EXISTENTIAL MEDICINE

Written by Janine Roberts

Monday, 18 August 2008 21:34

Extract from Fear of The Invisible

Why our cells make Retroviruses

I had earlier learnt that viruses invade cells in order to reproduce, and that cells are the victims of this process. I also learnt that HIV was a retrovirus that hijacked’ cells. But I now realised there was an entirely different way to see this process.  Biology, a science that unlike virology has no focus on illness, has taught me that no virus exists that is not made by a cell, that these are produced by all healthy cells, whether of plants, fungi, birds, fish or animals, and that cells apparently consider retroviruses so harmless that they will trustingly incorporate codes brought by them into their very genomes, into the protected centres of their being.

Why do we presume that viruses take the initiative when they enter a cell, when viruses are universally recognised to be inert? What if it is the other way around? What if cells actively attract the passing retrovirus because they need the information they carry? They reportedly carry markers that enable cells to recognize them.

Viruses are essentially, as we will see, cell-made transport particles, similar to the vesicles that cells make to use internally as transports. If cells send out such particles to share information between themselves, perhaps this explains why cell evolved viral receptors and can absorb many such particles without ill effect? It would even explain why cells invest a great deal of energy in making them. Since these transport particles seem to be no more living than a text message, this explains also why we cannot kill them with antibiotics.

What then of the viruses said to kill cells, the ‘cytolytic’?  Cytolysis in biology is defined as the death of a cell due to osmotic imbalance, due to the pressure being too great or low within. Could viruses jointly mount enough pressure to burst a human cell?  Today, this is thought very unlikely.  The virus is simply too small and the cell’s self-protection too able.  Currently, medical courses teach instead that viral-infected cells die mostly because of an ‘allergic’ reaction; or because our immune system will naturally kill a sick cell. We will look more at this in the next chapter.

If ‘viruses’ are essentially messenger particles, this may explain why the genomes of viruses can vary so much. It might not be always because they are ‘unstable’, or subject to chance ‘mutations’. It could be because they are designed by cells to carry a wide variety of messages. This would explain why the NIH reports that it is hard to classify them into ‘species’: ‘Since viruses can evolve with enormous rapidity under selective pressure, it is difficult to define by sequence precisely what a virus ‘species’ is;’ so it has been decided that they may be identified as of a particular species if less than 20% of their genetic code is different from other members of that species. (Yet the genetic codes of monkeys and humans have only a 2% difference.)

I wondered if the ‘selective pressure’ the NIH mentioned may be a factor that leads cells to alter the code in a virus? Once it is produced, then its codes are static – so its variability is due to the cell that makes it.

We still know remarkably little about why cells make retroviruses and their possible usefulness.  The same applies to other viruses. The cells of our world clearly invest a vast amount of energy in making milliards of them.  We live in a sea of them.  Every breath we take is full of them; but they rarely hurt us.  We evolved among them.  But two characteristics they all have by definition:  viruses are all produced by cells, whether from animals, fungi, plants or bacteria, and they all contain short segments of genetic code.

Comparing a cell’s size with that of a virus is, to put it simplistically, like comparing an elephant to a scrap of paper.  A virus is approximately a billion times smaller than a cell. In contrast, each cell of a multicellular organisms, whether animal, plant or fungi, contain nearly two metres of genetic code in its core – with even more in its mitochondria and organelles (small organs). And they are certainly not inert.  Today techniques like X-ray crystallography reveal that living cells pulsate with energy, colour and movement. 

The vast difference between the amount of genetic code in a virus and in a cell reflects the immensely greater capability of the latter. Yet much too often some scientists and journalists have permitted themselves to speak of viruses as if they are as intelligent as cells and have the same survival instincts.  

If we want to better understand why cells make retroviruses, and other viruses, it might help to first look at the similar particles cells make to use internally as transports. Our cells are constantly making vast numbers of hollow transport particles (‘vesicles’) in their ‘ribosome’ organelles;’ sending these out carrying cargo along intricate ‘road-systems’ of ‘microtubules.’ This system is extremely busy – supplying materials for some 100,000 chemical reactions a second including cargoes of protein, enzymes and even DNA:  ‘The final proof that DNA actually entered the vesicles came from import experiments, in which radiolabeled ssDNA [single stranded DNA] was shown to accumulate inside the vesicles.’  Communication systems are thus as vital for our cells as they are for the largest of our hi-tech factories. 

The ‘Golgi Apparatus’ is the cell’s ‘post office.’ It directs these cargoes to where they are most needed. This system also supplies the materials for the assembly of retroviruses on internal membranes. It can transport large mitochondria organelles and is of vital importance to the division of cells into new cells.

Extraordinarily these vesicles, with their cargo inside them, are carried along the ‘roads’ of microtubules by motors that walk!  They ‘walk’ with ‘two legs … stepping along just like a porter carrying some cellular material as cargo.’ ‘Kinesin, one of the best-studied molecular motors, walks with precise steps of 8 nanometres.’ They can go forwards and backwards. Some molecular stepping motors even have a gear system!

The sketch below, not drawn to scale, illustrates how vesicles are carried along the 25nm-wide microtubules. They move at about 100 steps a second. The Kinesin particle, some 70nm long, has two ‘feet’ powered by ATP molecules. It also has two ‘hands’ that grip its cargo – sometimes a vesicle that may well be over 500nm wide. The cargoes thus can be far wider than the ‘roads.’ Several kinesin working together can drag whole strands of DNA.  Illustrations of this may be found on a Max-Planck-Institute of Molecular Cell Biology and Genetics website. 

The microtubule ‘roads’ are linked to a finer network of actin threads. A scientist researching them said: ‘Microtubules (MT) are like freeways and actin filaments are like local streets.’ Both networks constantly carry thousands of moving particles.‘

Another study reported: ‘Eukaryotic cells create internal order by using protein motors to transport molecules and organelles along cytoskeletal tracks. Recent genomic and functional studies suggest that five cargo-carrying motors emerged in primitive eukaryotes and have been widely used throughout evolution.’

In 2006 Drs. Andrew Z. Fire and Craig C. Mello jointly won their Nobel Prize for Medicine for describing how cells use another extremely important messenger – the ‘messenger RNA’ (mRNA) particle. It carries information in the form of double-stranded RNA to control the making of proteins ‘involved in all processes of life, for instance as enzymes digesting our food, receptors receiving signals in the brain, and as antibodies defending us against bacteria.’ The cells also make smaller  ‘microRNAs’ capable of carrying only 20 or so base pairs of code.   Our cells thus control and regulate the genes that were previously thought to reign supreme.

In 2006, discoveries in cellular biology won not only the Nobel Prize for Medicine but also the Nobel Prize for Chemistry. The latter went to Roger D. Kornberg: ‘for his fundamental studies on how the information stored in the genes is copied and transferred to the parts of the cells that produce proteins.’ In his Nobel speech he emphasised that, if this communication ‘is interrupted, the organism will soon die, since all protein production in the cells ceases.’ He added: ‘Many illnesses – like cancer, heart disease, and different kinds of inflammation are linked to disturbances in the transcription process’ that is vital to these cellular communications.

Cells also make larger elements known as transposons and retrotransposons. These can travel within their nuclei. They are the tools by which the cells adjust their genetic codes to meet environmental challenges and may contain  up to five thousand ‘base-pairs’ of code. They can carry the code for cellular genes, help regulate our genes – and even re-wire our gene control systems! For example: ‘Many retrotransposons carry enhancer sequences responsive to host gene regulatory systems so that they are capable of rewiring the regulation of adjacent genes-perhaps another example of ‘genomic altruism.’ (A contradiction to the theory of the selfish gene?)

The genomes of plants are mostly made of such transported codes.  As for humans, it seems from the evidence left within our genome, that around 42% of it has been moved around by transposons or retrotransposons at one stage or another of our evolution.

THE CREATION OF RETROVIRUSES

Cells do not only make internal transports. We now know that they also make particles that travel through ‘extra-cellular space’ to other cells, not to ‘perniciously infect’ them, but to pass information to them. We are multi-cellular organisms and inter-cell communication is absolutely vital to us. An adult human contains approximately 100,000 billion cells and for us to survive, these must ‘talk’ to one another, learn from each other, share and cooperate.

A cell can transform its retrotransposons, give them the ability to travel between cells with their variable load of genetic codes, make them retroviruses, by simply appending to each an additional piece of code.  An ‘intracellular, non-infectious retrotransposon’ becomes ‘a budding, infectious retrovirus merely by appending a retroviral MA domain.’ This may well be how retroviruses first evolved.

They leave their home cell by ‘budding’ from it.  On arrival at another cell, the codes they carry are incorporated into that cell’s DNA – and with this these retroviruses as such cease to exist. They have served their function. They are strictly one-use vehicles. Our genome has been thus constructed in part from codes created by other cells over a very long period of time. In other words, this system is vital to evolution.

Retroviruses can carry a wide variety of messages. ‘Retroviral particles contain a variety of cellular RNA’s’. The scientists who noted this, added that these  ‘are presumed to be packaged fortuitously during virion [virus] assembly.’   But this is only a presumption made because the authors did not see a reason for the presence of these codes.   I would ask if the ‘parent’ cell put these RNAs into retroviruses to have them taken as cargo to other cells?

The vesicles leaving cells do not all swim free. Recent micrographs, such as that reproduced below, reveal slender ‘nanotube’ bridges slung from cell to cell, carrying proteins, organelles and encoded information. ‘Transfer of molecules and organelles can occur directly from the cytoplasm of one cell to that of the other.’ Note the particle on the connecting nanotube. From 50 to 200nm wide, these connections might accommodate a retrovirus.  We do not yet know if retroviruses do, but vesicles have been observed moving along them from one cell to another.

These nanotube ‘highways’ are made of actin (once thought to be a constituent of HIV) and can link immune cells together. It is reported: ‘We think the cells make these connections so that they can work in a coordinated fashion to collect antigens from pathogens rather than working as individuals. This would seem to make the likelihood of successful delivery of the antigen to a distant lymph node much more likely.’

This nanotube network also extends within the cell to carry the vesicles that move by ‘stepping.’ ‘Mitochondria and intracellular vesicles, including late endosomes and lysosomes, could be detected within thick, but not thin, membrane nanotubes. Analysis from kymographs demonstrated that vesicles moved in a stepwise, bidirectional manner at 1 µm/s, consistent with their traffic being mediated by the microtubules found only in thick nanotubes.’

It seems every year we are learning more about how vital and complex is the cell’s transport system.  Some of the particles they send out mineralise the spaces between cells to create bone and cartilage. So surely it is not too far fetched to suggest that retroviruses also play a vital role in carrying codes from one cell to another?

The National Institutes of Health (NIH) reported that retroviruses ‘are so irregular and so labile that we have been unable to apply the tools of structural analysis to good effect.’ It also reported that retroviral DNA ‘closely resembles a cellular mRNA’ messenger vesicle. Retroviruses are also said to be ‘unique among animal viruses in that some groups exhibit considerable polymorphism in receptor usage.’  They are thus particularly well suited for carrying messages – as they can deliver ‘irregular’, or varying, code ‘similar to’ mRNA to many kinds of cellular receptors.

Today, I am glad to say, many biologists are no longer automatically naming all such travelling elements as ‘viruses’.  Many of these now called ‘exosome vesicles,’ a name first given them in 1997. These are generically described as ‘cargo-loaded small vesicles released into extra-cellular space,’ a description that surely applies to all viruses? Scientists varyingly describe them as particles of a width ‘up to 120nm’, ‘from 40 to 100nm’, from ‘60 to 90nm’ or even ‘up to 150nm’. They thus include vesicles of the size of the typical retrovirus, from 80 to 120nm wide, as well as sizes pertaining to other viruses.  In 2006-7 several scientists placed the retroviral family itself among the exosomes.

All kinds of cells make exosomes, including T-Cells – and, it seems, often for very good reasons.  In an important experiment, ‘exosomes secreted by bone-marrow-derived dendritic cells were challenged with tumour-derived peptides, activated CTLs, causing the eradication of established tumours.’  When near tumour cells, exosomes are reported to sometimes produce very strong anti-tumour reactions. Radiation-damaged cells also produce exosomes, perhaps as a genetic code repair mechanism.  They are also thought to help against streptococcus pneumonia bacteria and to generally stimulate our immune systems, including T-cells. They can transport antigens that protect us.

When I read the above reports, it made me wonder if this explains why retroviruses were initially discovered near tumour cells. They could be there to help cells repair themselves. It might also explained why they are found entering and leaving T-cells – a phenomenon long associated with ‘HIV infection.’ It could be that they are there for a very different reason – to help these cells fight pathogens.

Exosomes are now called ‘one of the most important protein complexes’ involved in controlling the ‘RNA-processing machinery’ in mammals. They vitally help ensure accuracy in the reading of RNA messages and help deactivate old RNA messages that are no longer needed. Because of their many functions, they are also called ‘secreted organelles’ – external cellular organs. (Yet, ‘anti-retroviral therapy’ is based around the out-of-date notion that all retroviruses are useless or dangerous particles.)

In 2007 it was described how ‘cells send RNA messages to each other by packing these into exosomes’ and how exosomes can carry a ‘large amounts of RNA’ from one cell to another – up to 1,300 different mRNAs! Among these are vital messages that ‘regulate cellular development and protein synthesis’ – in other words regulate some of the most important functions in our bodies and in all multicellular organisms.

Professor Peter Duesberg at Berkley was the first to describe the genome of retroviruses as `I have mentioned. He wrote of how their genetic codes ‘are integrated as proviruses [viral DNA] into the germ line of most if not all vertebrates’ after being carried from one cell to another. He also described them as totally harmless.

In summary:  retroviruses, like retrotransposons and messenger RNA (mRNA), carry information encoded into double-stranded RNA. They are formed inside cells on membranes.  They are then budded out through the cell wall, which, on the way through, donates to them their protective coating of proteins.  On arrival at another cell, their RNA is passed inside, converted and incorporated into that cell’s library of DNA.

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For plant retroviruses, see Plant retroviruses: structure, evolution and future applications, https://tspace.library.utoronto.ca/handle/1807/1315 For retroviruses from several fish species, see http://www.ncbi.nlm.nih.gov/books/bv.fcgi?rid=rv.section.8063

‘Exosomes express cell recognition molecules on their surface that facilitate their selective targeting and uptake into recipient cells.’ From ‘Exosomal transfer of proteins and RNAs at synapses in the nervous system.’ Neil R Smalheiser. Biol. Direct; published online Nov. 30th. 2007

  Definition of cytolysis from Biology Online.  http://www.biology-online.org/dictionary/Cytolysis

  For example, see http://www.kcom.edu/faculty/chamberlain/Website/Lects/MECHANIS.HTM

and also  http://mansfield.osu.edu/~sabedon/biol2065.htm

http://www.ncbi.nlm.nih.gov/books/bv.fcgi?rid=rv.section.285

Mae-Wan Ho This e-mail address is being protected from spambots. You need JavaScript enabled to view it Bioenergetics and the Coherence of Organisms   Neuronetwork World 5, 733-750, 1995.

LabNotes  http://www.mbl.edu/publications/pub_archive/labnotes/2.3/Langford.html
In recent video tapes made by Langford and colleagues with high-powered light microscopes, packages could be seen travelling along microtubules.

Eugen A. et al.  Puzzles of the living cell… Digest Journal of Nanomaterials and Biostructures Vol. 1, No. 3, September 2006, p. 81 – 92 http://www.chalcogen.infim.ro/Preoteasa.pdf

Fabrice Dumas  et al. An Agrobacterium VirE2 channel for transferred-DNA transport into plant cells. Proceeding of the National Academy of Science, USA January 2001.

  Nature Cell Biology  3, 473 – 483 (2001) Published online: 18 April 2001; | Caveolar endocytosis of simian virus 40 reveals a new two-step vesicular-transport pathway to the ER Lucas Pelkmans, Jürgen Kartenbeck & Ari Helenius

http://www.ncbi.nlm.nih.gov/books/bv.fcgi?highlight=virus,golgi&rid=rv.section.2497

http://www.tifr.res.in/~roop/NaturesNanotech.htm

http://www.emccd.com/downloads/?doc=Watching%20Single%20Motor%20Molecules%20at%20Work.

How Does Intracellular Molecular-Motor-Driven Transport Work?
http://eiffel.ps.uci.edu/cyu/talks/actin04.ppt

  LabNotes Fall 1992. http://www.mbl.edu/publications/pub_archive/labnotes/2.3/Langford.html

http://www.ncbi.nlm.nih.gov/pubmed/12600311?dopt=AbstractPlus

Processing of Pre-microRNAs by the Dicer-1-Loquacious Complex in Drosophila Cells.  Kuniaki Saito. Akira Ishizuka Haruhiko Siomi Mikiko C. Siomi

1 Institute for Genome Research, University of Tokushima, Kuramoto, Tokushima, Japan.  PLOS Biology.

  Our double stranded code is made up of nucleotides that pair themselves with nucleotides on the other strand. Each of these couples is called a  ‘base-pair’.

White et al. 1994

Lander ES, Linton LM, Birren B, Nusbaum C, et al. Initial sequencing and analysis of the human genome. Nature, 2001; 409(6822): 860-921

Higher-Order Oligomerization Targets Plasma Membrane Proteins and HIV Gag to Exosomes
Yi Fang, Ning Wu, Xin Gan, Wanhua Yan, James C Morrell, and Stephen J Gould. Department of Biological Chemistry, Johns Hopkins University School of Medicine, full text available at
http://www.pubmedcentral.nih.gov/articlerender.fcgi?artid=1885833#pbio-0050158-b058 – also Denzer K, Kleijmeer MJ, Heijnen HF, Stoorvogel W, Geuze HJ. Exosome: from internal vesicle of the multivesicular body to intercellular signaling device. Journal of Cell Science 113, 3365-3374 (2000). The MA domain is added to the ‘N-terminus of its Gag-like protein.’

David J Griffiths  Endogenous retroviruses in the human genome sequence. Genetic Biol. 2001; 2(6): reviews1017.1-reviews1017.5. Published online 2001 June 5.

http://www.ncbi.nlm.nih.gov/books/bv.fcgi?highlight=ribosomes,cell&rid=rv.section.2542#2547

Similar arguments also apply to the exosomes as noted later.

Demontis F (2004) Nanotubes Make Big Science. PLoS Biol  Published: July 13, 2004

Demontis F (2004) Nanotubes Make Big Science. PLoS Biol 2(7): e215 doi:10.1371/journal.pbio.0020215 Image courtesy of Hans-Hermann Gerdes

Nanotrubes link immune cells. The Scientist, 20 September 2005.

Björn Önfelt et al ‘Structurally Distinct Membrane Nanotubes between Human Macrophages Support Long-Distance Vesicular Traffic or Surfing of Bacteria The Journal of Immunology, 2006, 177: 8476-8483

Bonucci, E. (1967), “Fine structure of early cartilage calcification”, Journal Ultrastructure Research, 20: 33-50

Exosome ‘vesicles’ are not be confused with the unrelated ‘exosome complex’ that helps break down RNA within cells.

Martin P. Bard, Joost P. Hegmans, Annabrita Hemmes, Theo M. Luider, Rob Willemsen, Lies-Anne A. Severijnen, Jan P. van Meerbeeck, Sjaak A. Burgers, Henk C. Hoogsteden and Bart N. Lambrecht; Proteomic Analysis of Exosomes Isolated from Human Malignant Pleural Effusions American Journal of Respiratory Cell and Molecular Biology. Vol. 31, pp. 114-121, 2004

Nicolas Blanchard*, Danielle Lankar*, Florence Faure*, Armelle Regnault, Céline Dumont*, Graça Raposo and Claire Hivroz2,* TCR Activation of Human T Cells Induces the Production of Exosomes Bearing the TCR/CD3/ Complex1 The Journal of Immunology, 2002, 168: 3235-3241.

The Journal of Immunology, 2001, 166: 7309-7318. Proteomic Analysis of Dendritic Cell-Derived Exosomes: A Secreted Subcellular Compartment Distinct from Apoptotic Vesicles Clotilde Théry,, Muriel Boussac,, Philippe Véron*, Paola Ricciardi-Castagnoli, Graça Raposo, Jerôme Garin and Sebastian Amigorena*

http://www.ncbi.nlm.nih.gov/books/bv.fcgi?rid=rv.section.285

Denzer et al. op.cit. – cites this experiment as carried out by Zitvogel et al., 1998.

Banchard et al. 2002.  Cited above.

Infection and Immunity, January 2007, p. 220-230, Vol. 75, No. 1

Dendritic Cell-Derived Exosomes Express a Streptococcus pneumoniae Capsular Polysaccharide Type 14 Cross-Reactive Antigen That Induces Protective Immunoglobulin Responses against Pneumococcal Infection in Mice

Jesus Colino and Clifford M. Snapper*

Houseley J, LaCava J, Tollervey D. RNA-quality control by the exosome. Nat Rev Mol Cell Biol. 2006 Jul;7(7):529-39. PMID: 16829983

Raijmakers R, Shilders G, Pruijn G.  The Exosome: a molecular machine for controlled RNA degradation in both nucleus and cytoplasm.  European Journal of Cell Biology, Vol. 83, 5 July 2004. Pp 175-183

Yi Fang, Ning Wu, Xin Gan, Wanhua Yan, James C Morrell, and Stephen J Gould; Higher-Order Oligomerization Targets Plasma Membrane Proteins and HIV Gag to Exosomes. PLoS Biol. 2007 June; 5(6): e158. Published online 2007 June 5

‘The Exosome Exchange.‘ The Journal of Cell Biology. 21 May 2007.  Also, Denzer K et al. Exosome: from internal vesicle of the multivesicular body to intercellular signaling device’ Journal of Cell Science, Vol 113, Issue 19 3365-3374,

Retroviruses as Carcinogens and Pathogens: Expectations and Reality By Peter H. Duesberg.  Cancer Research, Vol. 47, pp. 1199-1220, (Perspectives in Cancer Research), March 1, 1987.

Graziella Griffith, and Marie-Christine Dokhélar Arielle R. Rosenberg, Lélia Delamarre, Claudine Pique, Isabelle Le Blanc,  Early Assembly Step of a Retroviral Envelope Glycoprotein: Analysis Using a Dominant Negative Assay

Cell Biol., Volume 145, Number 1, April 5, 1999 57-68