Project Grant R00GM149822

STUDYING THE MODULATORS AND THE PHYSIOLOGICAL FUNCTIONS OF RNA TAILING IN THE C. ELEGANS OOCYTE - ABSTRACT THE DECREASING QUALITY OF OOCYTES, ASSOCIATED WITH MATERNAL AGE, IS A MAJOR COMPONENT OF REDUCED FERTILITY IN OLDER WOMEN. MANY MOLECULAR MARKERS ARE OBSERVED IN DETERIORATING OOCYTES, INCLUDING TRANSCRIPTOMIC CHANGES. INTERESTINGLY, IN MOST ANIMAL MODELS, THE END OF OOGENESIS AND EARLY EMBRYOGENESIS PROCEED IN THE ABSENCE OF TRANSCRIPTION, RELYING ON STORED MESSENGER RNAS (MRNAS). THE DEVELOPING OOCYTE CAREFULLY MANAGES DIFFERENT POPULATIONS OF RNA DURING MATURATION, FERTILIZATION, AND ZYGOTIC GENOME ACTIVATION TO SUCCESSFULLY JUMPSTART EMBRYOGENESIS. THE ACCUMULATION OF THESE MATERNAL TRANSCRIPTS DURING OOGENESIS IS THEREFORE ESSENTIAL. YET WE KNOW VERY LITTLE OF HOW THE TRANSCRIPTOME IS SCULPTED AFTER THE TRANSCRIPTIONAL MACHINERY IS INACTIVATED. WHAT DRIVES TRANSCRIPTOMIC CHANGES AND HOW ARE THEY CONTRIBUTING TO OOCYTE QUALITY? RNA TAILING, OR THE ADDITION OF UNTEMPLATED NUCLEOTIDES TO THE 3' END OF RNA MOLECULES, IS A POST- TRANSCRIPTIONAL PROCESS THAT HAS LONG BEEN ASSOCIATED WITH THE REGULATION OF RNA STABILITY AND TRANSLATION. BOTH NUCLEOTIDE COMPOSITION AND LENGTH OF TAILS CAN DETERMINE TAIL FUNCTION. RNA TAILS ARE ESPECIALLY DYNAMIC IN THE GERMLINE AND THE EARLY EMBRYO. THE OBJECTIVE OF THIS WORK IS TO IDENTIFY THE TERMINAL NUCLEOTIDYL TRANSFERASES AND EXONUCLEASE THAT MODULATE RNA TAILS AND TO ELUCIDATE THE MECHANISM THAT MEDIATE THE DOWNSTREAM EFFECTS ON RNA STABILITY AND TRANSLATION IN THE OOCYTE. FIRST, I WILL EXAMINE THE PHYSIOLOGICAL FUNCTION OF TNTS AND EXONUCLEASES IN C. ELEGANS FERTILITY. SECOND, I WILL CHARACTERIZE CHANGES IN RNA TAIL LENGTH AND COMPOSITION DURING OOGENESIS AND EARLY EMBRYOGENESIS TO SHED LIGHT ON CONSERVED PATHWAYS INVOLVED IN GENERATING VIABLE AND COMPETENT OOCYTES. THIRD, I WILL IDENTIFY THE CO-FACTORS THAT ACT UPSTREAM AND DOWNSTREAM OF TNTS AND EXONUCLEASES, TO COORDINATE THEIR ACTIVITY, AND MODULATE TAIL-MEDIATED REGULATION OF RNA STABILITY AND TRANSLATION. I AM UNIQUELY QUALIFIED TO CONDUCT THIS RESEARCH, HAVING STUDIED DIFFERENT TYPES OF RNA AND THEIR BIOLOGY THROUGHOUT MY GRADUATE AND POSTDOCTORAL TRAINING. IN KATHERINE MCJUNKIN'S LABORATORY, I HAVE USED THE C. ELEGANS MODEL SYSTEM AND LARGE-SCALE GENOMIC SCREENS TO IDENTIFY TNTS RESPONSIBLE FOR MIRNA TAILING AND TO ASSESS ITS IMPACT ON MICRORNA TURNOVER. IN THE PROPOSED WORK, I WILL APPLY STATE-OF-THE-ART TECHNIQUES TO DISSECT THE MACHINERY RESPONSIBLE FOR MRNA TAILING IN THE CONTEXT OF REPRODUCTION AND FERTILITY. THIS WORK SEEKS TO EXPLORE POST-TRANSCRIPTIONAL MECHANISMS REGULATING GENE EXPRESSION DURING THE OOCYTE-TO-EMBRYO TRANSITION AND THEIR CONTRIBUTION TO OOCYTE QUALITY.