THE PRIMARY RESEARCH PRODUCT THIS PROJECT WILL PRODUCE IS IDENTIFICATION OF THE INSERTION STRAINS OF CHLAMYDOMONAS REINHARDTII A SINGLE-CELL GREEN ALGA WHICH OPTIMIZE SURVIVAL DURING EXPOSURE TO THE COMBINED IMPACTS OF SPACE RADIATION AND MICROGRAVITY IN THE COSMIC SPACE ENVIRONMENT BEYOND THE VAN ALLEN BELTS. THE CHLAMYDOMONAS REINHARDTII STRAINS IN A MAPPED INSERTIONAL MUTANT LIBRARY ARE A TRANSFORMATIVE TOOL ALLOWING NEAR GENOME WIDE ANALYSIS OF THE GENES MEDIATING SURVIVAL ADVANTAGE DURING STIMULI SUCH AS COSMIC RADIATION AND MICROGRAVITY. THE SECONDARY SET OF RESEARCH PRODUCTS THIS PROJECT WILL PRODUCE ARE IDENTIFICATION OF THE PATHWAYS MEDIATING THE GENE CHANGES VISUALIZATION OF THE PATHWAY INTERACTIONS AND IDENTIFICATION OF ANY ENVIRONMENTS GIVING SIMILAR GENE PATHWAY SIGNATURES IN OTHER MODEL SYSTEMS. OUR GOAL IS TO IDENTIFY STRAINS OF CHLAMYDOMONAS REINHARDTII WITH GENE INSERTIONS WHICH CONVEY THE BEST SURVIVAL ADVANTAGE DURING THE EXPOSURE TO THE COMBINED IMPACTS OF SPACE RADIATION AND MICROGRAVITY FOR FUTURE USE AS PARENT STRAINS IN HYDROGEN PRODUCTION INITIATIVES. HYDROGEN IS THE LEAD CONTENDER FOR FUEL PRODUCTION FOR RETURN SPACE FLIGHT MISSIONS FROM ORBITAL BODIES BEYOND LOW EARTH ORBIT. THE METABOLISM OF SELECT MICROORGANISMS MAKES HYDROGEN FUEL. THE BEST-KNOWN AND MOST EFFICIENT HYDROGEN PRODUCER IS CHLAMYDOMONAS REINHARDTII WHICH IS POISED FOR EXPLOITATION AS AN INDUSTRIAL BIOTECHNOLOGY PLATFORM FOR BIOFUEL PRODUCTION. THE TYPE OF INVESTIGATION WE PROPOSE IS EXPERIMENTS THAT WILL BE CONDUCTED ABOARD THE EM-1 LUNAR ORBIT FLIGHT WITH ASYNCHRONOUS CONTROLS IN KENNEDY SPACE CENTER ISSES CHAMBERS (ISS ENVIRONMENTAL SIMULATORS). THE PROPOSED HARDWARE IS THE NASA BRIC-100VC. HYPOTHESIS: THE HYPOTHESIS TO BE TESTED IS THAT THE MAPPED INSERTIONAL MUTANT LIBRARY OF CHLAMYDOMONAS REINHARDTII WILL ALLOW IDENTIFICATION OF THE GENES THAT MEDIATE OPTIMAL RADIATION RESISTANCE AND IMPERVIOUSNESS TO MICROGRAVITY IN THE COSMIC SPACE ENVIRONMENT BEYOND THE VAN ALLEN BELTS. THE SPECIFIC AIMS OF THE PROPOSAL ARE: SPECIFIC AIM 1: TO EXPERIMENTALLY DETERMINE WHICH OF THE THOUSANDS OF MAPPED INSERTION MUTANT STRAINS OF CHLAMYDOMONAS REINHARDTII HAVE THE BEST LONG-TERM SURVIVAL ADVANTAGE DURING COMBINED SPACE RADIATION AND MICROGRAVITY IMPACTS IN THE COSMIC SPACE ENVIRONMENT BEYOND THE VAN ALLEN BELTS. SPECIFIC AIM 2: TO ANALYZE THE DATASET OF INSERTION STRAINS WITH THE BEST LONG-TERM SURVIVAL ADVANTAGE DURING COMBINED SPACE RADIATION AND MICROGRAVITY IMPACTS TO: PERFORM PATHWAY ANALYSIS USING GENE SET ENRICHMENT ANALYSIS (GSEA) VISUALIZE THE PATHWAY INTERACTIONS USING CYTOSCAPE/CLUEGO MAKE COMPARISON TO HOMOLOGOUS GENE SETS IN CHLAMYDOMAS SACCHAROMYCES CEREVISIAE AND OTHER MODEL ORGANISM DATABASES FOR ANY ENVIRONMENTS GIVING SIMILAR GENE PATHWAY SIGNATURES. ALTHOUGH FOCUSED ON GENE PATHWAYS FOR SURVIVAL FOR FUTURE APPLICATIONS IN HYDROGEN FUEL PRODUCTION THE DATA GENERATED WILL PROVIDE GENOME LEVEL INSIGHTS INTO THE CANCER RISKS OF PROLONGED EXPOSURE TO COSMIC RADIATION AND MICROGRAVITY. THE SPECTRUM OF CANCER RISK GENES IS KNOWN AND THIS STUDY WILL PARSE THE IMPORTANCE OF RISK GENES AND THE PATTERNS OF RADIATION RISK CHANGE WITH EXPOSURE TO THE COSMIC SPACE ENVIRONMENT BEYOND THE VAN ALLEN BELTS.