Project Grant F30HL184499

REGULATION OF TRANSPOSABLE ELEMENTS IN AGING HEMATOPOIETIC STEM AND PROGENITOR CELLS VIA SINGLE-CELL MULTI-OMICS - PROJECT SUMMARY CLONAL HEMATOPOIESIS (CH), THE CLONAL EXPANSION OF MUTANT HEMATOPOIETIC STEM AND PROGENITOR CELLS (HSPCS), HAS BEEN CLOSELY LINKED TO AGING AND IMPLICATED IN HEMATOLOGICAL MALIGNANCIES AND CARDIOVASCULAR DISEASES. GENETIC AND EPIGENETIC DYSFUNCTION ARE WELL-KNOWN IN CH, YET THE CONTRIBUTIONS OF HETEROCHROMATIN DYSREGULATION AND TRANSPOSABLE ELEMENTS (TES) REMAIN UNDER EXPLORED. AMONG THE MOST FREQUENT GENETIC ALTERATIONS IN CH ARE MUTATIONS IN TET2, A DNA METHYLATION REGULATOR, HIGHLIGHTING THE IMPORTANCE OF EPIGENETIC INTEGRITY IN HSPCS. RECENT EVIDENCE INDICATES THAT AGE-RELATED HETEROCHROMATIN DYSREGULATION DRIVES TE ACTIVATION IN HSPCS WHERE TES CAN MIMIC VIRAL ELEMENTS, INDUCE INFLAMMATORY SIGNALING, AND DEGRADE NORMAL HSPC FUNCTION, ENABLING MUTANT HSPCS TO OUTCOMPETE THEIR NORMAL COUNTERPARTS. THIS PROPOSAL SEEKS TO DELINEATE THE PRECISE ROLES OF TES IN CH PATHOGENESIS. WE HYPOTHESIZE THAT THE EPIGENETIC DYSREGULATION ACCOMPANYING AGING NOT ONLY TRIGGERS ABNORMAL TE EXPRESSION ACROSS DISTINCT HSPC SUBSETS BUT ALSO CONTRIBUTES TO THEIR FUNCTIONAL DECLINE, THEREBY FOSTERING CONDITIONS FAVORABLE FOR TET2 MUTANT CELL EXPANSION. TO TEST THIS HYPOTHESIS, SINGLE-CELL MULTI-OMICS APPROACHES, AND IN-VIVO MOUSE MODELS WILL BE USED. IN AIM 1, THE EPIGENETIC AND TRANSCRIPTIONAL LANDSCAPES OF TES IN HSPCS WILL BE MAPPED AT SINGLE-CELL RESOLUTION DURING NORMAL AGING. SINGLE-CELL DATASETS FROM YOUNG AND AGED WILD-TYPE (WT) MICE WILL BE EMPLOYED TO REVEAL CELL-TYPE AND AGE-SPECIFIC TE EXPRESSION PATTERNS. EXPERIMENTAL VALIDATION WILL BE USED TO DETERMINE HOW TE DYSREGULATION IMPACTS HSPC SELF-RENEWAL AND LINEAGE SPECIFICATION. IN AIM 2, HOW TET2 LOSS OF FUNCTION (LOF) RESHAPES TE EXPRESSION DYNAMICS AND ACCELERATES CH ONSET WILL BE ASSESSED. BY INTEGRATING SINGLE-CELL OMICS FROM YOUNG AND OLD TET2 KNOCKOUT MICE AND EXPERIMENTAL VALIDATION, THE GENOMIC LOCI SUSCEPTIBLE TO TET2 DYSREGULATION AND TE UPREGULATION WILL BE PINPOINTED. THIS WILL ILLUMINATE HOW TET2 LOF AT DIFFERENT AGES INFLUENCES CH PATHOGENESIS VIA ALTERED TE EXPRESSION AND EPIGENETIC INSTABILITY. THIS PROJECT'S INNOVATION LIES IN ITS COMPREHENSIVE, SINGLE-CELL-BASED INVESTIGATION OF TE REGULATION DURING HSPC AGING AND ITS DIRECT INTERROGATION OF TET2 LOF THROUGH A CHEMOGENETIC PLATFORM. BY INTEGRATING MOLECULAR BIOLOGY WITH COMPUTATIONAL METHODS, OUR WORK WILL OFFER UNPRECEDENTED INSIGHT INTO HOW EPIGENETIC ABERRATIONS AND TE DYSREGULATION DRIVE CH, INFORMING NOVEL THERAPEUTICS TO PRESERVE THE INTEGRITY OF THE AGING HSPCS AND MITIGATE THE PROGRESSION OF CH. ADDITIONALLY, THIS F30 FELLOWSHIP APPLICATION INCORPORATES A PLAN FOR LOGAN'S MD/PHD TRAINING AT TEXAS A & M IN THE LAB OF DR. HUANG, A LEADING EXPERT ON EPIGENETICS AND CH. COMBINING DUAL TRAINING IN WET AND DRY LAB TECHNIQUES WILL ALLOW LOGAN TO DEVELOP A ROBUST SKILLSET TO INTEGRATE FUNDAMENTAL RESEARCH IN CLINICAL APPLICATION. HIS MULTIDISCIPLINARY MENTORING ENVIRONMENT WILL ENSURE HIS DEVELOPMENT AS AN INDEPENDENT PHYSICIAN-SCIENTIST.