hiv structure, life cycle, and replication (1) dr. matthew d. marsden, ph.d. ucla, department of...
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HIV Structure, Life Cycle, and Replication (1)
Dr. Matthew D. Marsden, Ph.D.UCLA, Department of Medicine
mmarsden@ucla.edu
http://www.ck12.org/concept/HIV/
HIV Structure, Lifecycle and Replication (1)
Background: Basic Virology and Pathogenesis
Structure: Virion structure, genomic structure, and accessory molecules
Lifecycle: Infection and Expression
On http://aids.gov/hiv-aids-basics/hiv-aids-101/aids-timeline/
A new disease…
On http://aids.gov/hiv-aids-basics/hiv-aids-101/aids-timeline/
A new disease…
•By the end of 1981, there was a cumulative total of 270 reported cases of severe immune deficiency among gay men, and 121 of those individuals had died.
•In 1983, Luc Montagnier and Françoise Barré-Sinoussi reported the discovery of a new virus (later called HIV) that is the cause of AIDS.
•The first commercial blood test for HIV was licensed in 1985, allowing screening of the U.S. blood supply.
•In 1987 the first anti-HIV drug (AZT) was approved by the U.S. Food and Drug Administration.
•The first potent combination of anti-HIV drugs became available in 1995.
Pneumocystis pneumonia—Los Angeles.Gottlieb M S, Schanker H M, Fan P T, Saxon A, Weisman J D & Polzalski“In the period October 1980-May 1981, 5 young men, all active homosexuals, were treated for biopsy confirmed Pneumocystis carinii pneumonia at 3 different hospitals in Los Angeles, California. Two of the patients died. All 5 patients had laboratoryconfirmed previous or current cytomegalovirus (CMV) infection and candidal mucosal infection”.Morbid. Mortal, Weekly Rep. 30:250-2. 1981.
Pneumocystis carinii Pneumonia and Mucosal Candidiasis in Previously Healthy Homosexual Men — Evidence of a New Acquired Cellular ImmunodeficiencyMichael S. Gottlieb, M.D., Robert Schroff, Ph.D., Howard M. Schanker, M.D., Joel D. Weisman, D.O., Peng Thim Fan, M.D., Robert A. Wolf, M.D., and Andrew Saxon, M.D.N Engl J Med 1981; 305:1425-1431December 10, 1981
Isolation of a T-Lymphotropic Retrovirus from a Patient at Risk for Acquired Immune Deficiency Syndrome (AIDS)F. Barré-Sinoussi; J. C. Chermann; F. Rey; M. T. Nugeyre; S. Chamaret; J. Gruest; C. Dauguet; C. Axler-Blin; F. Vézinet-Brun; C. Rouzioux; W. Rozenbaum; L. Montagnier. (May 20, 1983)Science, New Series, Vol. 220, No. 4599., pp. 868-871
Detection, isolation, and continuous production of cytopathic retroviruses (HTLV-III) from patients with AIDS and pre-AIDS
Frequent detection and isolation of cytopathic retroviruses (HTLV-III) from patients with AIDS and at risk for AIDS
Serological analysis of a subgroup of human T-lymphotropic retroviruses (HTLV-III) associated with AIDS
Antibodies reactive with human T-lymphotropic retroviruses (HTLV-III) in the serum of patients with AIDSGallo RC. et al. (May 1984)Science 224 (4648): 497–508
(I) Identification
of AIDS
(II)Isolation
of the virus
(III)Link
Virus-AIDS
Human immunodeficiency virus (HIV) is a retrovirus that causes acquired immunodeficiency syndrome (AIDS).
Since the epidemic was identified in 1981, more than 60 million people have contracted HIV and nearly 30 million have died of HIV-related causes.
At the end of 2011, an estimated 34 million people, an estimated 0.8% of adults aged 15-49 years worldwide, are living with HIV.
2.5 million new infections in 2011; 330,000 were children. 7,000 people contract HIV everyday, nearly 300 every hour.
In 2011 alone, AIDS claimed an estimated 1.7 million lives, of which 230,000 were children.
HIV primarily infects vital cells in the human immune system such as helper T cells (CD4+ T cells), macrophages and dendritic cells. HIV infection leads to low levels of CD4+ T cells.
http://www.unaids.org/en/
Region (lower- and middle-income countries) Antiretroviral therapy coverage
Estimated number of people receiving antiretroviral therapy
Estimated number of people needing antiretroviral therapy
Sub-Saharan Africa 37% 3,911,000 10,600,000
Eastern and Southern Africa 41% 3,203,000 7,700,000
Western and Central Africa 25% 709,000 2,900,000
Latin America and the Caribbean 50% 478,000 950,000
Latin America 51% 425,000 840,000
The Caribbean 48% 52,400 110,000
East, South and South-East Asia 31% 739,000 2,400,000
Europe and Central Asia 19% 114,000 610,000
North Africa and the Middle East 11% 12,000 100,000
Total 36% 5,254,000 14,600,000
Clinton Health Access Initiative (CHAI)$25 billion total needed to achieve all targets in low and middle-income countries (2009)On average it cost $380 to keep a patient on the medication for a year in Global Fund countries
HIV is responsible for a catastrophic pandemic:
http://aids.gov
http://aids.gov
MSM = Men who have sex with menIDU = Intravenous drug users
ESTIMATES OF NEW HIV INFECTIONS IN THE U.S., 2009, BY TRANSMISSION CATEGORY
What is HIV?
To understand what HIV and AIDS are, let’s break it down:
Causative agent:
H – Human – This particular virus can only infect human beings.
I – Immunodeficiency – HIV weakens your immune system by destroying important cells that fight disease and infection. A "deficient" immune system can't protect you.
V – Virus – A virus can only reproduce itself by taking over a cell in the body of its host.
http://aids.gov/hiv-aids-basics/hiv-aids-101/what-is-hiv-aids/
What is HIV?
To understand what HIV and AIDS are, let’s break it down:
Causative agent:
H – Human – This particular virus can only infect human beings.
I – Immunodeficiency – HIV weakens your immune system by destroying important cells that fight disease and infection. A "deficient" immune system can't protect you.
V – Virus – A virus can only reproduce itself by taking over a cell in the body of its host.
http://aids.gov/hiv-aids-basics/hiv-aids-101/what-is-hiv-aids/
What is HIV?
Disease:
A – Acquired – AIDS is not something you inherit from your parents. You acquire AIDS after birth.
I – Immuno – Your body's immune system includes all the organs and cells that work to fight off infection or disease.
D – Deficiency – You get AIDS when your immune system is "deficient," or isn't working the way it should.
S – Syndrome – A syndrome is a collection of symptoms and signs of disease. AIDS is a syndrome, rather than a single disease. It is a complex illness with a wide range of symptoms.
What is HIV?
Human immunodeficiency virus or HIV is a type of virus (from the Latin “virus” referring to poison).
Viruses are:
Small -Generally too small to see with a regular light microscope (20 - 300 nm diameter) If a cell was a football stadium then a small virus would be around the size of a football.
What is HIV?
Human immunodeficiency virus or HIV is a type of virus (from the Latin “virus” referring to poison).
Viruses are:
Small -Generally too small to see with a regular light microscope (20 - 300 nm diameter) If a cell was a football stadium then a small virus would be around the size of a football.
Can only replicate in living cells - Some can survive for long periods of time outside cells, but cannot replicate that way.
What is HIV?
Human immunodeficiency virus or HIV is a type of virus (from the Latin “virus” referring to poison).
Viruses are:
Small -Generally too small to see with a regular light microscope (20 - 300 nm diameter) If a cell was a football stadium then a small virus would be around the size of a football.
Can only replicate in living cells - Some can survive for long periods of time outside cells, but cannot replicate that way.
Made up of Nucleic acids (DNA/RNA) and proteins -different from protein-only “prions” or nucleic acid-only “viroids”.
Thousands of very different types of virus exist and HIV is a particular type termed a “retrovirus”.
Viruses
Microscopic infectious agents that can infect the cells of a biological organism.
Viruses can only replicate themselves by infecting a host cell and are incapable of reproducing on their own.
A complete viral particle, known as a virion consists of nucleic acid surrounded by a protective coat of protein called a capsid.
http://tcf.epfl.ch/page-20833-en.html
The HIV genome is composed of 9 genes, which encode 15 proteins.
For comparison, the E. Coli bacterium contains around 4,377 genes and the human genome encodes around 21,000 genes.
http://biology.kenyon.edu/slonc/gene-web/Lentiviral/Lentivi2.html
How does HIV cause disease?
HIV is a virus that infects and destroys cells of the immune system (CD4+ cells).
http://aids.gov/hiv-aids-basics/hiv-aids-101/what-is-hiv-aids/
HIV is a virus that infects and destroys cells of the immune system (CD4+ cells).
Approximately 8-10 years
Initial infectionAsymptomatic period
(clinical latency) AIDSOften (not always) accompanied by severe flu like symptoms: Opportunistic infections and cancer:
AIDS (acquired immunodeficiency syndrome) is the late-stage HIV disease. This occurs when immune system becomes so damaged that it cannot fight off diseases and certain types of cancer.
http://aids.gov/hiv-aids-basics/hiv-aids-101/what-is-hiv-aids/
HIV Life cycle
http://preprod.www.tibotec.com/content/backgrounders/www.tibotec.com/hiv_lifecycle.html
http://www.youtube.com/watch?v=9leO28ydyfU
Movie of the HIV Life Cycle
CD4
CXCR4
Binding
Fusion & Entry
Nuclear localization & entry
Reverse transcription
Integration
Infection
gp120
p24
Viral RNA
RT & othervirion proteins
Expression
gp120
p24
Viral RNA
RT & othervirion prteins
Budding
Assembly
Viral Gene Transcription
Translation
Post-translationalprocessing
Cellular Activation
HIV Structure
Virion
Genomic
Proteomic
http://www.readcube.com/articles/10.1038/nrmicro2596
http://upload.wikimedia.org/wikipedia/commons/0/00/HIV_Mature_and_Immature.PNG
HIV Structure
SIV HIV
HIV Structure
Virion
Genomic
Proteomic
CD4
CXCR4
Binding
Fusion & Entry
Nuclear localization & entry
Reverse transcription
Integration
Infection
gp120
p24
2 copies of viral RNA
RT & othervirion proteins
Referred to as:•Provirus•Proviral DNA•Integrated Provirus
R U5 RU3
HIV RNA
HIV DNA
R U5U3 R U5U3
LTR LTR
Host DNAHost DNA
Reverse transcription
Integration
HIV Genome
Nature Reviews Microbiology 2, 461-472 (June 2004)
HIV-1 RNA
R U5 RU3
RRE
Nature 460, 711-716 (6 August 2009)
Modified from : The AmFAR AIDS Handbook, D Ward, pp. 348
HIV-1 Integrated DNA
R U5U3 R U5U3
LTR LTR
Host DNAHost DNA
CD4 & MHC downregulation
RNA Splicing
TranslationalFrameshift
Source: The Molecular Biology of HIV/AIDS, D Ward, pp. 19
These are the major spliced RNA species, but HIV can actually produce at least 109 different spliced RNAs (Nucleic Acids Res. 2012 Nov 1;40[20])
Source: Atlas of Infectious Diseases, Mandell & Mildvan (ed.), pp. 3.13
Initiation of HIV transcription is performed by cellular factors
Dr. Isabelle BOUALLAGA Institut Pasteur. http://www.123bio.net/
Source: Atlas of Infectious
Diseases, Mandell & Mildvan (ed.),
pp. 3.13
HIV Structure
Virion
Genomic
Proteomic
HIV Proteins
Structural ProteinsGag: Matrix, Capsid, NC, p6 Pol: Protease, Reverse Transcriptase, Integrase Env: gp120, gp41
Regulatory ProteinsTatRevNef
Accessory proteinsVifVprVpu
CD4 & MHC downregulation
Structural protein expression and function
Source: The Molecular Biology of HIV/AIDS, D Ward, pp. 19
Gag and Gag-Pol
Source: BioAfrica Bioinformatics for HIV Research. http://bioafrica.mrc.ac.za/
Gag and Gag-Pol
Gene/Protein Mass, KDa Function
gag
(Pr) 55gag p55 Gag precursor protein
MA - Matrix p17 Aids nuclear import and viral assembly
CA - Capsid p24 HIV central core – contains HIV genome and enzymes
Matrix p17 Capsid p24 NC p9 p6p1p2
myristate
Association withmembrane
Formation of Gagmultimers
RNA bindingRNA packing
Viral Entry
EnvelopeIncorporation
Viral Budding
NC - Nucleocapsid p7/p9
Precise location in virion unknown, not generally present inother retroviruses, may aid in incorporation of Vpr into virion
Assoc. with HIV RNA, involved in packaging of HIVRNA into virions
p6 p6
Source: BioAfrica Bioinformatics for HIV Research. http://bioafrica.mrc.ac.za/
PR p10 RT IN p32p6
Gag
p51
p66
pol
(Pr) 160gag-pol p160 Gag-Pol precursor protein
PR - protease p10 Cleaves Gag and Gag-Pol
IN - integrase p32 Integrates viral genome into host DNA
RT – reversetranscriptase
p66/p51 p66 – copies RNA genome. RNAse H
p51 – regulatory?
Gene/Protein Mass, KDa Function
Source: The Molecular Biology of HIV/AIDS, D Ward, pp. 19
Envelope
env(Pr) 160 env gp160 Env precursor protein
SU – Envelope gp120 Surface glycoprotein that binds CD4
TM - Transmembrane gp41 Transmembrane protein that anchors gp120 to virus,responsible for fusion between virus and host cell membrane.
Gene/Protein Mass, KDa Function
Gene/Protein Mass, KDa Functiongag
(Pr) 55gag p55 Gag precursor protein
MA - Matrix p17 Aids nuclear import and viral assembly
CA - Capsid p24 HIV central core – contains HIV genome and enzymes
NC - Nucleocapsid p7/p9
Precise location in virion unknown, not generally present inother retroviruses, may aid in incorporation of Vpr into virion
Assoc. with HIV RNA, involved in packaging of HIVRNA into virions
pol(Pr) 160gag-pol p160 Gag-Pol precursor protein
PR - protease p10 Cleaves Gag and Gag-Pol precursor proteins
IN - integrase p31 Integrates viral genome into host DNA
RT – reversetranscriptase
p66/p51 p66 – copies RNA genome. RNAse H
p51 – functions in RT heterodimerenv
(Pr) 160 env gp160 Env precursor protein
SU – Envelope gp120 Surface glycoprotein that binds CD4 and coreceptors
TM - Transmembrane gp41 Transmembrane protein that anchors gp120 to virus,responsible for fusion between virus and host cell membrane
p6 p6
Source: The Molecular Biology of HIV/AIDS, D Ward, pp. 19
Regulatory proteins
Regulatory Proteins:TAT: Trans-Activator of TranscriptionREV: Regulator of Virion protein expressionNEF: Negative Regulatory Factor
Accessory Proteins:VIF: Virion Infectivity FactorVPU: Viral Protein UVPR: Viral Protein R
TAR
Poor transcription
TAT: Trans-Activator of Transcription
TAR
Poor transcription
Recruitment ofCDK9
Enhanced transcription
CDK9
Tat
Tat
Tat
TatTat
TAT: Trans-Activator of Transcription
Positive-feedback loop
RRE = rev-response element
REV: Regulator of Virion protein expression
NEF: Negative Regulatory Factor
* Downmodulation the expression of several surface molecules
important in host immune function:
MHC I, MHC II, CD4
* T-cell activation (activates the production of MIP-1alpha and
MIP-1beta in macrophages)
VIF: Virion Infectivity Factor
www.hivmedicine.com/textbook/pathogen.htm
VPU: Viral Protein U
• Involved in viral budding, enhancing virion release from the cell• Counteracts Tetherin (Bst2/CD317/HM1.24)
• Downregulation of CD4
http://www.plospathogens.org/article/info%3Adoi%2F10.1371%2Fjournal.ppat.1003299
Tetherin
VPU: Viral Protein U
* Involved in viral budding, enhancing virion release from the cell
* Downregulation of CD4
VPR: Viral Protein R
* Regulation of nuclear import of the HIV-1 pre-integration complex
* Required for virus replication in non-dividing cells such as macrophages.
* Induces cell cycle arrest and apoptosis in proliferating cells
G ene/Pro te in M ass , K D a Functionta t
Tat p14 Transactiva tes H IV transcrip tion (k inase adaptor to R N A P 2).
revR ev p19 N uclear export o f unsp liced & s ing le -sp liced R N A (v ia R R E ).
nef
N ef p27 In terna lizes ce ll-surface C D 4 & M H C C lass I m o lecu les .
M ay enhance ce llu la r ac tiva tion .
v if
Vif p23 O vercom es a post-en try b lock to in fec tion .
M ay in fluence v irion assem bly .
vpu
Vpu p16 Fac ilita tes v irion re lease v ia ion channe l fo rm ation .
Targets C D 4 fo r destruction in ER (free ing bound env).
vprVpr p15 Targets the H IV p re -in tegra tion com plex to the nuc leus .
A rres ts activa ted ce lls in G 2 phase o f ce ll cyc le .
Anti-HIV drugs
Nucleoside/Nucleotide Reverse Transcriptase Inhibitors
lamivudine (3TC), zalcitabine (ddC), zidovudine (AZT), didanosine (ddI), stavudine (d4T),
tenofovir
Non-Nucleoside Reverse Transcriptase Inhibitors
Delavirdine, efavirenz, nevirapine
Protease Inhibitors
Amprenavir , indinavir, saquinavir, saquinavir, lopinavir/ritonavir, ritonavir, nelfinavir
Integrase InhibitorsIsentress (Raltegravir or MK-0518) , JTK303/GS-9137
Fusion or Entry Inhibitors
Enfurvitide (Fuzeon or T20), Maraviroc (Selzentry -CCR5 antagonist-)
HAART (Highly Active Antiretroviral Therapy): three or more anti-HIV drugs (antiretrovirals) from
at least 2 different classes in combination allows potent inhibition of HIV replication.
Take-home points:
• Structure: – Env is only exposed viral protein in virion (neutralization resistant)– Binding/fusion with host cell is mediated by gp120 & gp41 (CD4 &
CCR5 or CXCR4)– RNA genome -- requires HIV reverse transcription to DNA– integration requires HIV integrase– LTR/promoter requires cellular transcription factors– HIV protease activity is needed for generation of infectious virions– accessory genes modulate:
1) cellular function (e.g., nef, vpr)
2) viral gene expression (e.g., tat, rev)– Capsid (p24) represents the primary structural component of virion– small molecule inhibitors of these structure’s functional activity
represent the primary current antiviral therapeutic strategies
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