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Casey Finnerty

HHMI's BioInteractive - Infectious Diseases Lecture Series - 2 views

www.hhmi.org/...lectures.html

virology HIV AIDS Dengue virus genomics SARS media

shared by Casey Finnerty on 16 Nov 12 - Cached
  • Casey Finnerty on 16 Nov 12
     
    This page has links to all the HHMI Holiday Lectures on infectious disease topics.Currently these include: "2000 and Beyond: Confronting the Microbe Menace", "AIDS: Evolution of an Epidemic", and "Viral Outbreak: The Science of Emerging Disease".
Casey Finnerty

Dr. Donald A. Henderson, Who Helped End Smallpox, Dies at 87 - The New York Times - 1 views

www.nytimes.com/...d-end-smallpox-dies-at-87.html

smallpox vaccine

shared by Casey Finnerty on 22 Aug 16 - No Cached
  • Dr. Donald A. Henderson, a leader of one of mankind’s greatest public health triumphs, the eradication of smallpox, died on Friday in Towson, Md. He was 87.
  • died in a hospice of complications of a hip fracture, including infection with antibiotic-resistant staphylococcus, a dangerous pathogen he had himself researched and raised alarms about
  • The last known case was found in a hospital cook in Somalia in 1977.
  • ...14 more annotations...
  • The only other disease to have been banished from the earth is rinderpest, a little-known relative of measles that kills hoofed animals and once caused widespread starvation in Africa; it was eradicated in 2011.
  • Smallpox, caused by the variola virus, was long one of mankind’s most terrifying scourges.
  • it killed almost a third of its victims, often through pneumonia or brain inflammation.
  • Many others were left blind from corneal ulcerations or severely disfigured by pockmarks.
  • Because it killed 80 percent of the American Indians who caught it, it was a major factor in the European conquest of the New World.
  • In 1796, Dr. Edward Jenner, an English physician, infected a young boy with cowpox taken from a blister on a milkmaid’s hand. Cowpox, a mild disease, protected those who had it from smallpox, and the modern vaccine era began.
  • Dr. Henderson quickly realized that trying to vaccinate vast populations was futile and switched to “ring vaccination.”
  • Dr. Foege, who is considered the father of this tactic, said it was “invented by accident” during a 1967 Nigerian outbreak when he had very little vaccine on hand.
  • “The first night, we asked ourselves what we would do if we were a virus bent on immortality,”
  • In 1977, success in hand, Dr. Henderson became dean of the Johns Hopkins University School of Hygiene and Public Health.“He was an imposing guy — physically big and very confident,” said Dr. Michael J. Klag, the school’s current dean, who was a student in that era. “He did not suffer fools gladly, and you were never sure if you were a fool or not.”
  • He gloomily foresaw failure for most other disease-elimination campaigns. The “siren song of eradication,” he once wrote, had led to goals that were more “evangelical” than attainable.
  • The problem, he would explain, was that each viral foe was so different. Smallpox had many weaknesses to exploit; it had no animal host. Every case can be found because victims have pox on their faces, and one vaccination provides lifetime immunity.
  • In 2011, when Bill Gates threw the full weight of his foundation into fighting polio, he struggled to explain how he would overcome such obstacles and, at the end of an interview, turned to an aide and said aloud, “I’ve got to get my D. A. Henderson response down better.”
  • The campaign, many experts have noted, succeeded just in time. A few years later, the virus that causes AIDS spread across Africa. Because the live smallpox vaccine can grow in an immune-compromised person into a huge, rotting, ultimately fatal lesion, it would have been impossible to deploy it.
Trevor F

Unique molecular signatures of disease brain endothelia provide a novel site for CNS-di... - 5 views

www.ncbi.nlm.nih.gov/...PMC3181494

Adeno-associated virus gene therapy

shared by Trevor F on 27 Sep 12 - No Cached
  • Trevor F on 27 Sep 12
     
    Great paper for introductory look at adapting adeno-associated viruses (AAV) to different cell tropisms.  Although the scope of this article is the CNS, and more specifically the brain.  Also goes over how brain vasculature expresses different characteristics in disease states that allows for specification of AAVs to have tropism for the diseased epithelial beds.
Casey Finnerty

Panel Recommends HPV Vaccine for Boys and Young Men - NYTimes.com - 0 views

www.nytimes.com/...26vaccine.html

microbiology HPV vaccination virology

shared by Casey Finnerty on 29 Aug 12 - No Cached
  • HPV infection is the most common sexually transmitted disease — between 75 percent and 80 percent of females and males in the United States will be infected at some point in their lives. Most will overcome the infection with no ill effects. But in some people, infections lead to cellular changes that cause warts or cancer, including cervical, vaginal and vulvar cancers in women and anal cancers in men and women. A growing body of evidence suggests that HPV also causes throat cancers in men and women as a result of oral sex. HPV infections cause about 15,000 cancers in women and 7,000 cancers in men each year. And while cervical cancer rates have plunged over the past four decades because of widespread screening, anal cancer rates in men and women have been increasing. Head and neck cancers have also been increasing, with the share associated with HPV infection increasing rapidly — perhaps because oral sex has increased in popularity.
Casey Finnerty

New Infection, Not Relapse, Brings Back Symptoms of Lyme Disease, Study Finds - NYTimes... - 4 views

www.nytimes.com/...-lyme-disease-study-finds.html

microbiology Lyme disease

shared by Casey Finnerty on 14 Nov 12 - No Cached
  • a new study finds that repeat symptoms are from new infections, not from relapses.
    • Sarah Muncy
      Sarah Muncy on 15 Nov 12
       
      But why are the patient so susceptible to getting the infections over and over again?
  • Dr. Steere acknowledged that symptoms, sometimes disabling ones, do linger for months after treatment in as many as 10 percent of patients. Doctors do not know why. But, Dr. Steere said, “antibiotics are not the answer.”
  • Casey Finnerty on 15 Nov 12
     
    Great question! I was wondering the same thing. I think the answer is actually in Chap. 6 of Cann! (Toward the end. Ask me again if you are unsure.) It's all in that little book, isn't it?!!!
Casey Finnerty

Flu Deaths Reach Epidemic Level, but May Be at Peak - NYTimes.com - 0 views

www.nytimes.com/...levels-but-may-be-peaking.html

virology influenza microbiology

shared by Casey Finnerty on 13 Jan 13 - Cached
  • Although the report supported getting flu shots, it said that new vaccines offering lifelong protection against all flu strains, instead of annual partial protection against a mix-and-match set, must be created.
  • “Vaccine effectiveness” is a very different metric from vaccine-virus match, which is done in a lab. Vaccine efficacy is measured by interviewing hundreds of sick or recovering patients who had positive flu tests and asking whether and when they had received shots.
  • During the 2009 swine flu pandemic, many elderly Americans had natural protection, presumably from flus they caught in the 1930s or ’40s.
  • ...12 more annotations...
  • “Think about that,” Dr. Osterholm said. “Even though they were old, they were still protected. We’ve got to figure out how to capture that kind of immunity — which current vaccines do not.”
  • Dr. Bresee acknowledged the difficulties, saying: “If I had the perfect answer as to how to make a better flu vaccine, I’d probably get a Nobel Prize.”
  • a preliminary study rated this year’s vaccine as 62 percent effective, even though it is a good match for the most worrisome virus circulating.
  • urged Americans to keep getting flu shots.
  • Even though deaths stepped — barely — into epidemic territory for the first time last Saturday, the C.D.C. officials expressed no alarm, and said it was possible that new flu infections were peaking in some parts of the country.
  • Epidemiologists count how many death certificates are filed in a flu year, compare the number with normal years, and estimate what percentage were probably flu-related.
  • The C.D.C.’s vaccine effectiveness study bore out the point of view of a report released last year by the University of Minnesota’s Center for Infectious Disease Research and Policy. It said that the shot’s effectiveness had been “overpromoted and overhyped,” said Michael T. Osterholm, the center’s director.
  • At the same time, he praised the C.D.C. for measuring vaccine effectiveness in midseason. “We’re the only ones in the world who have data like that,” he said.
  • “To get a vaccine across the ‘Valley of Death’ is likely to cost $1 billion,”
  • the metric means the shot “reduces by 62 percent your chance of getting a flu so bad that you have to go to a doctor or hospital.”
  • “far from perfect, but by far the best tool we have to prevent influenza.”
  • Most vaccinations given in childhood for threats like measles and diphtheria are 90 percent effective or better. But flu viruses mutate so fast that they must be remade annually.
Casey Finnerty

Powassan Virus (POW) Basics - Minnesota Dept. of Health - 0 views

www.health.state.mn.us/...basics.html

Powassan minnesota virology

shared by Casey Finnerty on 29 Aug 12 - No Cached
  • Initial laboratory testing in 2009-2010 found blacklegged ticks infected with POW virus in parts of north-central, east-central, and southeastern Minnesota, areas highly endemic for other tick-borne diseases such as Lyme disease.
Casey Finnerty

Questions Rise on Preparations at Hospitals to Deal With Ebola - NYTimes.com - 0 views

www.nytimes.com/...pitals-to-deal-with-ebola.html

ebolavirus

shared by Casey Finnerty on 14 Oct 14 - No Cached
  • “Usually, an individual is not sick for three to five days after the onset of symptoms, which will fool you,” Dr. Ribner said. “You say, ‘Oh, you’re not going to be that sick.’ Then, around Day 5 to 7, they really crash. Their blood pressure goes down, they become stuporous to unresponsive, and they start to have renal and liver failure. This correlates with the enormous viral load, which is just attacking every organ in the body.”
  • A concern for health workers is that as patients grow sicker, the levels of virus in their blood rise and they become more and more contagious. The researchers at Emory tested patients and found high levels of the virus in their body fluids and even on their skin.At the peak of illness, an Ebola patient can have 10 billion viral particles in one-fifth of a teaspoon of blood. That compares with 50,000 to 100,000 particles in an untreated H.I.V. patient, and five million to 20 million in someone with untreated hepatitis C.
  • “That helped us to understand why, if this is only spread by body fluids, why it is more contagious than hepatitis A, B and C, and H.I.V.,” Dr. Ribner said. “It’s just that there’s so much more virus in the fluids they put out.”
apopp10

The Major Genetic Determinants of HIV-1 Control Affect HLA Class I Peptide Presentation - 11 views

www.ncbi.nlm.nih.gov/...PMC3235490

shared by apopp10 on 14 Nov 14 - No Cached
  • Yet a small number of people demonstrate sustained ability to control HIV replication without therapy. Such individuals, referred to as HIV controllers, typically maintain stable CD4+ cell counts, do not develop clinical disease, and are less likely to transmit HIV to others (2).
    • laceemarie
      laceemarie on 14 Nov 14
       
      This is really cool! I have not heard of this until now. Have these people been studied to find out why this occurs? Is it because of the genetics of the person or a certain mutation in the virus? And the key words here are "less likely." That's a pretty broad statement - "less likely" meaning how likely?
    • slgoogin8981
      slgoogin8981 on 24 Nov 14
       
      It is interesting to see that there are 3 allelic variants that correlate with disease prognosis, but how does this information benefit society? We can't change peoples genetic so to alter their prognosis. Is this just a benefit to know what medications would be most beneficial with the least amount of side effects?
  • Abstract
    • Sean Hogan
      Sean Hogan on 02 Dec 14
       
      Presentation Paper 11/14
  • ...2 more annotations...
  • Although variation in the entire HLA protein is involved in the differential response to HIV across HLA allotypes, the major genetic effects are condensed to the positions highlighted in this study, indicating a structural basis for the HLA association with disease progression that is probably mediated by the conformation of the peptide within the class I binding groove.
    • becky214
      becky214 on 06 Dec 14
       
      If variation in the HLA protein results in different responses to HIV, does it also cause different responses to antiviral HIV drugs?
    • apopp10
      apopp10 on 08 Dec 14
       
      So it's only the amino acid position that accounts for the SNP and HLA association signals? It does not matter what amino acid you place there? Individual amino acid chemistry will have no affect on it?
sdahlseng10

In Vivo Gene Delivery and Stable Transduction of Nondividing Cells by a Lentiviral Vector - 0 views

www.jstor.org.wsuproxy.mnpals.net/...2889637.pdf

gene therapy nondividing cells lentivirus vector

shared by sdahlseng10 on 02 Oct 12 - No Cached
  • sdahlseng10 on 02 Oct 12
     
    Supplemental Article to Lesch et al. (2011); Journal club 1: This article provides interesting support to the effectiveness of lentivirus vectors for gene therapy treatment. Not only is HIV or lentivirus efficient at transduction to rapidly dividing cells, it is also capable of transduction to nondividing cells; making it an exciting vector for gene therapy treatment. The range of diseases that could be treatable with a gene therapy vector that can target other cell types than hematopoietic cells could transform the science of gene therapy.
Casey Finnerty

To Fight a Virus, Get a Virus: Military Bets on Mutant Pathogen - Bloomberg - 1 views

www.bloomberg.com/...nfections-in-two-brothers-labs

shared by Casey Finnerty on 15 Jul 16 - No Cached
  • Casey Finnerty on 15 Jul 16
     
    Describes the use of therapeutic interfering particles to treat viral disease.
Casey Finnerty

Calls/Webinars|Clinician Outreach and Communication Activity (COCA) - 0 views

emergency.cdc.gov/calls

shared by Casey Finnerty on 03 Oct 14 - No Cached
  • Casey Finnerty on 03 Oct 14
     
    I *highly* recommend you check out the CDC COCA calls if you are interested in medicine and infectious disease. Today's call on focal limb paralysis co-incident with the current outbreak of EV-D68 was fascinating. You may find my notes on it at this link: https://www.evernote.com/shard/s7/sh/01e57d8c-bc3f-4e86-85c6-476117fa5731/80a13752cdcfd28a5a4224d1c9df882f
Casey Finnerty

What Would Keep Ebola from Spreading in the US? Investing in Simple Research Years Ago.... - 0 views

www.wired.com/...ebola-ic-research

ebolavirus

shared by Casey Finnerty on 16 Oct 14 - No Cached
  • Casey Finnerty on 16 Oct 14
     
    Maryn McKenna writes frequently and eloquently on infectious disease in her blog "Superbug". I recommend you check out her latest post on the Ebola cases in the US.
apopp10

Norovirus Translation Requires an Interaction between the C Terminus of the Genome-link... - 75 views

www.jbc.org/...21738.long

shared by apopp10 on 12 Sep 14 - No Cached
  • Importantly, however, despite the interaction between the norovirus VPg protein and eIF4E, it appears to be dispensable for MNV translation initiation, at least in vitro
    • ameliaobert
      ameliaobert on 15 Sep 14
       
      I find it interesting that the MNV is depensable if incorporated in vitro. Im curious as to why this could be.
    • Casey Finnerty
      Casey Finnerty on 26 Sep 14
       
      They are saying the VPg-eIF4E interaction is dispensable in vitro, not the virus itself.
  • Mass spectrometry was used to identify proteins present within the samples with a minimum of 2 unique peptides and >90% identification probability
    • laceemarie
      laceemarie on 16 Sep 14
       
      I'm confused as to how exactly mass spectrometry was used in this experiment. If the proteins are already separated by molecular weight after the SDS-PAGE, what precisely are they looking for when running a mass spec? It article says, ".... a minimum of 2 unique peptides and >90% identification probability," but this doesn't mean much to me. I understand the identification part because based on the way the molecule splits after being shot with ions, you can get a basic idea of the structure of the molecule. My question with the 2 unique peptides is that each amino acid has a different molecular weight right? So if the proteins were all the same except at the 2 unique peptides, the mass spec would show a different mass to charge ratio for each protein with the varying peptides correct? Would the charge of the amino acids in the peptide have anything to do with the charge portion of the mass to charge ratio? I hope this question came out okay. 
    • abachman12
      abachman12 on 22 Sep 14
       
      I also found this confusing. I understand what your saying and it makes your question makes sense to me. I think that the charge of the amino acids in the peptide would have something to do with the charge portion of the mass to charge ration.
    • Casey Finnerty
      Casey Finnerty on 22 Sep 14
       
      I'll address this in class. In the meantime, check out http://en.wikipedia.org/wiki/Peptide_mass_fingerprinting
  • inability to culture many members of the virus family in immortalized cells.
    • slgoogin8981
      slgoogin8981 on 17 Sep 14
       
      What are the other options to culture these viruses?
    • Casey Finnerty
      Casey Finnerty on 22 Sep 14
       
      This is the subject of intense study. See ref 18.
  • ...34 more annotations...
  • The mutation F123A ablated the ability of VPg to co-purify eIF4G, eIF4A, and PABP but did not affect the ability of NTAP-MNV VPg to co-purify eIF4E (Fig. 5), suggesting that the eIF4E and eIF4G binding regions on VPg are distinct
    • ameliaobert
      ameliaobert on 17 Sep 14
       
      If F123A mutation is observed through data analysis (co-purification) that the eIF4E and eIF4G are distinct, so differ from each other. Then, why does the data suggest that the direct interacts of VPg with eIF4G determines the eIF4F binding capacity from the data obtained? As well as how do these purification datas are able to determine if the binding capacities for the virus VPg proteins are direct interactions?
    • Casey Finnerty
      Casey Finnerty on 22 Sep 14
       
      As written, your question is unclear. Please rewrite it.
  • These data would suggest that the direct interaction of VPg with eIF4G largely determines the eIF4F binding capacity of the norovirus VPg protein.
  • some of which function directly in translation initiation, whereas others may contribute to the regulation of host cell translation by virus infection.
    • ameliaobert
      ameliaobert on 17 Sep 14
       
      So here, they are saying that the initiation factors, some directly interact with the viruses translation to mRNA, whereas the other initiation factors present in teh virus contribute there translation factors specifically to when the host cells get involved with the virus translation factors? Basically specifically encoding for specific initiation translation factors based on the area of translation (i.e. from the virus or from the host cell)?
  • using the number of unique peptides identified for each protein (Table 1) as a semiquantitative indirect measure of protein abundance in the purified complex, it was also apparent that eIF4G was enriched in the complex with respect to the other proteins isolated.
    • becky214
      becky214 on 18 Sep 14
       
      I was confused by what they are suggesting with this data. Is it true that elF4G is a binding partner, or is it just suggested?
    • Casey Finnerty
      Casey Finnerty on 22 Sep 14
       
      Suggested. What evidence do you think would be conclusive?
    • Casey Finnerty
      Casey Finnerty on 22 Sep 14
       
      BTW, is that a golden eagle in your picture?
  • we attempted
  • to identify
  • components of the complex that interacted with VPg directly and contribute to viral translation
    • slgoogin8981
      slgoogin8981 on 18 Sep 14
       
      Is this the same as saying, they were looking for promoters?
    • Casey Finnerty
      Casey Finnerty on 22 Sep 14
       
      No. Promoters are involved with transcription. In this paper they are looking for eIF's (of translation) that interact with VPg.
  • Independent observations indicate that eIF4G phosphorylation is stimulated during norovirus replication in cell culture and that this phosphorylated form of eIF4G is associated with MNV VPg in infected cells (29). The impact and functional relevance of initiation factor phosphorylation on the norovirus life cycle is currently under study.
    • becky214
      becky214 on 19 Sep 14
       
      Maybe it is possible that this phosphorylation helps initiate and enhance translation.
    • alexridesducati
      alexridesducati on 19 Sep 14
       
      It seems in order for this virus to replicate effectively, it has to bind several different ways, at least, just to initiate protein synthesis. Is there any way to synthesize a molecule that can target any of these proteins for degradation or at least something that can inhibit the binding site (similar to a neutralizing antibody :) . It seems that if you nip this right at the bud, that its possible to shut this virus down for the most part, except for the fact that in vitro, elF4E can be dispensed for translation initiation. Makes me wonder what the key differences are between an in vitro and in vivo environment that allows this to happen.
  • high affinity interaction
    • smolizon11
      smolizon11 on 19 Sep 14
       
      Im confused to as what high affinity means?
    • Casey Finnerty
      Casey Finnerty on 22 Sep 14
       
      High affinity would mean a tighter interaction between two molecules. The high affinity nature of the VPg-eIF4G interaction is suggested by the fact that eIF4G is carried through the TAP procedure, and that it persists despite washing with 1M salt.
  • Noroviruses, members of the Caliciviridae family of small positive-strand RNA viruses, are a major cause of acute gastroenteritis
    • smolizon11
      smolizon11 on 19 Sep 14
       
      Its really interesting to be able to put a virus with being the cause of certain diseases. Even when talking about the disease in a "non-medical" setting its almost as if you have "higher hand" at the information about it. Makes for fun and interesting conversation with other people besides virology students.
  • As these proteins were not identified in both experimental systems and the focus of this study was to determine the role of canonical initiation factors in VPg-dependent translation, these additional host cell proteins factors were not examined in more detail.
    • Sean Hogan
      Sean Hogan on 19 Sep 14
       
          Is it possible that any of these are binding far upstream or downstream from the IF complex yet still affect affinity of complex compoenets such as with the eIF4E interaction with VPg protein in vitro?
    • Casey Finnerty
      Casey Finnerty on 22 Sep 14
       
      Yes.
  • recombinant MNV was performed by infecting baby hamster kidney cells with fowlpox virus expressing T7 RNA polymerase followed by transfection of MNV cDNA expression constructs as described previously
    • slgoogin8981
      slgoogin8981 on 19 Sep 14
       
      Was the fowlpox used just for the purpose of its polymerase? What is cDNA?
    • Casey Finnerty
      Casey Finnerty on 22 Sep 14
       
      cDNA stands for complementary DNA. Technically, that would mean the DNA strand that is complementary in sequence to the RNA. In practice, after the first cDNA strand is synthesized (typically using an RNA template and reverse transcriptase), the second strand is synthesized with a DNA polymerase as well. This double-stranded cDNA can then be cloned in a plasmid, which is then introduced into cells via transfection as done by the Goodfellow group. The fowlpox encoding RNA polymerase from T7 phage (this RNA pol is not error prone and highly specific for the T7 transcription start site) is used for two things: the T7 RNA polymerase drives transcription of the MNV RNA from the plasmid containing the cDNA; AND it also promotes capping of the MNV genome/transcript, which promotes translation of MNV proteins. See http://www.ncbi.nlm.nih.gov/pmc/articles/PMC3471295/, Fig. 4.
    • slgoogin8981
      slgoogin8981 on 26 Sep 14
       
      Thank you for the link! I am very unskilled with the lab results and procedures done in lab. I appreciate the guidance and opportunity to learn. I have only had three biology labs: 241, 242 and Cell.
    • Casey Finnerty
      Casey Finnerty on 01 Oct 14
       
      You're welcome! And thanks for the question!
  • human noroviruses have yet to be cultivated in the laboratory (18).
  • Our understanding of norovirus biology has been greatly enhanced by the discovery of murine norovirus (MNV) in 2003 (19),
  • Previous reports have also highlighted a potential association of the norovirus VPg protein with components of the eIF3 complex (28) and eIF4E, PABP, and eIF4G as well as the ribosomal protein S6 (29), although the functional relevance of these interactions has yet to be demonstrated. Here we describe the proteomic characterization of the murine norovirus translation initiation factor complex, demonstrating that VPg associates directly with the core components of the eIF4F complex and PABP. We further demonstrate that the interaction between eIF4G and VPg is essential for norovirus translation. Furthermore, we demonstrate that eIF4G is required for efficient virus replication in cell culture.
  • although these cells are not permissive to MNV infection due to the lack of a suitable receptor, robust MNV translation and replication occurs upon transfection of viral RNA into the cytoplasm (22).
    • slgoogin8981
      slgoogin8981 on 19 Sep 14
       
      It is very cool that this virus can still use human translation equipment even though it is a marine from of the virus. How did they enter the virus genome into the cytoplasm?
  • It is worth noting, however, that in addition to disrupting electrostatic interactions, increasing sodium chloride concentrations stabilize hydrophobic interactions.
  • It is important to note that in both these approaches low levels of eIF4E remained (Fig. 8C) and 4E-BP1 expression in the MNV permissive cells did not completely block the eIF4E-4G interaction. Therefore, further studies on the role of eIF4E during the norovirus life cycle are clearly warranted, but it is worth noting that in addition to a direct role in cap binding for translation initiation, eIF4E plays numerous roles in the regulation of gene expression.
    • jpolanco10
      jpolanco10 on 19 Sep 14
       
      Why did this not block the eIF4E-4G interaction? Is there a really high affinity or was it able to recruit other proteins to aide it? It seems that the low level of eIF4E was still able to do its job, mainly regulation of the genes
  • Murine macrophage RAW264.7 and microglial BV2 cell lines were cultured in Dulbecco's modified Eagle's medium (DMEM) with 10% (v/v) fetal calf serum (FCS), penicillin (100 units/ml), streptomycin (100 μg/ml), and 10 mm HEPES buffer. Baby hamster kidney cells (BHK-21) expressing T7 RNA polymerase (BSRT7 cells) were cultured in similar media lacking HEPES but containing 1 mg/ml G418. Similarly, HEK 293T cells were maintained in media lacking HEPES. HEK 293T cells stably expressing pMEP4-NTAP or pMEP4-NTAP MNV VPg plasmids were supplemented with 50 μg/ml hygromycin B and nonessential amino acids. The HEK293 TREX cells stably expressing pcDNA4/TO-NTAP derivatives of MNV VPg were supplemented with 5 μg/ml Blasticidin and 200 μg/ml Zeocin. All cell lines were maintained at 37 °C and 10% CO2.
    • nkasdagly
      nkasdagly on 19 Sep 14
       
      Are they setting up the primary and secondary antibodies here or am i totally off? Just wondering because i see they used fetal calf serum and baby hampster kidney cells... 
  • Noroviruses, members of the Caliciviridae family of small positive-strand RNA viruses, are a major cause of acute gastroenteritis in man (8) but have also been identified in a number of other species including dogs (9, 10), cats (11), sheep (12), and cattle (13
    • Sean Hogan
      Sean Hogan on 19 Sep 14
       
      Work?
  • the first step in translation is the binding of the initiation factor eIF4E, a component of the eIF4F complex, to the 5′ cap structure.
  • eIF4F is a complex of three initiation factors; eIF4E is the cap-binding protein, eIF4A functions as an RNA helicase, and eIF4G acts as a scaffold to bridge the mRNA to the 40 S ribosomal subunit via its interaction with eIF3 (3).
  • 14 individual point mutations in VPg were introduced into the MNV infectious clone,
  • the effect of mutations on the ability of VPg to associate with initiation factors was assessed by the enrichment of the eIF4F complex using m7GTP-Sepharose followed by Western blotting for VPg.
  • the mutations V115A, D116A, and F123A altered the levels of infectious virus produced;
  • the mutants V115A, D116A, and F123A showed a consistently reduced interaction with eIF4F (Fig. 4).
  • The degree to which the mutation affected virus recovery appeared to correlate with the ability of VPg to be co-purified with initiation factors (Fig. 4).
  • These data would indicate that the disordered C terminus of the norovirus VPg protein contains amino acids involved in the interaction with eIF4G.
  • We recently analyzed the solution structure of the core domain of VPg using nuclear magnetic resonance (NMR) spectroscopy and found that the MNV VPg protein consists of a compact structured core formed by a pair of α-helices that is flanked by long, flexible N and C termini (35). The region we have identified as being involved in the direct interaction with eIF4G, namely the C-terminal domain, is disordered and, therefore, is likely to adopt a fixed structure only upon interaction with eIF4G.
    • rmeloche10
      rmeloche10 on 22 Sep 14
       
      Using this advanced spectroscopy to find the structured core is an awesome break in technology. It shows the C-terminal is disordered until interaction with the initiation factor, does this disorder provide any advantage within the virus?
    • Casey Finnerty
      Casey Finnerty on 22 Sep 14
  • eIF4GI
    • Casey Finnerty
      Casey Finnerty on 22 Sep 14
       
      eIF4GI and eIF4GII are related forms of eIF4G.
  • This approach was used in place of authentic infection to enable the use of the F123A VPg mutant as a specificity control as this cDNA clone does not produce infectious virus (Fig. 4).
    • Casey Finnerty
      Casey Finnerty on 22 Sep 14
       
      Throughout the paper, the authors have included helpful explanatory text like this.
  • These data indicate that the VPg binding site lies within residues 654–1131 of eIF4GI, a region known to contain both eIF4A and eIF3 binding sites (7).
    • Casey Finnerty
      Casey Finnerty on 22 Sep 14
       
      Couldn't the binding site be narrowed further to 675-1131?
  • Noroviruses, a major cause of gastroenteritis in man, have evolved a mechanism that relies on the interaction of translation initiation factors with the virus-encoded VPg protein covalently linked to the 5′ end of the viral RNA.
    • apopp10
      apopp10 on 08 Dec 14
       
      I guess this comment never got adhered to my original highlight. Here it is once more. As someone who has gastrointestinal issues including inflammation, i wonder if this virus could have played a role in the cause of my condition, Ulcerative Colitis. I think interrupting this interaction could provide an effective treatment against this virus.
  • Casey Finnerty on 12 Sep 14
     
    Journal Article for our Week 4 discussion.
jiyoung yoon

Association of HIV Diversity and Survival in HIV-Infected Ugandan Infants - 10 views

www.plosone.org/...10.1371%2Fjournal.pone.0018642

hiv genetic disease virology

shared by jiyoung yoon on 09 Sep 12 - No Cached
    • Sarah Muncy
      Sarah Muncy on 10 Sep 12
       
      Ha, so again, like an organism. Where a population has a great deal of genetic diversity, there are more traits onto which natural selection can act. The more diverse the HIV is at this given stage means a failure in this case though, as the host dies and cannot transmit the disease. Strange.
  • Casey Finnerty on 10 Sep 12
     
    That's a really cool observation, Sarah! It's really all about transmission. Since HIV is (primarily) sexually transmitted, infection in infants (via transovarial transmission) may be considered a dead end for the virus. Perhaps vertical transmission is an aberrance. If HIV would evolve into a less virulent form, perhaps vertical transmission would become more important. Just sayin'...
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