Reduction In Particle Production Research Articles

Mutations of Conserved Residues in the Major Homology Region Arrest Assembling HIV-1 Gag as a Membrane-Targeted Intermediate Containing Genomic RNA and Cellular Proteins.

ABSTRACTThe major homology region (MHR) is a highly conserved motif that is found within the Gag protein of all orthoretroviruses and some retrotransposons. While it is widely accepted that the MHR is critical for assembly of HIV-1 and other retroviruses, how the MHR functions and why it is so highly conserved are not understood. Moreover, consensus is lacking on when HIV-1 MHR residues function during assembly. Here, we first addressed previous conflicting reports by confirming that MHR deletion, like conserved MHR residue substitution, leads to a dramatic reduction in particle production in human and nonhuman primate cells expressing HIV-1 proviruses. Next, we used biochemical analyses and immunoelectron microscopy to demonstrate that conserved residues in the MHR are required after assembling Gag has associated with genomic RNA, recruited critical host factors involved in assembly, and targeted to the plasma membrane. The exact point of inhibition at the plasma membrane differed depending on the specific mutation, with one MHR mutant arrested as a membrane-associated intermediate that is stable upon high-salt treatment and other MHR mutants arrested as labile, membrane-associated intermediates. Finally, we observed the same assembly-defective phenotypes when the MHR deletion or conserved MHR residue substitutions were engineered into Gag from a subtype B, lab-adapted provirus or Gag from a subtype C primary isolate that was codon optimized. Together, our data support a model in which MHR residues act just after membrane targeting, with some MHR residues promoting stability and another promoting multimerization of the membrane-targeted assembling Gag oligomer.IMPORTANCE The retroviral Gag protein exhibits extensive amino acid sequence variation overall; however, one region of Gag, termed the major homology region, is conserved among all retroviruses and even some yeast retrotransposons, although the reason for this conservation remains poorly understood. Highly conserved residues in the major homology region are required for assembly of retroviruses; however, when these residues are required during assembly is not clear. Here, we used biochemical and electron microscopic analyses to demonstrate that these conserved residues function after assembling HIV-1 Gag has associated with genomic RNA, recruited critical host factors involved in assembly, and targeted to the plasma membrane but before Gag has completed the assembly process. By revealing precisely when conserved residues in the major homology region are required during assembly, these studies resolve existing controversies and set the stage for future experiments aimed at a more complete understanding of how the major homology region functions.

On the seawater temperature dependence of the sea spray aerosol generated by a continuous plunging jet

AbstractBreaking waves on the ocean surface produce bubbles which, upon bursting, deliver seawater constituents into the atmosphere as sea spray aerosol particles. One way of investigating this process in the laboratory is to generate a bubble plume by a continuous plunging jet. We performed a series of laboratory experiments to elucidate the role of seawater temperature on aerosol production from artificial seawater free from organic contamination using a plunging jet. The seawater temperature was varied from −1.3°C to 30.1°C, while the volume of air entrained by the jet, surface bubble size distributions, and size distribution of the aerosol particles produced was monitored. We observed that the volume of air entrained decreased as the seawater temperature was increased. The number of surface bubbles with film radius smaller than 2 mm decreased nonlinearly with seawater temperature. This decrease was coincident with a substantial reduction in particle production. The number concentrations of particles with dry diameter less than ∼1 μm decreased substantially as the seawater temperature was increased from −1.3°C to ∼9°C. With further increase in seawater temperature (up to 30°C), a small increase in the number concentration of larger particles (dry diameter >∼0.3 μm) was observed. Based on these observations, we infer that as seawater temperature increases, the process of bubble fragmentation changes, resulting in decreased air entrainment by the plunging jet, as well as the number of bubbles with film radius smaller than 2 mm. This again results in decreased particle production with increasing seawater temperature.

Vps4 and the ESCRT-III complex are required for the release of infectious hepatitis C virus particles.

The mechanisms by which infectious hepatitis C virus (HCV) particles are assembled and released from infected cells remain poorly characterized. In this regard, many other enveloped viruses, notably human immunodeficiency virus type 1, have been shown to utilize the host vacuolar protein sorting machinery (also known as the endosomal sorting complex required for transport; ESCRT) to traffic through the cell and effect the membrane rearrangements required for the formation of enveloped particles. We postulated that this might also apply to HCV. To test this hypothesis, we established a method of conditional virus-like particle assembly involving trans-complementation of an envelope-deleted JFH-1 genome using plasmid transfection. This system reliably produced virus particles that were infectious and could be enumerated easily by focus-forming assay in Huh7 cells. Following co-transfection with plasmids expressing various dominant-negative forms of either components of the ESCRT-III complex or Vps4 (the AAA ATPase that recycles the ESCRT complexes), a reduction in particle production was seen. No significant effect was observed after co-transfection of dominant-negative ESCRT-I or Alix, an ESCRT associated protein. Dominant-negative Vps4 or ESCRT-III components had no effect on either virus genome replication or the accumulation of intracellular infectious particles. These data were confirmed using cell culture infectious HCV and we conclude that HCV requires late components of the ESCRT pathway for release of infectious virus particles.

Multiple production in the Monte Carlo string fusion model

The influence of the string fusion on the basic features of relativistic heavy-ion collisions is studied with the help of a Monte Carlo string fusion model. The model is based on the parton picture of strong interactions and includes the string fusion phenomenon. It is demonstrated that string fusion leads to a considerable reduction of particle production with a relative enhancement of the baryon component in AA collisions at ultra-relativistic energies