Leader RNA of Rinderpest virus binds specifically with cellular La protein: a possible role in virus replication

Rinderpest virus (RPV) is an important member of the Morbillivirus genus in the family Paramyxoviridae and employs a similar strategy for transcription and replication of its genome as that of other negative sense RNA viruses. Cellular proteins have earlier been shown to stimulate viral RNA synthesis by isolated nucleocapsids from purified virus or from virus-infected cells. In the present work, we show that plus sense leader RNA of RPV, transcribed from 3' end of genomic RNA, specifically interacts with cellular La protein employing gel mobility shift assay as well as UV cross-linking of leader RNA with La protein. The leader RNA synthesized in virus-infected cells was shown to interact with La protein by immunoprecipitation of leader RNA bound to La protein and detecting the leader RNA in the immunoprecipitate by Northern hybridization with labeled antisense leader RNA. Employing a minireplicon system, we demonstrate that transiently expressed La protein enhances the replication/transcription of the RPV minigenome in cells. Sub-cellular immunolocalization shows that La protein is redistributed from nucleus to the cytoplasm upon infection. Our results strongly suggest that La protein may be involved in regulation of Rinderpest virus replication

Role of Polypyrimidine Tract Binding Protein in Mediating Internal Initiation of Translation of Interferon Regulatory Factor 2 RNA

BACKGROUND: Earlier we have reported translational control of interferon regulatory factor 2 (IRF2) by internal initiation (Dhar et al, Nucleic Acids Res, 2007). The results implied possible role of IRF2 in controlling the intricate balance of cellular gene expression under stress conditions in general. Here we have investigated the secondary structure of the Internal Ribosome Entry Site of IRF2 RNA and demonstrated the role of PTB protein in ribosome assembly to facilitate internal initiation. METHODOLOGY/PRINCIPAL FINDINGS: We have probed the putative secondary structure of the IRF2 5'UTR RNA using various enzymatic and chemical modification agents to constrain the secondary structure predicted from RNA folding algorithm Mfold. The IRES activity was found to be influenced by the interaction of trans-acting factor, polypyrimidine tract binding protein (PTB). Deletion of 25 nts from the 3'terminus of the 5'untranslated region resulted in reduced binding with PTB protein and also showed significant decrease in IRES activity compared to the wild type. We have also demonstrated putative contact points of PTB on the IRF2-5'UTR using primer extension inhibition assay. Majority of the PTB toe-prints were found to be restricted to the 3'end of the IRES. Additionally, Circular Dichroism (CD) spectra analysis suggested change in the conformation of the RNA upon PTB binding. Further, binding studies using S10 extract from HeLa cells, partially silenced for PTB gene expression, resulted in reduced binding by other trans-acting factors. Finally, we have demonstrated that addition of recombinant PTB enhances ribosome assembly on IRF2 IRES suggesting possible role of PTB in mediating internal initiation of translation of IRF2 RNA. CONCLUSION/SIGNIFICANCE: It appears that PTB binding to multiple sites within IRF2 5'UTR leads to a conformational change in the RNA that facilitate binding of other trans-acting factors to mediate internal initiation of translation

SARS-CoV-2 B.1.617.2 Delta variant replication and immune evasion

Abstract: The B.1.617.2 (Delta) variant of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) was first identified in the state of Maharashtra in late 2020 and spread throughout India, outcompeting pre-existing lineages including B.1.617.1 (Kappa) and B.1.1.7 (Alpha)1. In vitro, B.1.617.2 is sixfold less sensitive to serum neutralizing antibodies from recovered individuals, and eightfold less sensitive to vaccine-elicited antibodies, compared with wild-type Wuhan-1 bearing D614G. Serum neutralizing titres against B.1.617.2 were lower in ChAdOx1 vaccinees than in BNT162b2 vaccinees. B.1.617.2 spike pseudotyped viruses exhibited compromised sensitivity to monoclonal antibodies to the receptor-binding domain and the amino-terminal domain. B.1.617.2 demonstrated higher replication efficiency than B.1.1.7 in both airway organoid and human airway epithelial systems, associated with B.1.617.2 spike being in a predominantly cleaved state compared with B.1.1.7 spike. The B.1.617.2 spike protein was able to mediate highly efficient syncytium formation that was less sensitive to inhibition by neutralizing antibody, compared with that of wild-type spike. We also observed that B.1.617.2 had higher replication and spike-mediated entry than B.1.617.1, potentially explaining the B.1.617.2 dominance. In an analysis of more than 130 SARS-CoV-2-infected health care workers across three centres in India during a period of mixed lineage circulation, we observed reduced ChAdOx1 vaccine effectiveness against B.1.617.2 relative to non-B.1.617.2, with the caveat of possible residual confounding. Compromised vaccine efficacy against the highly fit and immune-evasive B.1.617.2 Delta variant warrants continued infection control measures in the post-vaccination era

Mitigating the Gateway Bottleneck via Transparent Cooperative Caching in Wireless Mesh Networks

Wireless mesh networks (WMNs) have been proposed to provide cheap, easily deployable and robust Internet access. The dominant Internet-bound traffic from clients causes a congestion bottleneck around the gateway, which can significantly limit the throughput of the WMN clients in accessing the Internet. In this paper, we present MeshCache, a transparent caching system for WMNs that exploits the locality in client Internet-bound traffic to mitigate the bottleneck effect at the gateway, thereby improving client perceived performance. MeshCache leverages the fact that a WMN typically spans a small geographic area and hence mesh routers are easily over-provisioned with CPU, memory, and disk storage, and extends the individual wireless mesh routers in a WMN with built-in content caching functionality. It then performs cooperative caching among the wireless mesh routers. We explore two architecture designs for MeshCache: (1) caching at every client access mesh router upon file download, and (2) caching at each mesh router along the route the Internet traffic travels, which requires breaking a single end-to-end transport connection into multiple single-hop transport connections along the route. We also leverage the abundant research results from cooperative web caching in the Internet in designing cache selection protocols for efficiently locating caches containing data objects for these two architectures. We further compare these two MeshCache designs with caching at the gateway router only. Through extensive simulations and evaluations using a prototype implementation on a testbed, we find that MeshCache can significantly improve the performance of client nodes in WMNs. In particular, our experiments with a Squid-based MeshCache implementation deployed on the MAP mesh network testbed with 15 routers show that compared to caching at the gateway only, the MeshCache architecture with hop-by-hop caching reduces the load at the gateway by 38%, improves the average client throughput by 170%, and increases the number of transfers that achieve a throughput greater than 1 Mbps by a factor of 3

Distributed Hashing for Scalable Multicast in Wireless Ad Hoc Networks

Several multicast protocols for mobile ad hoc networks (MANETs) have been proposed that build multicast trees using location information available from GPS or localization algorithms and use geographic forwarding to forward packets down the multicast trees. These stateless multicast protocols carry encoded membership, location and tree information in each packet. Stateless protocols are more efficient and robust than stateful protocols (ADMR, ODMRP) as they avoid the difficulty of maintaining distributed states in the presence of frequent topology changes in MANETs. However, stateless locationbased multicast protocols are not scalable to large groups because they encode group membership in the header of each data packet, i.e. they incur a per-packet encoding overhead. Additionally, such protocols involve centralized group membership and location management, either at the tree root or the traffic source. In this work, we present the Hierarchical Rendezvous Point Multicast (HRPM) protocol which significantly improves the scalability of stateless location-based multicast with respect to the group size. HRPM incorporates two key design ideas: (1) hierarchical decomposition of multicast groups, and (2) use of distributed geographic hashing to construct and maintain such a hierarchy efficiently. HRPM organizes a large group into a hierarchy of recursively organized manageable-sized subgroups in an effort to reduce per-packet encoding overhead. More importantly, HRPM constructs and maintains this hierarchy at virtually no cost using distributed hashing; distributed hashing is recursively applied at each subgroup for group management and avoids the potentially high cost associated with maintaining distributed state at mobile nodes. The hierarchical organization and the distributed hashing property also allows HRPM to scale to large networks and large numbers of groups. Performance results obtained via detailed simulations demonstrate that HRPM achieves enhanced scalability and performance. Coupled with its leverage of stateless geographic forwarding, HRPM scales well in terms of the group size, the number of groups, the number of sources, as well as the size of the network. In particular, HRPM maintains close to 95% multicast delivery ratio while incurring on average 5.5% per packet tree-encoding overhead for up to 250 group members in a 500-node network. Furthermore, it achieves a steady 95% delivery ratio while incurring nearly constant overhead as the number of groups increases from 2 to 45, while keeping the total number of receivers constant at 180, in a 500-node network. Lastly, it steadily achieves above 90% delivery ratio as the network scales up to 1000 nodes with up to 30% group members. As a reference, we also compared HRPM to ODMRP, a state-of-the-art topology-based multicast protocol that is scalable to large groups. HRPM performs comparably to ODMRP across a wide range of group sizes. More over, HRPM outperforms ODMRP when the network size, the number of groups, or the number of sources increases

DynCoDe: An Architecture for Transparent Dynamic Content Delivery

Delivery of web content is increasingly using dynamic and personalized content. Caching has been extensively studied for reducing the client latency and bandwidth requirements for static content. There has been recent interest in schemes to exploit locality in dynamic web content [1, 2]. We propose a novel scheme that integrates the distribution and caching of personalized content which rely heavily on dynamic generation of content. In the proposed architecture, resource intensive processes involved in content generation are pushed to the network edges. We have performed a preliminary evaluation of the architecture under real world network conditions and have noticed significant improvements in bandwidth consumption, user response time and server scalability showing the feasibility of such a scheme

How to Implement DHTs in Mobile Ad Hoc Networks?

and of Pastry routing state. However, a straightforward layering is not pragmatic, and three modifications are made to accommodate the shared medium access nature of MANETs: (1) Pastry's node joining process is modified to use expanding ring search for locating a bootstrap node to join the network; (2) The original Pastry uses an expensive "ping" mechanism with a delay metric to measure and maintain the proximity of nodes in its routing tables. Since delay is affected by many factors and has a high variability in MANETs, we modified Pastry to use a hop count metric for proximity; (3) To reduce the cost of this proximity probing, we modified DSR to export an API that allows Pastry to inquire about the proximity values for nodes it is interested in. DSR can then use its cache to reply to "pings" from Pastry if there is a cached path to the node being pinged. In the absence of such a cached path, a ROUTE REQUEST is initiated by DSR. In summary, the layered design is similar to implemen

DMesh: Incorporating Practical Directional Antennas in Multi-Channel Wireless Mesh Networks

Wireless mesh networks (WMNs) have been proposed as an effective solution for ubiquitous lastmile broadband access. Three key factors that affect the usability of WMNs are high throughput, costeffectiveness and ease of deployability. Recent research has focused on increasing WMN throughput assuming the use of multiple radios equipped with omnidirectional antennas accompanied with channel assignment to enable frequency separation between contending transmissions. Compared to omni antennas, directional antennas offer spatial separation between contending transmissions and have the potential to further enhance the throughput of WMNs. In this paper, we propose DMesh, a WMN architecture that combines spatial separation from directional antennas with frequency separation from orthogonal channels to improve the throughput of WMNs. An important requirement in DMesh is to accomplish this throughput improvement without inhibiting the other two key WMN requirements: cost-effectiveness and ease of deployability. The high cost of smart beamforming directional antennas and their form factor make it difficult to achieve these two requirements. Thus, in DMesh, we focus our effort on incorporating practical directional antennas that are widely and cheaply available (e.g. patch and yagi). The key challenge in DMesh is to exploit spatial separation from such practical directional antennas despite their lack of electronic steerability and interference nulling as well as the presence of significant sidelobes and backlobes. In this paper, we study how such practical directional antennas can improve the throughput of a WMN. Central to our architecture is a distributed, directional channel assignment algorithm for mesh routers that effectively exploits the spatial and frequency separation opportunities in a DMesh network. Simulation results show that DMesh improves the throughput of WMNs by up to 231% and reduces packet delay drastically compared to a multi-radio multi-channel omni network. A DMesh implementation in our 802.11b WMN testbed using commercially available practical directional antennas provides TCP throughput gains ranging from 31% to 57%

On optimal TTL sequence-based route discovery in MANETs

In on-demand multi-hop routing protocols for MANETs such as DSR and AODV, a fundamental requirement for peer-to-peer connectivity is to discover routes to a remote node via flooding of route request messages. Historically, such floodings of requests have used a TTL (time-to-live) large enough to reach all nodes in the network to ensure successful route discovery in one round of flooding. Recently [1], it was shown that the generic minimal cost flooding search problem can be solved via a sequence of floodings with an optimally chosen set of TTLs. The theoretical result, when applied to DSR route discovery, does not take into account optimizations such as route caching and overhearing, which can significantly reduce the frequency and the propagation range of route discovery operations. Equally importantly, the impact of using a sequence of floodings on the packet delivery delay is not clear. In this paper, we study the impact of using the optimal TTL sequence-based route discovery on DSR routing performance. Our results show when caching and overhearing are considered, the route discovery enhanced by an optimal TTL sequence has very similar overhead but higher delay than the basic route discovery mechanism.