THE OVERARCHING QUESTION THAT THIS PROJECT ADDRESSES IS: HOW CAN EVOLUTIONARY PROCESSES ALLOW LIFE TO EMERGE AND THEN EXPAND TO DIFFERENT ENVIRONMENTS? EVOLUTION THAT DRIVES INANIMATE MATTER TO BECOME ANIMATE HAS TO BE OPEN-ENDED I.E. CONTINUALLY PRODUCE NOVEL AND ADAPTIVE FORMS. ALSO ANY FORM OF LIFE THAT CANNOT LIBERATE ITSELF FROM THE INITIAL PREBIOTIC NICHE IS UNLIKELY TO SURVIVE. THUS IN ORDER FOR LIFE TO BECOME A PLANETARY PHENOMENON IT NEEDS TO EVOLVE A MECHANISM TO CONTROL ITS FUNCTIONS IN DIFFERENT ENVIRONMENTS WHILE PRESERVING THE MACHINERY OF OPEN-ENDED EVOLUTION. HOW THIS HAPPENS REMAINS LARGELY UNKNOWN. RECENT ADVANCES IN IN VITRO EVOLUTION TECHNIQUES HIGH THROUGHPUT SEQUENCING AND COMPUTATIONAL METHODS OF BIOINFORMATICS AND MOLECULAR MODELING WILL ALLOW US TO EXPLORE THIS ISSUE THROUGH SYSTEMATIC INTEGRATED SCIENTIFIC INQUIRY. WE INTEND TO UNDERTAKE THIS EFFORT COMBINING FOR THE FIRST TIME THESE THREE TECHNIQUES INTO A SINGLE APPROACH. EVOLUTIONARY PROCESSES IN EMERGING AND ESTABLISHED BIOLOGICAL SYSTEMS ARE DIFFERENT EVEN THOUGH THEY ARE BOTH BASED ON THE SAME PRINCIPLES OF PHYSICS CHEMISTRY AND NATURAL SELECTION. EARLY EVOLUTION IS CHARACTERIZED BY A RAPID RATE OF INNOVATION TIGHT COEVOLUTION OF DIFFERENT CELLULAR COMPONENTS AND A CLOSE RELATIONSHIP BETWEEN EVOLUTION OF FUNCTIONAL MOLECULES AND PHENOTYPE. THE PROLIFERATION OF LIFE DEPENDS ON A PROCESS OF OPEN-ENDED EVOLUTION IN WHICH THE EMERGING FUNCTIONS ARE CARRIED OUT BY BIOPOLYMERS (RNA PROTEINS). UNDERSTANDING HOW EARLY BIOPOLYMERS ACQUIRED AND IMPROVED FUNCTIONS INVOLVES ANSWERING FUNDAMENTAL QUESTIONS SUCH AS: WHEN IS EVOLUTIONARY OPTIMIZATION POSSIBLE AND WHEN ARE THE PRODUCTS OF EVOLUTION GOVERNED BY CHANCE? (Q1) WHAT IS THE ROLE OF NEUTRAL OR NEAR NEUTRAL SINGLE-POINT MUTATIONS IN DRIVING EVOLUTION OF FUNCTIONAL MOLECULES? (Q2) HOW DOES INCREASING POLYMER LENGTH AN EVOLUTIONARY MECHANISM ALTERNATIVE TO POINT MUTATIONS LEAD TO INNOVATION ON BOTH STRUCTURAL AND FUNCTIONAL LEVELS? (Q3) EVOLUTION OF FUNCTIONAL MOLECULES SHOULD NOT BE CONSIDERED IN ISOLATION. WHAT EVOLVES IS A SYSTEM WHICH IN EARLY EVOLUTION WE IDENTIFY WITH A PROTOCELL. IN THIS EVOLVING ENTITY ALL COMPONENTS WORK IN CONCERT. THIS FEATURE WHICH IS NOT PRESENT IN EVOLUTIONARY STUDIES ON FUNCTIONAL MOLECULES LEADS TO A DIFFERENT EVOLUTIONARY CRITERION. FIT PROTOCELLS ARE THOSE THAT GROW AND DIVIDE LEAVING MORE PROGENY CAPABLE OF REPRODUCTION THAN THEIR COMPETITION. THIS CREATES AN IMPERATIVE FOR COEVOLUTION OF DIFFERENT FUNCTIONS THAT INVOLVES NOT ONLY FUNCTIONAL POLYMERS BUT ALSO A NUMBER OF PHYSICAL AND CHEMICAL PROCESSES. THESE PROCESSES INCLUDE THE FORMATION AND MAINTENANCE OF SEMIPERMEABLE CELL BOUNDARIES ASSOCIATION OF MOLECULES TO SUPRAMOLECULAR STRUCTURES AND SELFORGANIZATION OF SUBCELLULAR COMPONENTS. THE REQUIREMENT OF INTEGRATION AND COORDINATION RAISES SEVERAL QUESTIONS CENTRAL TO UNDERSTANDING EVOLUTION OF PROTOCELLULAR SYSTEMS: WHAT MOLECULAR MECHANISMS OF COEVOLUTION OF PROTEINS NUCLEIC ACIDS AND THE CELLULAR MEMBRANES FACILITATE PROTOCELLULAR FUNCTIONS STABILITY GROWTH AND DIVISION? (Q4) WHICH FUNCTIONS FACILITATE ADAPTATION OF NASCENT LIFE TO DIFFERENT ENVIRONMENTAL CONDITIONS AND HOW DO THEY COEVOLVE? (Q5) UNDER WHAT CONDITIONS IS OPEN-ENDED EVOLUTION POSSIBLE AND WHAT CONSTRAINTS DOES IT IMPOSE ON THE DIFFERENT PROCESSES INVOLVED? (Q6)