New insights into HTLV entry

International audiencen.

Type-specific radioimmunoassays for the gp70s of mink cell focus-inducing murine leukemia viruses: expression of a cross-reacting antigen in cells infected with the friend strain of the spleen focus-forming virus

We have isolated the gp70 of a helper-independent strain of a Friend mink cell focus-inducing (MCF) virus, Fr-MCF-1. This recombinant virus, like the previously described AKR-MCF viruses, has been shown by both biological and biochemical means to be an envelope gene recombinant between Friend murine leukemia virus (F-MuLV) and a mouse xenotropic virus. Utilizing (125)I- labeled Fr-MCF-1 gp70 and antiserum prepared against an MCF strain of Moloney type-C virus (Mol-MCF(83)), we have developed a radioimmunoassay which detects immunological determinant (s)contained in the gp70s of MCF viruses derived from F-MuLV, Mol-MuLV, and AKR-MuLV. This MCF determinant(s) is not detected in the ecotropic parents of each of these MCF viruses, nor in helper-independent murine xenotropic viruses derived from Swiss or BALB/c mice. A protein partially cross-reactive with the MCF gp70 determinant(s) is detected in a replicating xenotropic virus derived from NZB mice. Utilizing this MCF gp70 specific immunoassay, we can detect a cross-reacting gene product coded for by the Friend strain of the spleen focus-forming virus (SFFV) in rat fibroblasts nonproductively infected with SFFV. The results support earlier molecular hybridization studies which indicated that the genome of SFFV contains genetic information derived from both F-MuLV and xenotropic virus, and that the xenotropic-related sequences in SFFV are highly related to those found in MCF murine type-C viruses

Cholesterol dependence of HTLV-I infection

Cholesterol-rich plasma membrane microdomains are important for entry of many viruses, including retro-viruses. Depletion of cholesterol with 2-hydroxypropyl-β-cyclodextrin inhibits entry of human T cell leukemia virus type I (HTLV-1) and HTLV-I envelope pseudotyped lentivirus particles. Using a soluble fusion protein of the HTLV-I surface envelope protein with the immunoglobulin Fc domain, the HTLV-I receptor was found to colocalize with a raft-associated marker and to cluster in specific plasma membrane microdomains. Depletion of cholesterol did not alter receptor binding activity, suggesting a requirement for cholesterol in a postbinding virus entry step

Molecular Aspects of HTLV-1 Entry: Functional Domains of the HTLV-1 Surface Subunit (SU) and Their Relationships to the Entry Receptors

The initial step in retroviral infection involves specific interactions between viral envelope proteins (Env) and specific receptors on the surface of target cells. For many years, little was known about the entry receptors for HTLV-1. During this time, however, functional domains of the HTLV-1 Env were identified by analyzing the effects of neutralizing antibodies and specific mutations in Env on HTLV-1 infectivity. More recent studies have revealed that HTLV-1 infectivity involves interactions with three different molecules: heparan sulfate proteoglycans (HSPG), the VEGF-165 receptor Neuropilin 1 (NRP-1) and glucose transporter type 1 (GLUT1). Here, we revisit previously published data on the functional domains of Env in regard to the recent knowledge acquired about this multi-receptor complex. We also discuss the similarities and differences between HTLV-1 and other deltaretroviruses in regards to receptor usage

A high confidence, manually validated human blood plasma protein reference set

<p>Abstract</p> <p>Background</p> <p>The immense diagnostic potential of human plasma has prompted great interest and effort in cataloging its contents, exemplified by the Human Proteome Organization (HUPO) Plasma Proteome Project (PPP) pilot project. Due to challenges in obtaining a reliable blood plasma protein list, HUPO later re-analysed their own original dataset with a more stringent statistical treatment that resulted in a much reduced list of high confidence (at least 95%) proteins compared with their original findings. In order to facilitate the discovery of novel biomarkers in the future and to realize the full diagnostic potential of blood plasma, we feel that there is still a need for an ultra-high confidence reference list (at least 99% confidence) of blood plasma proteins.</p> <p>Methods</p> <p>To address the complexity and dynamic protein concentration range of the plasma proteome, we employed a linear ion-trap-Fourier transform (LTQ-FT) and a linear ion trap-Orbitrap (LTQ-Orbitrap) for mass spectrometry (MS) analysis. Both instruments allow the measurement of peptide masses in the low ppm range. Furthermore, we employed a statistical score that allows database peptide identification searching using the products of two consecutive stages of tandem mass spectrometry (MS3). The combination of MS3 with very high mass accuracy in the parent peptide allows peptide identification with orders of magnitude more confidence than that typically achieved.</p> <p>Results</p> <p>Herein we established a high confidence set of 697 blood plasma proteins and achieved a high 'average sequence coverage' of more than 14 peptides per protein and a median of 6 peptides per protein. All proteins annotated as belonging to the immunoglobulin family as well as all hypothetical proteins whose peptides completely matched immunoglobulin sequences were excluded from this protein list. We also compared the results of using two high-end MS instruments as well as the use of various peptide and protein separation approaches. Furthermore, we characterized the plasma proteins using cellular localization information, as well as comparing our list of proteins to data from other sources, including the HUPO PPP dataset.</p> <p>Conclusion</p> <p>Superior instrumentation combined with rigorous validation criteria gave rise to a set of 697 plasma proteins in which we have very high confidence, demonstrated by an exceptionally low false peptide identification rate of 0.29%.</p