Welcome To Website IAS

Hot news
Achievement

Independence Award

- First Rank - Second Rank - Third Rank

Labour Award

- First Rank - Second Rank -Third Rank

National Award

 - Study on food stuff for animal(2005)

 - Study on rice breeding for export and domestic consumption(2005)

VIFOTEC Award

- Hybrid Maize by Single Cross V2002 (2003)

- Tomato Grafting to Manage Ralstonia Disease(2005)

- Cassava variety KM140(2010)

Centres
Website links
Vietnamese calendar
Library
Visitors summary
 Curently online :  1
 Total visitors :  4165563

Watching a virus grow
Thursday, 2019/11/07 | 08:18:08

Bogdan Dragnea

PNAS November 5, 2019 116 (45) 22420-22422

Figure: Cartoon representation of intermediates specific to 4 proposed mechanisms for icosahedral ssRNA virus growth. (A) Nucleation−elongation model (79). (B) Micellar condensation (“en masse”) (10). (C) RNA antenna model (11). (D) PS-mediated sequential assembly model (14). Arrows indicate impending binding events, vivid hues indicate interfaces activated for binding, and subdued hues represent subunits stabilized in postassembly conformations. CP, coat protein.

 

One of the most powerful molecular motors discovered, to date, belongs to a phage, a type of virus that infects bacteria (1). The motor serves to package stiff, double-stranded DNA (dsDNA) into a preassembled, proteinaceous polyhedral container called the procapsid. The DNA is about 50 times longer than the size of the capsid and gets packaged and compressed by the motor at near-crystalline densities. The capsid holds up its end of the bargain, withstanding positive pressure differentials of up to 40 atm (2). The level of current understanding of this fascinating biological process was made possible, in great measure, by the development of in singulo approaches to packaging dynamics, adapted to phages. However, what works for one virus may hardly apply to another. Thus, in a great number of single-stranded RNA (ssRNA) viruses—the largest virus group on Earth—nucleic acid packaging occurs concurrently with, and indeed is part of, spontaneous assembly. No ATP-powered motor is involved here. Instead of great positive pressures, the final pressure exerted by the RNA, at equilibrium, is modest and negative (3). Disentangling the interplay between the 2 types of concurrent driving interactions—those among coat proteins and those between coat proteins and RNA—is far from trivial. Adding to the challenge, intermediates have a fleeting existence (milliseconds), while the sizes of ssRNA viruses that can assemble in vitro are small (tens of nanometers). This is why, up to now, the principal features of ssRNA virus assembly have been mainly supplied by theoretical models, structural arguments from static data, and ensemble-averaged dynamic experiments. In singulo methods that would offer the same level of real-time dynamics detail and direct insight as those applied to dsDNA phages 2 decades ago have been scarce. In PNAS, Garmann et al. (4) provide a compelling demonstration of the potential held by a wide-field optical microscopy method for tightly controlled, real-time single-particle studies of the ssRNA bacteriophage MS2 assembly with near-molecular accuracy and broad temporal dynamic range.

 

To measure assembly kinetics in real time, Garmann et al. (4) adapt an imaging technique called interferometric scattering microscopy (5). In their approach, sparsely distributed single molecules of viral RNA are tethered via flexible linkers to the surface of a microscope coverslip, in solution. Upon injection of virus coat proteins, RNA−protein association results in an increase of the effective local protein density at tethered RNA locations. Since the optical polarizability of proteins is different from that of water, there is increased light scattering by the growing nucleoprotein complex. The scattered light is phase-shifted with respect to the incident light. This phase shift provides a way to boost contrast at the detector plane, where the coherent scattered light is made to interfere with light reflected by the coverslip. Thus, multiple growing virus particles can be imaged in parallel via a charge-coupled device in real time, as diffraction-limited spots of an intensity proportional to the local mass accumulation. The reported detection limit is 6 coat-protein dimers at 1-Hz bandwidth.

 

MS2 has a capsid formed of 180 identical proteins organized with icosahedral symmetry as 12 pentamers and 20 hexamers. A single genomic RNA molecule, 3,569 nucleotides long, is encapsulated inside the capsid. Like other ssRNA viruses, the genome does not occupy uniformly the entire available space. Instead, it forms 2 concentric layers adjacent to the luminal capsid surface and has relatively low density near center. However, while the genome of other simple T = 3 ssRNA viruses, such as the well-studied cowpea chlorotic mottle virus (CCMV), is smeared nearly isotropically across the lumenal surface, genome distribution in MS2 is anisotropic and correlates with capsid geometry (6). This difference in genome organization helped formulate the hypothesis that genome packaging during MS2 assembly may be guided by different constraints and therefore may follow qualitatively different pathways from those proposed for other simple viruses. For instance, 3 possible models exist for the broad class of icosahedral viruses represented by CCMV (Fig. 1 AC). The coat proteins of this class will readily assemble and form capsids in vitro around heterologous nucleic acids, other polyanions, and even nanoparticles, and thus the first driving interactions taken into consideration in building these models were nonspecific.

 

The size of the nucleus could not be estimated, since it appears to be smaller than the detection limit, that is, about 6 dimers (dimers, not monomers, are the more stable coat-protein state in solution, prior to assembly) (4). The findings are intriguing because the number of putative PS sites should be greater than that (14), yet it appears that the initiation step corresponding to proteins binding at PS sites could not be detected by the interferometric scattering approach. This could be due to a number of reasons: drift in the background intensity, and/or differences in the buffer ionic strength and protein concentration with respect to previous work (15). In those previous studies, protein concentration was in the nanomolar range, and total assembly times exceeded 103 s, which would be challenging to achieve with interferometric scattering, due to drift. In addition, lower ionic strength in ref. 15 favored stronger RNA−coat-protein interactions. Could it be that, like other viruses, MS2 is able to switch between mechanisms, for example, depending on the stage of infection? Might it be that PS activity depends on the kinetics of the nucleoprotein complex folding with PS sites fading in and out of existence through a slow ramping of protein concentration, as Dykeman et al. (16) have suggested? What would in singulo kinetics look like if the RNA of a different virus were used instead of MS2 RNA? What instrumentation improvements could lower the detection limit of interferometric scattering microscopy to the point where the nature of the critical nucleus could be determined? The path ahead is wide open and promises to add to an already fascinating story.

 

See https://www.pnas.org/content/116/45/22420

Back      Print      View: 24

[ Other News ]___________________________________________________
  • Egypt Holds Workshop on New Biotech Applications
  • UN Agencies Urge Transformation of Food Systems
  • Taiwan strongly supports management of brown planthopper—a major threat to rice production
  • IRRI Director General enjoins ASEAN states to invest in science for global food security
  • Rabies: Educate, vaccinate and eliminate
  • “As a wife I will help, manage, and love”: The value of qualitative research in understanding land tenure and gender in Ghana
  • CIP Director General Wells Reflects on CIP’s 45th Anniversary
  • Setting the record straight on oil palm and peat in SE Asia
  • Why insect pests love monocultures, and how plant diversity could change that
  • Researchers Modify Yeast to Show How Plants Respond to Auxin
  • GM Maize MIR162 Harvested in Large Scale Field Trial in Vinh Phuc, Vietnam
  • Conference Tackles Legal Obligations and Compensation on Biosafety Regulations in Vietnam
  • Iloilo Stakeholders Informed about New Biosafety Regulations in PH
  • Global wheat and rice harvests poised to set new record
  • GM Maize Harvested in Vietnam Field Trial Sites
  • New label for mountain products puts premium on biological and cultural diversity
  • The Nobel Prize in Physiology or Medicine 2016
  • Shalabh Dixit: The link between rice genes and rice farmers
  • People need affordable food, but prices must provide decent livelihoods for small-scale family farmers
  • GM Seeds Market Growth to Increase through 2020 Due to Rise in Biofuels Use
Designed & Powered by WEBSO CO.,LTD