HIV-1 Nef: a multifaceted modulator of T cell receptor signaling

Nef, an accessory protein of the Human Immunodeficiency Virus type 1 (HIV-1), is dispensable for viral replication in cell culture, but promotes virus replication and pathogenesis in the infected host. Acting as protein-interaction adaptor, HIV-1 Nef modulates numerous target cell activities including cell surface receptor expression, cytoskeletal remodeling, vesicular transport, and signal transduction. In infected T-lymphocytes, altering T-cell antigen receptor (TCR) signaling has long been recognized as one key function of the viral protein. However, reported effects of Nef range from inhibition to activation of this cascade. Recent advances in the field begin to explain these seemingly contradictory observations and suggest that Nef alters intracellular trafficking of TCR proximal machinery to disrupt plasma membrane bound TCR signaling while at the same time, the viral protein induces localized signal transduction at the trans-Golgi network. This review summarizes these new findings on how HIV-1 Nef reprograms TCR signalling output from a broad response to selective activation of the RAS-Erk pathway. We also discuss the implications of these alterations in the context of HIV-1 infection and in light of current concepts of TCR signal transduction

How HIV Takes Advantage of the Cytoskeleton in Entry and Replication

The host cell cytoskeleton plays a key role in the life cycle of viral pathogens whose propagation depends on mandatory intracellular steps. Accordingly, also the human immunodeficiency virus type 1 (HIV-1) has evolved strategies to exploit and modulate in particular the actin cytoskeleton for its purposes. This review will recapitulate recent findings on how HIV-1 hijacks the cytoskeleton to facilitate entry into, transport within and egress from host cells as well as to commandeer communication of infected with uninfected bystander cells

Cell motility through plasma membrane blebbing

Plasma membrane blebs are dynamic cytoskeleton-regulated cell protrusions that have been implicated in apoptosis, cytokinesis, and cell movement. Influencing Rho–guanosine triphosphatase activities and subsequent actomyosin dynamics appears to constitute a core component for bleb formation. In this paper, we discuss recent evidence in support of a central role of nonapoptotic membrane blebbing for cell migration and cancer cell invasion as well as advances in our understanding of the underlying molecular mechanisms. Based on these studies, we propose that in a physiological context, bleb-associated cell motility reflects a cell's response to reduced substratum adhesion. The importance of blebbing as a functional protrusion is underscored by the existence of multiple molecular mechanisms that govern actin-mediated bleb retraction

Live and Let Die Nef Functions beyond HIV Replication

AbstractThe viral Nef protein is important for the progression of the human and simian immunodeficiency virus (HIV/SIV) infection. So far, experimental evidence has suggested that Nef enhances viral replication and infectivity through a combination of different effects. Recent insights, however, indicate that its functions are more complex than previously anticipated. By targeting the T cell receptor, Nef may not only prime viral replication but, more importantly, ensure viral survival through distinct mechanisms of immune evasion and antiapoptosis

The Nef Protein of Human Immunodeficiency Virus Establishes Superinfection Immunity by a Dual Strategy to Downregulate Cell-Surface CCR5 and CD4

SummaryBackground: Viruses frequently render cells refractory to subsequent infection with the same virus. This state of superinfection immunity counteracts potentially detrimental consequences for the infected cell and facilitates high-level replication and viral spread in the host.Results: Here, we show that human immunodeficiency virus (HIV) employs its early gene product Nef to efficiently interfere with superinfection at the viral-entry step. In this context, we identify the downregulation of cell-surface CCR5, the major HIV coreceptor, as a novel and highly conserved activity of Nef. Nef targets the CCR5 coreceptor and the HIV binding receptor CD4 via distinct cellular machineries to enhance the endocytosis rate of both HIV receptor components and to accelerate their degradation. Functionally, these genetically separable actions by Nef synergized to efficiently protect cells from HIV superinfection at the level of fusion of the viral envelope with the plasma membrane.Conclusions: HIV has evolved two independent activities for Nef to downregulate the receptor complex and to facilitate its efficient replication and spread. This evasion strategy likely represents a mechanism by which the pathogenicity factor Nef elevates viral replication in vivo and thus promotes AIDS pathogenesis

Role of the C-terminal domain of the HIV-1 glycoprotein in cell-to-cell viral transmission between T lymphocytes

<p>Abstract</p> <p>Background</p> <p>Mutant HIV (HIV-Env-Tr712) lacking the cytoplasmic tail of the viral glycoprotein (Env-CT) exhibits a cell-type specific replication phenotype such that replicative spread occurs in some T-cell lines (referred to as permissive cells) but fails to do so in most T-cell lines or in PBMCs (referred to as non-permissive cells). We aim to gain insight on the underlying requirement for the Env-CT for viral spread in non-permissive cells.</p> <p>Results</p> <p>We established that in comparison to HIV-Wt, both cell-free and cell-to-cell transmission of mutant HIV-Env-Tr712 from non-permissive cells were severely impaired under naturally low infection conditions. This requirement for Env-CT could be largely overcome by using saturating amounts of virus for infection. We further observed that in permissive cells, which supported both routes of mutant virus transmission, viral gene expression levels, Gag processing and particle release were inherently higher than in non-permissive cells, a factor which may be significantly contributing to their permissivity phenotype. Additionally, and correlating with viral transfer efficiencies in these cell types, HIV-Gag accumulation at the virological synapse (VS) was reduced to background levels in the absence of the Env-CT in conjugates of non-permissive cells but not in permissive cells.</p> <p>Conclusions</p> <p>During natural infection conditions, the HIV-Env-CT is critically required for viral transmission in cultures of non-permissive cells by both cell-free and cell-to-cell routes and is instrumental for Gag accumulation to the VS. The requirement of the Env-CT for these related processes is abrogated in permissive cells, which exhibit higher HIV gene expression levels.</p

β-TrCP is dispensable for Vpu's ability to overcome the CD317/Tetherin-imposed restriction to HIV-1 release

<p>Abstract</p> <p>Background</p> <p>The cellular transmembrane protein CD317/BST-2/HM1.24/Tetherin restricts HIV-1 infection by physically tethering mature virions to the surface of infected cells. HIV-1 counteracts this restriction by expressing the accessory protein Vpu, yet the mechanism of this antagonism is incompletely understood. β-TrCP is the substrate recognition domain of an E3 ubiquitin ligase complex that interacts with the di-serine motif S52/S56 in the cytoplasmic tail of Vpu to target the CD4 receptor for proteasomal degradation. Recently, it has been suggested that β-TrCP is also critically involved in Vpu's ability to overcome the CD317-mediated virion release block.</p> <p>Results</p> <p>To test this model, we analyzed the consequences of several experimental strategies to interfere with the Vpu-β-TrCP protein-protein interaction. Under these conditions, we studied effects of Vpu on expression and localization of CD317 and CD4, as well as on its ability to promote HIV-1 release. Our results demonstrate a strict requirement for Vpu's di-serine motif for degradation of CD4 and also CD317, reduction of cell surface exposure of CD317, and HIV-1 release enhancement. We further show a critical role of β-TrCP2, but not of the structurally related β-TrCP1 isoform, for Vpu-mediated degradation of both receptors. Most importantly, Vpu remained active in downregulating CD317 from the cell surface and in overcoming the HIV-1 release restriction in β-TrCP-depleted cells.</p> <p>Conclusions</p> <p>These results demonstrate that β-TrCP is not strictly required for Vpu's ability to counteract the CD317-imposed virion release block and support the relevance of cell surface down-modulation of the restriction factor as a central mechanism of Vpu antagonism. Moreover, we propose the existence of a critical, yet to be identified cellular factor that interacts with Vpu via its di-serine motif to alter the trafficking of the restriction factor.</p

The Human Formin FHOD1 Contains a Bipartite Structure of FH3 and GTPase-Binding Domains Required for Activation

SummaryFormins induce the nucleation and polymerization of unbranched actin filaments. They share three homology domains required for profilin binding, actin polymerization, and regulation. Diaphanous-related formins (DRFs) are activated by GTPases of the Rho/Rac family, whose interaction with the N-terminal formin domain is thought to displace a C-terminal Diaphanous-autoregulatory domain (DAD). We have determined the structure of the N-terminal domains of FHOD1 consisting of a GTPase-binding domain (GBD) and the DAD-recognition domain FH3. In contrast to the formin mDia1, the FHOD1-GBD reveals a ubiquitin superfold as found similarly in c-Raf1 or PI3 kinase. This GBD is recruited by Rac and Ras GTPases in cells and plays an essential role for FHOD1-mediated actin remodeling. The FHOD1-FH3 domain is composed of five armadillo repeats, similarly to other formins. Mutation of one residue in the predicted DAD-interaction surface efficiently activates FHOD1 in cells. These results demonstrate that DRFs have evolved different molecular solutions to govern their autoregulation and GTPase specificity

Human endogenous retrovirus HERV-K(HML-2) encodes a stable signal peptide with biological properties distinct from Rec

<p>Abstract</p> <p>Background</p> <p>The human endogenous retrovirus HERV-K(HML-2) family is associated with testicular germ cell tumors (GCT). Various HML-2 proviruses encode viral proteins such as Env and Rec.</p> <p>Results</p> <p>We describe here that HML-2 Env gives rise to a 13 kDa signal peptide (SP) that harbors a different C-terminus compared to Rec. Subsequent to guiding Env to the endoplasmatic reticulum (ER), HML-2 SP is released into the cytosol. Biochemical analysis and confocal microscopy demonstrated that similar to Rec, SP efficiently translocates to the granular component of nucleoli. Unlike Rec, SP does not shuttle between nucleus and cytoplasm. SP is less stable than Rec as it is subjected to proteasomal degradation. Moreover, SP lacks export activity towards HML-2 genomic RNA, the main function of Rec in the original viral context, and SP does not interfere with Rec's RNA export activity.</p> <p>Conclusion</p> <p>SP is a previously unrecognized HML-2 protein that, besides targeting and translocation of Env into the ER lumen, may exert biological functions distinct from Rec. HML-2 SP represents another functional similarity with the closely related Mouse Mammary Tumor Virus that encodes an Env-derived SP named p14. Our findings furthermore support the emerging concept of bioactive SPs as a conserved retroviral strategy to modulate their host cell environment, evidenced here by a "retroviral fossil". While the specific role of HML-2 SP remains to be elucidated in the context of human biology, we speculate that it may be involved in immune evasion of GCT cells or tumorigenesis.</p

Microstructured blood vessel surrogates reveal structural tropism of motile malaria parasites

Plasmodium sporozoites, the highly motile forms of the malaria parasite, are transmitted naturally by mosquitoes and traverse the skin to find, associate with, and enter blood capillaries. Research aimed at understanding how sporozoites select blood vessels is hampered by the lack of a suitable experimental system. Arrays of uniform cylindrical pillars can be used to study small cells moving in controlled environments. Here, an array system displaying a variety of pillars with different diameters and shapes is developed in order to investigate how Plasmodium sporozoites associate to the pillars as blood vessel surrogates. Investigating the association of sporozoites to pillars in arrays displaying pillars of different diameters reveals that the crescent-shaped parasites prefer to associate with and migrate around pillars with a similar curvature. This suggests that after transmission by a mosquito, malaria parasites may use a structural tropism to recognize blood capillaries in the dermis in order to gain access to the blood stream