the differentiated cells. Consistent with this observation, higher levels of the phosphorylated translator inhibitor 4E-BP1 were observed in differentiated HPV-containing cells but not in differentiated non-HPV tumor cells or primary keratinocytes. The mTOR kinase inhibitor rapamycin blocked phosphorylation of 4E-BP1 and significantly decreased the level of E7 protein in Caski cells, suggesting that phosphorylation of 4E-BP1 is linked to E7 expression. Prevailing models for the molecular mechanisms underlying E7 expression have focused largely on transcriptional regulation. The results presented in this study demonstrate a significant role of the cellular translation machinery to maintain a high level of E7 protein in differentiated cells