Now that we have seen some of properties of living cells that are alive we can ask...

                                     What is the origins of cellular life? ? ?

  ORIGIN of LIFE ???  is a Paradigm Question of Cell & Molecular Biology?
  the origin of a Primordial Cell... was it chemical evolution, an astrobiological event,  or  what?  
       
 CURRENT PARADIGM...                                                                a text decscription of the origins of the first cells.*read this
     Most experimental evidence favors a Chemical Complexity-Evolutionary Adaptation Origins of Life... 
                     "chemical self-assembly has lead to complex self-replicating molecular systems "
 
          some timelines:     Age of Universe, the Milky Way, and the Earth*          
                                         between 4.5 to 4.0 bya - asteroids bombard & sterilize planet's surface
                                         then by  3.5 bya  - first fossil evidence of microscopic life is present [stromatolites]

     Initial chemical events were likely the: AGGREGATION of CARBON BASED MOLECULES...      
     Carbon shares 4 unpaired electrons easily forming carbs, lipids, proteins, & nucleic acids:

     The exact circumstances of the origin of life and the actual nature of first organisms may be
     forever lost to science, but research can at least help us understand what is possible because
      the hypothesis of chemical evolution of life conforms to the scientific method and is...
                                                 EXPERIMENTALLY TESTABLE... 
 
      Let's start with a Definition of Life & a look at Experimental Approaches to Origins of Life*  


        


 

 


 


 Definitions of Life:

   There is to date not a single agreed-upon definition of cellular life, even though there are
some 123 published definitions of life, many of which group defining terms into one short
statement:   Life is self-reproduction with variation (evolution).

   So how could it originate? This question lies at the core of understanding the cell
as the smallest living unit. Presumably, there wasn’t a cell right after the Big Bang,
so where did the first one really come from?        
   What did the molecules of Life (on Earth or elsewhere in Universe) look like before life
made its first appearance? & did they self-assemble & self-organize into a 1st cell-like entity? 

   Origin-of-life research has followed 2 main experimental approaches:   

     1)  a “METABOLISM FIRST” approach,
               in which chemical cycles may have led to the synthesis of reactive organic molecules. 
               This hypothesis is based upon speculative pre-biotic chemistry...

     2)  a “GENETICS FIRST” approach,
               which favors the formation of some sort of replicating information molecule as being 
               a necessary prerequisite for life.

               Did one precede the other or are both congruent?
   




   











  
   
    Let's look at the LABORATORY EXPERIMENTAL APPROACHES in Origin of Life Research...
  
    A)  Monomers are formed from a chemically reactive soup... in volcanic "ponds" on a primitive Earth?
                     a hypothesis that was 1st suggested by Charles Darwin.


           1828-1953 - Can we make biomolecules in the lab?   timeline of in vitro  organic syntheses*       

          1920s - Alexander Oparin proposes carbon based organics are dissolved in a primordial ocean soup &
                       an infant Earth possessed a strongly reducing atmosphere, containing CO2, CH4, NH3, H2-gas
                       and water vapor, and little O2....               Were these the raw materials for the evolution of life?
 
          1953 - Miller & Urey*...  achieve abiotic synthesis of organics in lab experiments  (Science 1953)
                   H2O, NH3, CH4, & H2 to make HCN & formaldehyde: then amino acids*, nucleotides & sugars

           2007 - repeat of Miller-Urey famous experiments*

           2008 - Jeffery Bada discovers 1953 samples*

           2011 - Primordial synthesis of amino acids in spark discharge experiment supports abiotic syntheses*       

      
   
              
                                                                         

 

 

 

 

 

 

 

 

 

 

 

 

 

       
 
B) 1977 - Deep dwelling (ocean) hydrothermal*vents...  (also known as deep sea volcanic plumes)  
               >  vents are full of organically rich carbon based molecules
                         - and constitute an ecosystems of life without light energy:  bacterial, worms, crabs, etc*
                               -  maybe even a LUCA? = a chemosynthetic thermophilic H2-consuming archaea?
                                               LUCA is assumed to be an anaerobic, extremeophile, CO2-fixing,
                                        H2-dependent, FeS containing thermophilic organism. Methanocaldococcus*
     1992 - Iron Sulfur World... C. Huber & G. Wachtershauser propose a beginning for metabolism...
                     key organic molecules may have originated at vents with superheated water (1000C +).
                          - a primary mechanism of abiogenesis might be H2S + FeS --> FeS2 [pyrite] reacting with
                            CO2, NH2 & H2S and passing over catalytic transition metals [FeS & NiS] favoring
                                    reduction of N2 to NH3 & CH3-SH [mercaptan] & possibly peptide bonds.

      Speculation*:  chemosynthesis may have helped life originate in vent-like regions... 
       
                                    a further controversy today is -      Vents    vs.     warm little ponds*.                                                      
    
                   a table of Prebiotic simulation experiments indicating it is a very active research area                                                                                   
                                                                            

 

 
 
 
 
 
 
 
 

 

 

 

 

 

   
  
   C) Evidence for an Astrobiological origin of Pre-Biomolecules...
 
                early Earth was constantly bombarded by asteroids & meteorites. It was a sterile habitat...  
    1          ideal for deposition of extraterrestrial organic materials  [bombarments & extinctions]
        SPACE DEBRIS...   may have deposited organic-like molecules, as carbon based aliphatics like
                             hexane or longer chains of C & H (space grease) on a newly formed Earth.
.....
           Comets in the inner solar system when heated by the Sun shed surface layers exposing prestellar  
 .....    
                     material that can be sampled remotely (spectroscopically) or in situ. Comets are mostly
                     ice crystals (40%) on cores of silicates (20%) & carbon as CO2 (5%) and also contain
                     about 20% organics [CO, HCN, CH4, CH3OH, NH3 & H2N-CH3 & H2N-CH2-CH3].
                    Xexon isotope signatures indicated that comets are not responsible for Earth's water.
                               a European Space agency probe of a comet -->  Rosetta & comet 67P  + results   
           Asteroids (rocks) containing organic molecules as...  PAH*- polycyclic aromatic hydrocarbon,
                    & nucleobases, quinones, quinones, COOH's, amines & amides = some 70 amino acids.
                             Collecting pristine asteroid sample materials*   and       Panspermia* 
       Meteorites are debris pieces of comets or asteroids that originated in space & hit Earth:
                             Murchison meteorite contents*  Australia 9/28/1969 - heavily analyzed. 
                .
    thus we have 3 possible sources for the initial abiotically produced metabolic bioorganic molecules:
                     1.  abiotic (Miller exp),      2.  hydrothermal vents,   or    3.  astrobiological.
   

 












 
 
2nd)  a 2nd Experimental Approach used in origin of Life studies:
            Build an artificial  MODEL  MOLECULAR  REPLICATIVE  SYSTEM
 
      the RNA WORLD Hypothesis...   1st proposed by J.B.S. Haldane (1963), who suggests that the first
                  living microbes used RNA as a guide to build proteins. In 1986 Walter Gilbert, a nucleic acid biochemist
                  coined the term RNA World to describe the hypothesis.
       the hpothesis emanates from debate of which came 1st DNA or RNA?
                  Gilbert argued for an emergence of RNA molecular replication as both an information-carrier molecule
                            and a catalytic molecule, as an important 1st step in making a replicative molecular  system...
       
        Observations related to replicative molecular systems:                                                               
                ►  in 1989  Sidney Altman & Tom Cech - received Nobel Prize for demonstrating in 1982
                                  that RNA molecules have CATALYTIC ACTIVITY...  (RIBOZYMES*).   
                                  some RNA's  catalyze polymeric cleavage in a sequence-specific way,
                                  by phosphoryl transfer reactions or hydrolysis of phosphate esters.
               ►  Maybe they might have also be capable of ligation* of RNA fragments together...
              ►  the roles* that cellular RNAs & Ribozymes have in today's cells supporting the RNA World?
              ►  Researchers have produced (in vitro) short chains of RNA 2000+ nucleotides long
              ►  clay minerals* might have enhanced a primitive way of concentrating reactive molecules
                                           facilitating formation of 1st RNAs & vesicles* leading to protocells...
     Hypothesis:   if chains of RNA acting as templates & also catalyze polymerization of like molecules,
                             then RNA molecules might have been the 1st SELF-REPLICATING molecular entity...
                            then followed by molecular selection*              
                                                                                     
 
 
 

        

  

 

 

 

 

 


  
     A long-standing weakness of the RNA-world hypothesis was the inability to
spontaneously generate the RNA nucleotides from the basic ingredients presumed to
be available on a prebiotic Earth.  Ribose, the 5-carbon sugar of the RNA backbone,
had been difficult to form under prebiotic conditions, as have the nitrogenous bases.

           in May 2016 Thomas Carell et al described a reaction mechanism using
           formic acid & aminopyrimidines (also found in comets) under early Earth
           conditions made high yields of adenosine*, from which guanosine* may be made.
                      
Another hurdle has been attempts at synthetic POLYMER syntheses...

           in RNA polymerizations there  isn’t only one way*   for two nucleotides to bind.
           in 'cellular life' RNA polymerase assembles a 3’–5’ phosphodiester linkages.
 
   Synthetc ribozyme RNA Polymerases*  may have been the first genetic information
                replicators. They do satisfy two of the criteria for the bare minimal requirements
                needed for an origin of life: replication and change over time.

   in 2019 a novel approach -  Hachimoji Engineered DNA* -   [ Hachi  =  8      moji  =  letters ]      
                why does DNA have only 4 bases and not 8 or more+? 

                                                

 

 

     

 

   

 

 

   

  

       

 

 

 

 

 

 

 

 

 

 


 Another Experimental Approach has been:   to build molecularly active vesiclized "Protenoids or Protocells".
                   aggregates of abiotically made organic molecules surrounded by a lipid membrane
                             i.e., an artificially made vesicular metabolic system...
                     Molecules in solution are randomized & unable to maintain an organized state (order), yet compartmentation
                     allows concentration and increased reactivity. Earliest forms of life may have been aggregates of prebiotic
                     macromolecules in a boundary to maintain an interior chemical environment distinct from "primordial soup"...
        Sidney W. Fox (1912-1998) - U.M. Director of NASA supported UM Institute for Molecular Evolution...
                     his laboratory conducted analyses of the first moon rock samples & he produced proteinoids & microspheres
                     by dropping amino acid solutions onto hot lava rock or clay.
Vesiclized experimental systems include:  coacervates,  proteinoid microspheres,  &  liposomes.  



Coacervate droplets formed by polypeptides, polysaccharides,  nucleic acids &/or lipids...
            have tough skin of water molecules holds hydrophobic interior with osmotic capability.
            Could they have an enzymatically active interiors [fig*]   Protocell Formation Research*   
           Can Protocells divide?*                   a side bar on Coacervates and drug delivery
Proteinoids - abiotic protein polymers can form microspheres* (1-2um dia)...
            abiotic amino acid polymer chains formed microspheres in water, which were
            osmotically active shells with selective permeability & have membrane potentials.
Liposomesg form spontaneously from phospholipids,   figure-1*  &  figure2*,  which
            are microscopic spherical vesicles that form when phospholipids are hydrated.
            they can engulf smaller proteinoids making more larger & active ones.
            Synthitic lab produced liposome can synthesize new proteins (hemolysin*)
 ...
A plausible route to 1st cells via...
           vesiclization, lipid enhanced polymerization, and combinatorial chemistry.
 
 

 

 

 

 

 













 
 
 
 
 
 
 
 

 
 
 
 
 
 
 

 
 
 
 
 
 
 
 
Hypothesis: Did the innate physiochemical properties of early-Earth biomolecules
                    acting within elementary PROTOCELL-LIKE entities give rise to the
                    essential metabolism of cellular behaviors of today's cells ??? 
   
Some supporting evidence includes:
    - fatty acid like molecules have been have been found in meteorites
                 &  under a variety of prebiotic conditions fatty-like molecules can self assemble.
  
    - artificial protocells can be made in the lab by encapsulating a self-replicating genomes
                inside a chemically simple self-replicating membrane vesicle.
   
    - artificial lab-made vesicles encapsulating known active genome ribozymes do generate
                an genomic-like activity, and some artificial vesicles can "steal" membrane fragments
                from other vesicles with less active genome pieces   (model figure*).
                            such "genomic fitness" may have evolved into "cellular fitness ".
     
     - artificial vesicles can grow larger micelles, a transmembrane pH gradient can be
                observed, due to faster flip-flop of protonated FA's on outer leaflet.
                Acidification of a vesicle's interior can store energy in form of a
                pH gradient, without enzymes, a primitive energy mechanism.

     - Could a vesiclized RNA genome have made proteins and lead to a cell???   (model figure*). 
  














   the  GENETIC  experimental approach to Origin of Life Research...

    Synthetic Biology  &  Minimal Genome Cell Research... 
         the concept of synthetic biology uses the approach of dissecting biological systems
         into modular functional units, or “bio-bricks,” that can ideally be recombined to assemble
         independently existing or wholly new biological systems...  
         its challenge may be to construct an artificial organism that reproduces & evolves.

     1) Synthetic Biology... uses a top-down rather than a bottom-up approach* and assumes one
                                          can't truly understand what one can't build from scratch (abiotically?). 
                                          it's the construction of fully functional cells (or parts) from scratch. 
         goals:  to design simplified versions of Life's processes synthetically that can produce
                                          desired products with high efficiency;
                     to artificially assemble the molecular components to model life processes;
                     to understand why & how matter can self-organize and exhibit living properties.
                                   i.e., to engineer new genetic circuits, entire genomes, or organisms
                                          to make complex biological machines.
                                          Taking man-made genetic elements to the level of ENGINEERING a CELL,
                                          and/or altering genes & arrangements to make novel designer genes. 
      
    Synthetic biology: artificial creation of DNA, genes, viri, cells that mimic/surpass natural systems.

     the roots of synthetic biology* via reconstitution experiments go way back in molecular biology...
   
  




    
   












 
 
 
 



  

     The Top-Down molecular approach is used in the making of a digital chromosome     
           
  2)  KNOCK-OUT Cells... looking for a minimalist essential genome required to make a cell,
                                       leads to synthesis of an artificial functional genome...
         
  J. Craig Venter, a principle investigator (P.I.) of Human Genome Project began by trying to make
  a new type of synthetic bacterium using DNA made in the lab from the sequenced genes of
  a bacterium Mycoplasma genitalium,  a gram-positive parasitic bacteria, whose primary infection
  site may be the human urogenital tract and causes non-gonococcal urethritis.  It has a circular
  chromosome of 582,970 base pairs, the smallest known genome of any independently replicating
  organism.   M.g. has some 525 genes (~476 protein encoding genes & 41 RNA genes).
  
     > How many genes does it take to make an organism?    What is the minimum genes a cell needs?
       As part of a Minimal Genome Project (researchers in Venter's group JCVI) sequenced
       the Mycoplasma genitalium genome & followed this work by systematically inactivating its genes
       by silencing them with microRNAs* [via so called knock-out cells*], to see which ones are
       essential to life & which are dispensable].

       Of the ~476+ protein-encoding genes, they concluded that only about 382 of them are essential to life.
  
   




 

 

 

 


  
     Can one synthesize an artificial chromosome?
   
  > 1st step: construct an artificial or synthetic chromosome that has "ALL" the genes (1 million bp)
           using lab-made chemicals, & TRANSPLANT it into bacterial cell = new synthetic life form.
  
           Venter's group developed a strategy for assembling viral sized pieces to produce large DNA
           molecules that enabled us to assemble a synthetic Mycoplasma mycoides chromosome in four
           stages from chemically synthesized DNA pieces averaging about 6 kb in size. This was
           accomplished through a combination of in vitro enzymatic methods and in vivo recombination
           in yeast cells. They hoped to transplant it into a cell with its DNA removed.
 > current Synthtic Biology Efforts:    Organs-on-a-Chip
     
 > next step:  transplant the artificial chromosome* of Mycoplasma mycoides into a Mycoplasma
          capricolum recipient cell, which has its chromosome removed, creating new artificial DNA cells.
           These new cells that are controlled only by the synthetic chromosome. The only DNA in these
           new bacterial cells is the designed synthetic DNA sequence, including “watermark”
  
  > final step:  new organism created*with smallest synthetic genome of any known cellular life form.
                        A minimal Cell computer Model*   and   J. Craig Venter's Digital Lifeview@home   

  > current Synthtic Biology Efforts:    Organs-on-a-Chip* 

                        Summary of Origins of Life Experimentation* 
   next    next - Microscopy & Cell Structure                         
                  




  

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 


 

 

 

 

 

 

 

 

 

 

 

 








    SKIP all the material BELOW this point.... 


 But what does it take to evolve a Eukaryotic Cell as we know it today ?
   
                             the evolution of the eucarya was single most important step in evolution 
                                                     of multicellular life forms & was a key step that lead to plant & animal life. 
         these experiments approaches have not yet been attempted...
        1. membrane encapsulates genetic DNA...   development of nucleus
                              greatest evolutionary invention - it internalized the genome
        2. loss of a rigid cell wall...
                               cells developed ability of phagocytosis - allowed engulfing of foods
                              also allowed cells to clump together -->   multi-cellularity  -->   tissues
        3. evolve a selectively permeable membrane...
                              protects cell, allows uptake gases & nutrients & exchange with environment
        4. evolve a cytoskeleton...
                              provides framework- allowed cell to grow larger, move, & permitted metabolism
                              eucarya are 10x larger that bacteria
        5. evolve aerobic respiration...   more efficient energy transformation
        6. develop various organelles... (maybe by endosymbiosis*)...
                              a sub-cell part that catalyzes a specific metabolic function
        7. development of sexual cell cycles... (transposons - moveable genes)...
                      a method to shuffle genes along chromosomes favored cellular evolution
   


 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 


 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

Great 20th Century Discoveries ?

Apollo 12 Mission


   SKIP THIS MATERIAL
In Jan 2009 Tracey Lincoln & Gerald Joyce demonstrated a pair of RNA ribozymes each of which
could assemble RNAs by joining together two shorter RNA strands [ligases]. In 30 hours, they
found a group of RNA molecules that grow 100 million times bigger. Unfortunately, success in the
experiments required presence of preexisting RNA pieces that were far too long & complex to
have accumulated spontaneously. Still, the results suggest that RNA has the raw catalytic power
to catalyze its own replication...
                         thus synthetic RNA molecules may be able to evolve just as natural ones are...
  
2nd Experimental Camp to building a replicative molecular system in compartments...
        2. membranes:  argues key factor in evolution of 1st life was emergence of cell membrane,
                                  self-assembling and self-replicating vesicles. 
Zostak's lab investigated how fatty acids (lipids) might have trapped RNA producing... protocells
the mere presence of RNA in fatty acid vesicles could promote their growth by their
appropriating membrane molecules from neighboring vesicles with less or no RNA (2003:PDF & 2009- PDF)
    Did he make an early "protocell"* ?    Jack Szostak video on Abiogenesis (2009)may view@home -55min 
 
          RNA World Hypothesis updates*read @ home.          Origins of   ife Prize - Abiogenesis 



  SKIP THIS PAGE... Some experimental examples of synthetic biology accomplished with Viri:

  A.   Synthetic Polio Virus - July 12 ,2002 : Molecular Origin of Life Research or Bioterrorism?  
         Eckard Wimmer from the U. of New York at Stony Brook used the polio virus widely known genetic sequence to
         synthesize a virus from shelf chemicals. They followed a recipe they downloaded from the internet and used gene
         sequences from a mail-order supplier. The artificially constructed virus appears identical to its natural counterpart;
         when injected it into mice the animals were paralyzed and died.             (Science 297, 1016-1018, 2002)

  B.    Phi X-174 virus consists of a circular DNA molecule of 11 genes wrapped in an icosahedral coat of proteins
         that was artificially synthesized in Nov.2003 by Craig Venter and colleagues [Phi X-174]. They pieced together 
         synthetic DNA sequences ordered from a biotechnology company & used a technique called polymerase cycle
         to assemble the strands of DNA together. [1st sequenced in 1978 Phi X-174 has 5,386 bp-11 genes &  ]

  C.   The 1918 Spanish Flu Virus is Reconstructed - October 2005 : Jeffery K. Taubenberger, a molecular pathologist
         at the old Armed Forces Institute of Pathology and his colleagues were able to piece together the virus's genes from
         2 unusual sources: 1) lung tissue removed at autopsy of  a 21-year-old soldier & 2) the frozen body if an Inuit woman,
         who died of influenza in November 1918 & was buried in the Alaskan permafrost.
         These sources provided intact pieces of viral RNA* that could be analysed and sequenced.
             The virus has 8 "RNA gene segments" & by gene sequencing & PCR, they reassembled the virus.
             2 of the 8 genes:     Hemagglutinin-A   and   Neuraminidase  which are surface coat proteins
             There are at least 16 different HA antigens, which binds the virus to the host cell. Hemagglutinin-A is a
             surface glycoprotein that binds virus to host cell.  Neuraminidase is a surface antigenic glycoprotein enzyme.
             9 neuraminidase subtypes are known, which aid in the efficiency of virus release from infected cells.
                     H5-N1 make up a subtype of human influenza virus A and the avian influenza virus type-A
                     H1-N1 is the subtype of swine flu            
          Engineering a deadly virus  +  Dual Use Research  &  NSABB  &  antiviral Tamiflu
  
           Many Earth-like planets have been found by Kepler satellite*  +   Exoplanets .       

          More recent ideas and hypothesis*:  could aminoacyl-tRNA synthetases may be a link?   

 
   in 2014 M. Robertson & G. Joyce developed an RNA enzyme (ribozyme) that catalyzes
           RNA-templated polymerization of RNA utilizing NTP-substrates. The enzyme
           polymerized RNA sequences up to 95 nucleotides in length, though it could not
           replicate RNA or the RNA enzyme that catalyzed the polymerization.

 
*

   Now that we have seen the attributes of a cell  that is alive*    


            Moons & water: Life requires water which favors analysis "locally" of:    Mars*  &  Enceladus* 

     
        an entire synthetic Yeast genome              Codon replacement to rewrite genomes 


      Intelligent life is Probably Very Rare*


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