Are Alive?

Patrick Forterre

Université Paris -Sud (Institut de Génétique et Microbiologie) Institut Pasteur (Département de Microbiologie) Spiraviridae Are Viruses Alive?

Is it an important issue? Most virologists assume « implicitely » that viruses are alive

They commonly speak of the « cycle » in their papers

Most biologists don’t care…………..many of them even don’t care about viruses at all!!

Biologists with philosophic background (and/or interested in exobiology ) usually consider that viruses are not living

(Marc Vanregenmortel, Michel Morange, Purificacion Lopez-Garcia, David Moreira, Antonio Lazcano, Testart)

Michel Morange Problems raised by the definition of life , In « Origins and evolution of life, an Astrobiology perspective », pp3-13 Gargaud, Lopez-Garcia and Martin, eds. Cambrige University press, 2011

Introduction: « I will show that the recent suggestion that viruses are alive makes no sense and obscures discussions about life »

« Examining the case of viruses, the absence of «viruses remain strict a definition leaves us defenceless when absurd parasites » statements are made on the living character of such objects » « there is no symmetry between a virocell and a «To consider viruses as living creates much ribocell. The first needs the useless confusion » second: the reverse is not true » There is no symmetrie between carnivors et les herbivors, The first needs the second: the reverse is not true

HYPOTHESIS: there is a psychological aspect in this debate

We don’t want to be classified in the same category than viruses

Ribocentrism

«To consider viruses as living creates much useless confusion » (Morange)

To consider viruses as non living, but by-products of life led to ignore them or to 513 pages underestimate their role in life history Nothing else on viruses??? My hypothesis: scientists who consider that viruses are not alive tend to underestimate (denigrate) viruses ……………and make false statements

% of in clusters of Virus- orgin

% of genes in gene clusters of unknown origin

Cortez, Forterre and Gribaldo % other genes Biology, 2009

6% 81%

13%

Ori C

a → c → g → a → t

About 20% of genes in archaeal and bacterial originated in viruses or DePerseval and Heidman, Cytogenet viral origin Genome Res, 2005

?

repeated

40% - derived elements) 8%% retrovirus

2% (exon) 15% eukaryotic genes (introns) Total sequences: 196 kb, 223 genes, 168 168 different proteins different proteins 103 singletons pGT5 (3.6) pRT1 (3.4 ) 36 proteins with homologue(s) in pTN1 (3.3) at least one other plasmid/virus* pTN2 (13) (6) PAV1 ( 18 ) pP12-1 (12) TPV1 ( 24 ) pT26-2 (25) 59 with homologues in cellular pAMT11 (20) genomes* pTN3 (18) PCIR10 (13) 58 of them being encoded by PIRI48 (13) genes present in integrated pEXT9a ( 10,5 ) viruses/plasmids (archaeal pIRI33 (11) proteins of viral origin ) pAMT7 (8.5) 1 protein of cellular origin (MCM )

•Mostly in Thermococcales or in Methanococcales or Thermococcales, one also in Sulfolobus (integrase), one in an Haloarchaeal plasmid The gene flux from viruses to cells is a very ancient on-going process : a lot of cellular proteins should have a viral origin

viruses

cells Cells are giant pick-pockets of viral genes Scientists systematically favour transfer from cells to viruses in the interpretation of phylogenetic analyses.

Implicit (false) assumption: new genes cannot originated in viral lineages RNA polymerase

Ancient cells (fourth domain) Megavirales

Boyer et al., Plos One, 2011 Williams et al., Plos One, 2011

Modern cells () Megavirales

Ancient Megavirales Modern cells (Eukaryotes/) Many viruses encode DNA replication proteins that are very divergent from their cellular homologues Escherichia coli Topo IV

Bacillus subtilis Prochlorococcus marinus Bactéries Deinococcus radiodurans Escherichia coli DNA gyrase Bacteriovirus T4

African Swine Fever (NCLDV) Chilo iridescent virus (NCLDV) Virus Trypanosoma brucei PBCV-1Chorella virus (NCLDV) Plasmodium falciparum Caenorhabditis elegans Eukaryotes Encephalitozoon cuniculi Saccharomyces cerevisiae Arabidopsis thaliana Type IIA DNA Caenorhabditis elegans Topoisomerase Homo sapiens Drosophila melanogaster s

Hs LE HILL RPDT YIG SVELVTQQMWVYDEDVG-INYREVTFVP GL YKIFD EILV NAA DNKQRDP-KMSC- IRVTMIRKQLISIWNNGKGIP E Tb IE HVLTR PEM YIG SLDTTATPMFIYDEQKGHMVWE TVKLNH GL LKIVD EILL NAS DNISNRSARMTY-IRVTITDTGEITIENDGAGIP T4 T4 IE HIKK RSGMYIG SSANETHERFMF----GK--WE SVGYVP GL VKLID EII DNSV DEGIRTKFKFANKINVTIKKNNQVTVEDNGRGIP Ec LDAVRK RPGMYIG DTDDGT------GL HHMVF EVV DNAI DEAL---AGHCKEIIVTIHADNSVSVQDDGRGIP B Bs LEAVRK RPGMYIG STNSK------GL HHLVW EIV DNSI DEAL---AGYCTDINIQIEKDNSITVVDNGRGIP Topo IB DNA topoisomerases

Poxviridae

Brochier-Armanet, Gribaldo, Forterre, Biology Direct, 2008 Eucarya Archaea

Indiani and O'Donnell Nature Reviews Molecular Cell Biology 7, 751–761 (October 2006) | doi:10.1038/ nrm2022 | Bacteria Archaea/ Eukarya

Baculovirus NCLDV Herpes T4

One transfer before LUCA followed by a Forterre, P. non-orthologous replacement in Current Opinion in Microbiol. 2002 bacteria

ancient virosphere

a DNA-LUCA?

BACTERIA ARCHAEA EUKARYA We know that many viruses have « invented » new forms of DNA

C

DNA HMC

HMC-DNA

To protect their genomes against nucleases encoded by their cellular « hosts Hypothesis: RNA viruses have invented DNA to protect their genomes. exactly as some modern viruses have modified their DNA.

ThyA RNA U-DNA T-DNA HMC-DNA RNR ThyA-like

If this is true, viruses have played a major role in the origin and evolution of modern cells My hypothesis: scientists who consider that viruses are not alive tend to underestimate (denigrate) viruses……………and make false statements The double-jelly-roll fold

The PRD1/Adenovirus lineage

The Hong-Kong fold

The HK97 lineage Viruses are ancient and co-evolved with their victims, (hosts?)

Prangishvili, Forterre and Garrett, Nature Rev Microbiol. 2006 It’s not because scientists who consider that viruses are not alive tend to underestimate them ……………that viruses are alive!!!! Metagenomic and genomic analyses data have shown that most genetic information in the biosphere is probably present in viral genomes and derived elements (plasmids)

If viruses are not living, this means that most genetic information on our planet has been created by non living entities!! Viruses and related elements (plasmids) are the major source of variations by manipulating genomes and creating new functions

Viruses put an enormous selection pressure. They control the number and type of cells in natural environments…… Viruses are the major agents of natural selection Raoult D. and Forterre, P. Nature Rev Microbiol, 2008

Ribosomes Encoding Organisms

Capsid Encoding Organisms

If you get the sequence of a 1 Mb genome from metagenomic data, this definition allows you to distinguish between cellular and a viral genomes Krupovic and Bamford, J Virol 2010

A virus is defined by its (the viral self sensu Bamford) Raoult D. and Forterre, P. Nature Rev Microbiol, 2008

Ribosomes Encoding Organisms

Capsid Encoding Organisms

If you get the sequence of a 1 Mb genome from metagenomic data, this definition allows you to distinguish between cellular and a viral genomes

are viruses organisms? are viruses alive? No, if we confuse the virus with the virion

Viruses ( virions ) are inert until they encounter cells, they have no metabolism, they cannot produce energy

Viruses ( virions ) can cristallize (Tobacco mosaic virus) Viewed in TV Chamberlain’s filter (1884) VIRUSES and GENES Jacob and Wollman Scientific American. 1961 Jun;204:93-107,

A virus may exist in three states:

-the extracellular infectious state

-the vegetative state of autonomous replication

-the proviral state »

«A virus can be considered a genetic element enclosed in a protein coat » This traditional view of “ viruses as virions ” has been criticized by Claudiu Bandea in 1983 and more recently by Jean-Michel Claverie

The virus should not be confused with the virion

Claudiu Bandea Jean-Michel Claverie (Journal of Theoretical Biology, (Genome Biology, 2006) 1983)

Bandea : The living phase of the virus is the intracellular phase of its life cycle Claverie : The virus is the viral factory

The viral factory reorganize cell metabolism, energetic, and all cell synthetic functions to produce virions

Mimivirus viral factory reorganize cell metabolism, energetic, and all cell synthetic functions to produce virions This is not a virus This is a virus

This are not Humans These are Humans Origin of the eucaryotic nucleus Takemura, J. Mol. Evol. 2001 Bell, J. Mol. Evol. 2001

Viral factory

the Nucleus of a virocell nucleus But…..the concept of viruses as viral factory cannot be generalized because viruses infecting archaea and bacteria dont produce obvious viral factories The virus transforms the cell into a viral factory (Lwoff, 1961)

André Lwoff The virus transforms the The organismal form of the cell into a virion factory virus is the virion factory

Forterre and Prangishvili, Research in Microbiology (2009) Ann N Y Acad Sci. , (2009)

The organismal form of the virus corresponds to the infected cell, which has been transformed into a new organism = a virocell The virocell: a cell whose dream is not to produce two cells but to produce virions

The virion : a vehicle to disseminate viral genes

The virocell : the living form of the virus Ribosome encoding organisms (REO) ribocells

Reproduce by cell division

Capsid encoding organisms (CEO) : virocells

Reproduce by production of virions Cells producing virions but that can still divide are « ribovirocells »

The virocell should not be confused with the virus

The virocell corresponds to one « existential state of the virus »

A virus may exist in three states:

-the extracellular infectious state

-the vegetative state of autonomous replication

-the proviral state »

Some viruses can infect different hosts, forming different types of virocells One should not confuse cells and organisms

Mamavirus

Amoeba

Sputnik

Bacteria Forterre: Giant viruses: conflict in revisiting the virus concept A dynamic RiboVirocell Intervirology, in press (2010)

A cell can harbor several different living organisms A living organism is always associated to one evolutionary lineage with an history that can be represented by a tree-like structure

Mamavirus

Amoeba

Sputnik

Bacteria marc van regenmortel

Some viruses that infect cyanobacteria and degrade their chromosomes encode proteins involved in photosynthesis to replace those of the cell

A photosynthetic virocell

Braggs and Chisholm, Plos One, 2008

Virions of photosynthetic viruses Genome can be amplified in the virocell from 100 to 1000 times

Mew viral genes (proteins) can originate by: point mutations, duplication, recombination, slippage, frameshift, overlapping

Viruses are not only vehicules New proteins also originated of cellular genes in viral lineages

Virus-first hypotheses for the origin of life cannot be correct Viruses cannot have appeared before cells……since they are cells themselves

But of course viruses might have appeared before modern cells , i.e. before LUCA Schematic history of life Origin of Life (first « cell »)

First RNA cell

« invention » of the ribosome First RNA -Protein cell

ORIGIN OF VIRUSES

« invention » of DNA LUCA

Eukarya Archaea Bacteria A simple hypothesis for the origin of viruses: the ancient escape hypothesis

Different mechanisms triggered the formation of different types of virions to disseminate RNA genomes in the late RNA world (after the invention of proteins).

Some ribocells became ribovirocells and later on virocells

Forterre and Krupovic, the origin of virocell, book chapter, in press Viruses are evolved by cells

Moreira and Lopez-Garcia, Nature Rev microbio (2009)

Viruses evolved within cells (in the virocell) Violent reactions to the virocell concepts

« conceptual trick » « artifice»

«epistemological cheating» The remore indeed does The remore stay not transform the shark outside!!!! Very different into a remora- from the virus situation shark…!!!!! A more interesting example: Chlamydiae

The cellular structure of the Chlamydiae is conserved all along its life cycle

Unlike viruses, he Chlamydiae ( a ribocell ) does not borrow the cellular structure of its victim

Viruses have unique life cycle properties that justify the class them in a categorie different from any other types of cellular parasite (capsid encoding organisms) What is life?

It’s not a scientific question

It’s a philosophical question

We need a philosophical framework My personal philosophical framework

Life is the mode of existence of albuminoïd bodies Friederich Engels (Dialectics of nature, 1883)

Life is the mode of existence of living organisms So…what is a living organisms ?

Two approaches:

We decide what is a living organism and We look if specific biological entities (viruses?) fit with our definition

We decide which biological entities are living and we propose a definition based on their properties

2008 : A collection of integrated organs (molecular machines) producing individuals evolving through natural selection

2012 A living organism should belong to a evolutionary lineage characterized by a life cycle with at least one cellular phase Life ? ?

?

Are plasmids living organisms ?

This is not a bacterial penis, but a plasmid made dissemination system

Plasmids are only made of nucleic acid. They probably originated from viruses that lost Bacteria have no sex! It’s a their capsid mode of plasmid dissemination