REGULATORY SMALL RNAS (SRNAS) REPRESENT A MAJOR CLASS OF REGULATORY MOLECULES THAT PLAY LARGE-SCALE AND ESSENTIAL ROLES IN MANY CELLULAR PROCESSES ACROSS ALL DOMAINS OF LIFE. HOWEVER MICROBIAL SRNAS HAVE BEEN PRIMARILY STUDIED IN MODEL ORGANISMS AND VERY LITTLE IS KNOWN ABOUT THE DYNAMICS OF SRNA SYNTHESIS IN NATURAL ENVIRONMENTS AND THE ROLES OF THESE SHORT TRANSCRIPTS AT THE COMMUNITY LEVEL. THE GOAL OF THIS PROJECT IS TO INVESTIGATE THE ROLES OF REGULATORY SRNAS IN THE STRESS RESPONSE OF EXTREMOPHILE COMMUNITIES AND THEIR ABILITY TO COPE WITH ENVIRONMENTAL CHANGES.THE OVERARCHING RESEARCH QUESTIONS WE WILL ADDRESS ARE: DO MICROORGANISMS ACTIVATE SPECIFIC SRNAS WHEN IN A COMMUNITY? WHAT ARE THE UNDERLYING MECHANISMS FOR EXTREMOPHILE MICROBIAL COMMUNITIES TO ADAPT TO ENVIRONMENTAL CHANGES? AND WHAT ARE THE ROLES OF REGULATORY SRNAS IN THIS ADAPTIVE RESPONSE AT THE COMMUNITY-LEVEL? WE WILL USE A WELL-CHARACTERIZED MICROBIAL COMMUNITY INHABITING HALITE NODULES (SALT ROCK) FROM SALAR GRANDE IN THE ATACAMA DESERT CHILE AS OUR MODEL. THESE COMMUNITIES ARE POLYEXTREMOPHILIC AND AS SUCH THEIR STUDY IS APPLICABLE TO A BROAD RANGE OF TERRESTRIAL AND EXTRATERRESTRIAL ENVIRONMENTS. THIS PROJECT HAS 2 MAJOR OBJECTIVES: IN OBJECTIVE 1 WE WILL SEQUENCE THE GENOMES AND TRANSCRIPTOMES OF ISOLATES AND ENRICHMENT CULTURES FROM SALAR GRANDE HALITE AND CHARACTERIZE THEIR REGULATORY SRNAS USING OUR MICROBIAL SRNA IDENTIFICATION PIPELINE. THE RESULTING SRNA CATALOG WILL PROVIDE A RESOURCE FOR THE DISCOVERY OF COMMUNITY SRNAS IN HALITE METAGENOMES AND WILL BE COMPLEMENTARY TO THE DE NOVO APPROACHES PROPOSED IN OBJECTIVE 2. IN OBJECTIVE 2 WE WILL INVESTIGATE THE MOLECULAR AND ECOLOGICAL RELEVANCE OF MICROBIAL SRNAS IN THE STRESS RESPONSE OF THE HALITE COMMUNITY. TO DO SO WE PROPOSE TO TRANSPLANT HALITE NODULES FROM SALAR GRANDE TO SEVERAL TEST LOCATIONS IN THE DESERT THAT HAVE CONTRASTING ENVIRONMENTAL CONDITIONS. WE WILL COMBINE ENVIRONMENTAL ECOLOGICAL AND MOLECULAR ANALYSES TO IDENTIFY DIFFERENTIALLY REGULATED SRNAS BETWEEN THESE COMMUNITIES AFTER 1 YEAR. STRUCTURE AND TARGET PREDICTION OF THE MOST DIFFERENTIALLY EXPRESSED COMMUNITY SRNAS WILL PROVIDE THE FIRST INSIGHTS INTO THEIR ROLES AT THE COMMUNITY LEVEL. WE WILL BUILD THE SRNA IDENTIFICATION AND TARGET PREDICTION ANALYTICAL APPROACH DEVELOPED DURING THIS PROJECT INTO AN EASY-TO-USE BIOINFORMATICS PIPELINE FOR THE COMMUNITY. RELEVANCE TO NASA: THE IDENTIFICATION OF PROCESSES REGULATED BY SRNAS AT THE COMMUNITY LEVEL WILL FURTHER OUR UNDERSTANDING OF CO-EVOLUTIONARY MECHANISMS OF MICROBIAL COMMUNITIES AND EXPAND OUR KNOWLEDGE OF THE DIVERSITY AND GENOMIC DISTRIBUTIONS OF MICROBIAL SRNAS. FURTHERMORE UNDERSTANDING RNA S ECOLOGICAL RELEVANCE AT THE REGULATORY LEVEL AND HOW IT MIGHT ALLOW LIFE TO SURVIVE IN EXTREME ENVIRONMENTS CAN SHED LIGHT INTO THE EARLY EVOLUTION OF LIFE AND THE BIOSPHERE AND HELP RECONSTRUCT THE NATURE OF THE MOST PRIMITIVE ORGANISMS. THIS PROJECT FOCUSES ON EXTREMOPHILES WHICH ARE MODEL ORGANISMS OF POSSIBLE ECOSYSTEMS BEYOND EARTH. THEREFORE UNDERSTANDING THE PHYSIOLOGICAL AND MOLECULAR ADAPTATIONS OF EXTREMOPHILES TO ENVIRONMENTAL STRESS AND CLIMATE CHANGE CAN ALSO IMPROVE PREDICTIONS OF WHERE LIFE MIGHT EXIST ELSEWHERE IN THE SOLAR SYSTEM AND WHERE TO SEARCH FOR EVIDENCE OF IT.