Special Issue "Structural and Molecular Biology of HIV"

Guest Editor
Prof. Dr. Andrew P. Rice
Nancy Chang Professor, Rice Lab - Molecular Virology & Microbiology, One Baylor Plaza, Mail Stop BCM-385, Houston, Texas 77030, USA
Website: http://www.bcm.edu/molvir/ricelab/?pmid=18017
E-Mail: arice@bcm.edu
Phone: 713-798-5774

Dear Colleagues,

Since the discovery of HIV almost 30 years ago, an enormous research effort has been conducted worldwide with the goal of conquering this infectious agent.  Although an effective HIV vaccine is not on the horizon, HIV research has partially achieved this goal, as effective anti-HIV drugs have been developed.  When used in combination, these drugs can suppress viral replication to almost undetectable levels in many patients.  Unfortunately, suppression of HIV replication requires lifelong adherence to the antiviral drugs, as a long-lived reservoir of latent viruses spontaneously reactivate when the drugs are discontinued.  A major effort in HIV research is now directed to discovering how this reservoir is established and maintained, with the long term goal of purging the reservoir and thereby curing HIV infection.
Basic research has provided the knowledge that lead to development of the existing anti-HIV drugs.   Continued research is required to produce more effective antiviral drugs, devise strategies to purge the viral reservoir, and develop an effective vaccine.  This special issue seeks to cover a broad range of HIV research in the areas of HIV molecular biology, host-virus interactions, and structural biology.  We welcome scientific perspectives, reviews, and original research papers.

Prof. Dr. Andrew P. Rice
Guest Editor

Submission

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Type of Paper: Review
Title: The Vesicular Budding of HIV and its Relationship to Exosome/Microvesicle Biogenesis
Author: Stephen J. Gould
Affiliation: Department of Biological Chemistry, Johns Hopkins University, Baltimore, MD 21205 USA; Tel: 443 847-9918; E-Mail: sgould@jhmi.edu
Abstract: Virus budding is an essential phase of the HIV lifecycle. HIV budding is driven by Gag, its primary structural protein. Two competing hypotheses have been put forward to explain the budding of HIV and its Gag protein. The prevailing, widely accepted view is that HIV budding requires the C-terminal p6 domain of Gag, which binds TSG101, recruits the ESCRT (endosomal sorting complexes required for transport) machinery to sites of virus assembly, and thereby induces the ESCRT-mediated catalysis of Gag/HIV budding. In contrast, the Trojan exosome hypothesis posits that HIV buds by the non-viral pathway of exosome/microvesicle (EMV) biogenesis, that Gag/HIV budding is mediated by the plasma membrane binding and higher-order oligomerization of the Gag protein, and that these elements lie upstream of p6 in the matrix, capsid, and nucleocapsid domains of Gag. Here I describe these competing models, review their ability to predict/explain the available data, and discuss areas of future investigation that would help elucidate which model best explains Gag/HIV budding.

Type of Paper: Article
Title: Global Conformational Dynamics of HIV-1 Reverse Transcriptase Bound to Non Nucleotide Inhibitors
Authors: D Wright, B Hall, P Kellam and P V Coveney *
Affiliation: Centre for Computational Science, University College London, 20 Gordon Street, London WC1H 0AJ, UK; E-Mail: p.v.coveney@ucl.ac.uk
Abstract: HIV-1 Reverse Transcriptase (RT) is a multifunctional enzyme responsible for the transcription of the RNA genome of the HIV virus into DNA suitable for incorporation within the DNA of human host cells. Its crucial role in the viral life cycle has made it one of the major targets for antiretroviral drug therapy. The Non Nucleotide RT Inhibitor (NNRTI) class of drugs binds allosterically to the enzyme, disrupting the polymerase active site and altering the dynamics of the enzyme. A popular theory of NNRTI finction is that the inhibitors lock the p66 thumb domain into position which creates an unaturally wide DNA binding cleft, preventing RT from correctly binding substrates and hence performing its polymerase function. However, coarse grained network model analysis, based on crystal structures, have indicated that different NNRTIs alter domain movements in distinctive ways. Here we use ensemble molecular dynamics alongside network models to explore global dynamics on a multinanosecond timescale. We find that, rather than inhibitor binding, the biggest determinant of equilibrium dynamics is the global conformational state of the enzyme. Furthermore we suggest that the dynamic variations seen in previous network models may be decoupled from the process of conformational change.

Type of Paper: Review
Title: The Surprising Role of Amyloid Fibrils in HIV Infection
Authors: Laura M. Castellano and James Shorter
Affiliation: Department of Biochemistry and Biophysics, Perelman School of Medicine at The University of Pennsylvania, 804 Stellar-Chance Laboratories, 422 Curie Boulevard, Philadelphia, PA 19104, USA; E-Mail: jshorter@mail.med.upenn.edu
Abstract: Despite its discovery over 30 years ago, human immunodeficiency virus (HIV) continues to be a pressing threat to public health worldwide.  Semen serves as the predominant vehicle promoting the spread of this retrovirus, and recently, several endogenous peptides in semen were found to boost HIV infection.  Interestingly, each of the identified peptides was found to be amyloidogenic, indicating that these agents exist naturally as amyloid fibrils in semen.  By facilitating virion attachment and fusion to target cells, these amyloid fibrils can greatly enhance HIV infectivity, and they present a novel target for the development of an innovative means to combat HIV transmission.  Here we review the identification of these amyloidogenic peptides, their mechanism of action, and current strategies for inhibiting their HIV-enhancing effects.

Type of Paper: Review
Title: Computer-Aided Approaches for Targeting HIVgp41
Authors: William J. Allen¹ and Robert C. Rizzo ^*,1,2,3
Affiliations: ¹ Department of Applied Mathematics and Statistics, Stony Brook University, Stony Brook, New York, 11794-3600, USA
² Institute of Chemical Biology & Drug Discovery, Stony Brook University, Stony Brook, New York, 11794-3600, USA
³ Laufer Center for Physical & Quantitative Biology, Stony Brook University, Stony Brook, New York, 11794-3600, USA; E-Mail: rizzorc@gmail.com
Abstract: Virus-cell fusion is the primary means by which the human immunodeficiency virus (HIV) delivers its genetic material into the human T-cell host. Fusion is mediated in large part by the viral glycoprotein 41 (gp41) which advances through four distinct conformational states: (1) native, (2) pre-hairpin intermediate, (3) fusion active (fusogenic), and (4) post-fusion. The pre-hairpin intermediate is a particularly attractive step for therapeutic intervention given that gp41 N-terminal (NHR) and C-terminal (CHR) heptad repeat domains are transiently exposed prior to the formation of a six-helix bundle required for fusion. Peptide-based inhibitors, including the FDA-approved drug T20, target the intermediate and there are significant efforts to develop small molecule alternatives. Here, we review current approaches to studying interactions of inhibitors with gp41 with an emphasis on atomic-level computer modeling including molecular dynamics, free energy analysis, and docking. Atomistic modeling yields a unique level of structural and energetic detail, complementary to experimental approaches, which will be important for the design of improved next generation anti-HIV drugs.

Type of Paper: Review
Title: Making a short story long: Regulation of P-TEFb and HIV-1 transcriptional elongation in CD4+ T lymphocytes and macrophages
Authors: Rajesh Ramakrishnan¹, Karen Chiang ^1,2, Hongbing Liu ¹, Sona Budhiraja ¹, Hart Donahue ¹ and Andrew P. Rice ^{1, 2}
Affiliations: ¹ Department of Molecular Virology & Microbiology, Baylor College of Medicine, Houston, TX, 77030, USA; E-Mail: arice@bcm.edu
² Interdepartmental Program in Translational Biology and Molecular Medicine, Baylor College of Medicine, Houston, TX, 77030, USA
Abstract: Productive transcription of the integrated HIV-1 provirus is restricted by cellular factors that inhibit RNA polymerase II elongation.  The viral Tat protein overcomes this by recruiting a general elongation factor, P-TEFb, to the TAR RNA element that forms at the 5’ end of nascent viral transcripts.  P-TEFb exists in multiple complexes in cells, and its core consists of a kinase, Cdk9, and a regulatory subunit, either Cyclin T1 or Cyclin T2.  Tat binds directly to Cyclin T1 and thereby targets the Cyclin T1/P-TEFb complex that phosphorylates the CTD of RNA polymerase II and the negative factors that inhibit elongation, resulting in efficient transcriptional elongation.   P-TEFb is tightly regulated in cells infected by HIV-1 – CD4+ T lymphocytes and monocytes/macrophages.  A number of mechanisms have been identified that inhibit P-TEFb in resting CD4+ T lymphocytes and monocytes, including miRNAs that repress Cyclin T1 protein expression and dephosphorylation of residue Thr186 in the Cdk9 T-loop. These repressive mechanisms are overcome upon T cell activation and macrophage differentiation when the permissivity for HIV-1 replication is also greatly increased.  This review will summarize what is currently known about mechanisms that regulate P-TEFb and how this regulation impacts HIV-1 replication and latency.

Type of Paper: Article
Title: A Comparison of Two Single-Stranded DNA Binding Models by Mutational Analysis of APOBEC3G
Authors: Keisuke Shindo ^1,2, Ming Li ¹, Phillip J. Gross ¹ , Elena Harjes ³, Yongjian Lu ³, Hiroshi Matsuo ³ and Reuben S Harris ^1,*
Affiliations: ¹ Department of Biochemistry, Molecular Biology and Biophysics, Institute for Molecular Virology and Center for Genome Engineering, University of Minnesota, Minneapolis, Minnesota, 55455 USA; E-Mail: rsh@umn.edu
² Kyoto
³ Department of Biochemistry, Molecular Biology and Biophysics, Institute for Molecular Virology, University of Minnesota, Minneapolis, Minnesota, 55455 USA
Abstract: APOBEC3G is a DNA cytidine deaminase that inhibits the replication of a variety of parasitic genetic elements including the lentivirus HIV-1. Several high-resolution structures of the APOBEC3G catalytic domain exist, but none demonstrate how this enzyme binds single-stranded DNA substrates. Here, we constructed a panel of APOBEC3G amino acid substitution mutants and performed a variety of assays to distinguish between “Brim” and “Kink” models for single-strand DNA binding. Each model predicts distinct sets of interactions between surface arginines and the negatively charged phosphodiester backbone of single-stranded DNA. Concordant with both models, changing the conserved arginine at position 313 to glutamate abolished catalytic and HIV-1 restriction activities. In support of the Brim model, arginine to glutamate substitutions at positions 213, 215, and 320 also compromised APOBEC3G activities. Analogous substitutions at Kink model residues 374 and 376 yielded discordant results, with a glutamate substitutions having modest to little effects, respectively. Our data are most consistent with the Brim model for single-stranded DNA binding by APOBEC3G.

Type of Paper: Review
Title: Dynamic regulation of HIV-1 gene expression
Author: Alessandro Marcello
Affiliation: Laboratory of Molecular Virology, International Centre for Genetic Engineering and Biotechnology (ICGEB), Padriciano, 99, 34149 Trieste, ITALY; E-Mail: marcello@icgeb.org
Abstract: Gene expression of the human immunodeficiency virus type 1 (HIV-1) is a highly regulated process with profound pathological implications. HIV-1 mRNAs are efficiently transcribed by Tat-mediated activity on RNA polymerase elongation but inefficiently processed allowing the Rev-mediated export of RNAs that contain the Rev-responsive RNA element (RRE). These non-terminally spliced RNAs are exported from the nucleus to allow expression of Gag-Pol and Env proteins and for the production of full-length genomic RNAs. Hence, a balance exists between efficiently processed mRNAs and RNAs that are retained in the nucleus awaiting Rev action. Here we discuss the dynamic regulation of this pathway in light of recent findings that implicate several novel cellular cofactors of Rev function.

Type of Paper: Review
Title: The AID/APOBEC family as intrinsic immunity against HIV and retroelements
Authors: Atsushi Koito ^1,*  Kazuhiko Maeda ²,  Terumasa Ikeda ¹,  Nobuo Sakaguchi ²
Affiliations: ¹ Department of Retrovirology and Self-Defense, Faculty of Life Science, Kumamoto University, Kumamoto 860-8556, Japan 
² Department of Immunology, Faculty of Life Science, Kumamoto University, Kumamoto 860-8556, Japan; E-Mail: akoito@kumamoto-u.ac.jp
Abstract: Retroviruses, including HIV and ancestral mobile retroelements are controlled by host restriction machineries that play key roles in intrinsic immunity, an example being members of the cytidine deaminases AID/APOBEC family. The AID/APOBECs employ multiple mechanisms to keep mobile elements and foreign DNAs under control. Although recent evidence indicates that the ancestor of the AID/APOBECs is a DNA mutator, their biological activities are modulated and controlled by interaction with RNA as well as other protein molecules.  This review discusses the current understanding of the mechanism of action of AID/APOBECs and their role in controlling their targets: retrovirus and retroelement.

Type of Paper: Article
Title: Free energy profile of the deaminase domain of HIV-1 restriction factor APOBEC3G calculated by a molecular dynamics simulation
Authors: Yoshifumi Fukunishi ^1,*, Saki Hongo ², Masami Lintuluoto ² and Hiroshi Matsuo ^3,*
Affiliations: ¹ Biomedicinal Information Research Center (BIRC), National Institute of Advanced Industrial Science and Technology (AIST), 2-3-26, Aomi, Koto-ku, Tokyo 135-0064, Japan; E-Mail: y-fukunishi@aist.go.jp
² Division of Applied Life Science, Graduate School of Life and Environmental Science, Kyoto Prefectural University, Kyoto 606-8522, Japan
³ Department of Biochemistry, Molecular Biology and Biophysics, University of Minnesota, Minneapolis, Minnesota, USA 55455; E-Mail: matsu029@umn.edu
Abstract: The human APOBEC3G protein (A3G) is a single-stranded DNA deaminase that inhibits the replication of HIV-1, other retroviruses, and retrotransposons. Atomic details of A3G’s catalytic mechanism are starting to emerge as the structures of the catalytic domain of A3G (A3Gctd) have been determined using NMR and X-ray crystallography. Even though overall tertiary structures are similar between the NMR and crystal structures, there is some disagreement over β-strand 2 (β2) and loops containing amino acids which are important for the catalytic reaction. In order to find which structure is more stable in solution, we calculated the free energy profile of the A3Gctd structures by using the Generalized Born/Solvent Accessible Surface (GBSA) method accompanied with a molecular dynamics simulation. In order to consider entropy term of free energy, we have developed a new method that takes into account the distribution of dihedral angles of protein. The crystal structures had lower energies than the NMR structures as long as only the enthalpy term was considered. When both enthalpy and entropy terms were considered, the NMR structures had lower energies than the crystal structures, suggesting that NMR structures were dominant in solution. In addition, the β2-loop-β2’ structure which had been observed in all NMR structures was stable throughout molecular dynamics simulation for 10 nanoseconds. Therefore, in 4 contrast to previous speculations, it is not likely that A3Gctd adapts the continuous β2 strand structure. The loops containing ssDNA-binding arginines stayed around within 4Å of their original position through the 10 nsec MD; keeping the catalytic Zn2+ inaccessible to the target cytosine. This may well explain why A3G does not bind or catalyze single cytosine nucleoside. In the A3Gctd crystal structures, the loops are away from the catalytic site; leaving Zn2+ accessible to the target cytosine. Therefore, we speculate that A3Gctd became more enthalpy driven structure similar to the crystal structures when it lost entropy upon binding to a DNA substrate, which enable the target cytosine to access to the catalytic Zn2+.

Type of Paper: Review
Title: Breaking Barriers to an AIDS Model with Macaque-tropic HIV-1 Derivatives
Authors: Rajesh Thippeshappa, Hongmei Ruan and Jason T. Kimata
Affiliation: Department of Molecular Virology and Microbiology, Baylor College of Medicine, Houston, TX 77030, USA; E-Mail: jkimata@bcm.edu
Abstract: The development of an animal model of human immunodeficiency virus type 1 (HIV-1)/AIDS for preclinical testing of antiretroviral therapy, vaccines, and curative strategies, and for studies of pathogenesis is hampered by the human-specific tropism of HIV-1. Although simian immunodeficiency virus (SIV) or HIV-1/SIV chimeric viruses (SHIVs)-rhesus macaque models are excellent surrogate models for AIDS research, the genetic difference between SIV and HIV-1 restricts their utility as model systems.  The identification of innate retroviral restriction factors has increased our understanding about blockades to HIV-1 replication in macaques and provided a guide for the construction of macaque-tropic HIV-1 clones.  However, while these viruses replicate in macaque cells in vitro, they are easily controlled in the macaque host and have not caused AIDS, indicating that we do not fully understand the restrictive barriers of innate immunity.  In this review, we discuss recent findings regarding HIV-1 restriction factors, particularly as they apply to cross-species transmission of primate lentiviruses and the development of a macaque model of HIV-1/AIDS.