Not listed CHARACTERIZING FATTY ACID UPTAKE, METABOLISM, AND STORAGE BY GUT BACTERIA FROM THE GENUS TURICIBACTER - PROJECT SUMMARY/ABSTRACT THE GASTROINTESTINAL MICROBIOTA IS A POTENT MEDIATOR OF HOST LIPID INTERACTIONS AND METABOLISM. GRAM POSITIVE BACTERIA FROM THE UNDERSTUDIED TURICIBACTER GENUS LIVE IN THE GUTS OF MAMMALS, BIRDS, AND FISH AND ARE PREDICTED TO INTERACT WITH LIPIDS IN HUMAN AND ANIMAL HOSTS. MONOCOLONIZATION OF MICE WITH DIFFERENT STRAINS OF TURICIBACTER ALTER HOST SERUM LIPIDS WITH A NOTABLE DECREASE IN SERUM TRIGLYCERIDES, PARTICULARLY THOSE CONTAINING SATURATED AND MONOUNSATURATED FATTY ACIDS (FAS). TO UNDERSTAND HOW TURICIBACTERS DIRECTLY INTERACT WITH FAS, I PERFORMED RNASEQ ON TURICIBACTER CULTURES PREINCUBATED WITH FAS AND FOUND THAT SATURATED, MONOUNSATURATED, AND POLYUNSATURATED FAS ELICIT DISTINCT TRANSCRIPTIONAL RESPONSES IN THESE CELLS. THESE RESULTS LED ME TO IDENTIFY PUTATIVE TURICIBACTER GENE HOMOLOGS OF A RECENTLY DISCOVERED GRAM POSITIVE FA UPTAKE SYSTEM CALLED THE FATTY ACID KINASE (FAK) SYSTEM. MY RNASEQ DATA INDICATED THAT DIFFERENT FA SUBSTRATES PROMPTED A DIFFERENT REGULATION PATTERN FOR THE FOUR FA-BINDING (FAKB) HOMOLOGS POSSESSED BY TURICIBACTERS, AND LATER AMINO ACID SEQUENCE ANALYSIS REVEALED THAT THREE OF THESE HOMOLOGS ARE DISTINCT FROM ALL PREVIOUSLY STUDIED FAK PROTEINS. AIM 1 OF MY PROPOSAL IS TO DEFINE THE MECHANISM OF FA UPTAKE BY TURICIBACTERS AND CHARACTERIZE FA BINDING AND SPECIFICITY OF NOVEL FAKB PROTEINS. TO DO THIS, I WILL PURIFY FAK KINASE AND FAKB HOMOLOGS IDENTIFIED IN TURICIBACTERS, THEN PROBE THEIR ABILITY TO PHOSPHORYLATE FAS (A CRUCIAL FIRST STEP DURING METABOLISM) AND CONDUCT FA BINDING AFFINITY AND COMPETITION ASSAYS TO DETERMINE THEIR BINDING SPECIFICITY. THIS CHARACTERIZATION OF THE TURICIBACTER FAK SYSTEM WILL IMPROVE OUR UNDERSTANDING OF FA UPTAKE ACROSS GRAM POSITIVE BACTERIA. AIM 2 OF MY PROPOSAL WILL DEFINE THE CONSEQUENCES OF TURICIBACTER FA UPTAKE BY STUDYING LIPIDOMIC AND METABOLOMIC CHANGES IN TURICIBACTER CELLS AND SUBCELLULAR LIPID DROPLET (LD)-LIKE STRUCTURES DURING GROWTH WITH FAS. INITIAL EXPERIMENTS USING FLUORESCENT LABELED FAS INDICATE THAT TURICIBACTERS INCORPORATE EXOGENOUS FAS INTO POLAR AND NEUTRAL LIPIDS, AND IMAGING ANALYSIS REVEALED THE PRESENCE OF LIPID-LABELED STRUCTURES WITHIN TURICIBACTER CELLS THAT ARE NOT WELL STUDIED IN GUT BACTERIA. I WILL FIRST STUDY CHANGES IN WHOLE CELLS WHEN TURICIBACTERS ARE INCUBATED WITH DIFFERENT FAS, THEN ISOLATE LD-LIKE STRUCTURES AND IDENTIFY THEIR SPECIFIC LIPID AND PROTEIN COMPONENTS. FINALLY, I WILL USE FLUORESCENT LABELING AND IMMUNOFLUORESCENCE TO STUDY THE CO-OCCURRENCE OF FAK PROTEINS WITH LIPID ACCUMULATION AND CELL GROWTH. THESE EXPERIMENTS WILL ALLOW US TO IDENTIFY SPECIFIC PRODUCTS OF FAS TAKEN UP BY TURICIBACTERS AND PROVIDE CLUES TO THE FUNCTION OF LIPID STORAGE STRUCTURES IN ENVIRONMENTS WHERE NUTRIENTS ARE ABUNDANT. TOGETHER, THESE AIMS WILL CHARACTERIZE HOW TURICIBACTERS TAKE UP, METABOLIZE, AND STORE EXOGENOUS FAS. THE RESULTS OF THESE STUDIES WILL LAY A FOUNDATION FOR FUTURE RESEARCH INTO HOW FA METABOLISM BY TURICIBACTERS AND RELATED BACTERIA INFLUENCES HOST LIPID METABOLISM AND INFORM PRODUCTION OF MICROBIOTA-BASED THERAPEUTICS FOR METABOLIC DISORDERS. $50.1k 5/7/26 Not listed CHARACTERIZING FATTY ACID UPTAKE, METABOLISM, AND STORAGE BY GUT BACTERIA FROM THE GENUS TURICIBACTER - PROJECT SUMMARY/ABSTRACT THE GASTROINTESTINAL MICROBIOTA IS A POTENT MEDIATOR OF HOST LIPID INTERACTIONS AND METABOLISM. GRAM POSITIVE BACTERIA FROM THE UNDERSTUDIED TURICIBACTER GENUS LIVE IN THE GUTS OF MAMMALS, BIRDS, AND FISH AND ARE PREDICTED TO INTERACT WITH LIPIDS IN HUMAN AND ANIMAL HOSTS. MONOCOLONIZATION OF MICE WITH DIFFERENT STRAINS OF TURICIBACTER ALTER HOST SERUM LIPIDS WITH A NOTABLE DECREASE IN SERUM TRIGLYCERIDES, PARTICULARLY THOSE CONTAINING SATURATED AND MONOUNSATURATED FATTY ACIDS (FAS). TO UNDERSTAND HOW TURICIBACTERS DIRECTLY INTERACT WITH FAS, I PERFORMED RNASEQ ON TURICIBACTER CULTURES PREINCUBATED WITH FAS AND FOUND THAT SATURATED, MONOUNSATURATED, AND POLYUNSATURATED FAS ELICIT DISTINCT TRANSCRIPTIONAL RESPONSES IN THESE CELLS. THESE RESULTS LED ME TO IDENTIFY PUTATIVE TURICIBACTER GENE HOMOLOGS OF A RECENTLY DISCOVERED GRAM POSITIVE FA UPTAKE SYSTEM CALLED THE FATTY ACID KINASE (FAK) SYSTEM. MY RNASEQ DATA INDICATED THAT DIFFERENT FA SUBSTRATES PROMPTED A DIFFERENT REGULATION PATTERN FOR THE FOUR FA-BINDING (FAKB) HOMOLOGS POSSESSED BY TURICIBACTERS, AND LATER AMINO ACID SEQUENCE ANALYSIS REVEALED THAT THREE OF THESE HOMOLOGS ARE DISTINCT FROM ALL PREVIOUSLY STUDIED FAK PROTEINS. AIM 1 OF MY PROPOSAL IS TO DEFINE THE MECHANISM OF FA UPTAKE BY TURICIBACTERS AND CHARACTERIZE FA BINDING AND SPECIFICITY OF NOVEL FAKB PROTEINS. TO DO THIS, I WILL PURIFY FAK KINASE AND FAKB HOMOLOGS IDENTIFIED IN TURICIBACTERS, THEN PROBE THEIR ABILITY TO PHOSPHORYLATE FAS (A CRUCIAL FIRST STEP DURING METABOLISM) AND CONDUCT FA BINDING AFFINITY AND COMPETITION ASSAYS TO DETERMINE THEIR BINDING SPECIFICITY. THIS CHARACTERIZATION OF THE TURICIBACTER FAK SYSTEM WILL IMPROVE OUR UNDERSTANDING OF FA UPTAKE ACROSS GRAM POSITIVE BACTERIA. AIM 2 OF MY PROPOSAL WILL DEFINE THE CONSEQUENCES OF TURICIBACTER FA UPTAKE BY STUDYING LIPIDOMIC AND METABOLOMIC CHANGES IN TURICIBACTER CELLS AND SUBCELLULAR LIPID DROPLET (LD)-LIKE STRUCTURES DURING GROWTH WITH FAS. INITIAL EXPERIMENTS USING FLUORESCENT LABELED FAS INDICATE THAT TURICIBACTERS INCORPORATE EXOGENOUS FAS INTO POLAR AND NEUTRAL LIPIDS, AND IMAGING ANALYSIS REVEALED THE PRESENCE OF LIPID-LABELED STRUCTURES WITHIN TURICIBACTER CELLS THAT ARE NOT WELL STUDIED IN GUT BACTERIA. I WILL FIRST STUDY CHANGES IN WHOLE CELLS WHEN TURICIBACTERS ARE INCUBATED WITH DIFFERENT FAS, THEN ISOLATE LD-LIKE STRUCTURES AND IDENTIFY THEIR SPECIFIC LIPID AND PROTEIN COMPONENTS. FINALLY, I WILL USE FLUORESCENT LABELING AND IMMUNOFLUORESCENCE TO STUDY THE CO-OCCURRENCE OF FAK PROTEINS WITH LIPID ACCUMULATION AND CELL GROWTH. THESE EXPERIMENTS WILL ALLOW US TO IDENTIFY SPECIFIC PRODUCTS OF FAS TAKEN UP BY TURICIBACTERS AND PROVIDE CLUES TO THE FUNCTION OF LIPID STORAGE STRUCTURES IN ENVIRONMENTS WHERE NUTRIENTS ARE ABUNDANT. TOGETHER, THESE AIMS WILL CHARACTERIZE HOW TURICIBACTERS TAKE UP, METABOLIZE, AND STORE EXOGENOUS FAS. THE RESULTS OF THESE STUDIES WILL LAY A FOUNDATION FOR FUTURE RESEARCH INTO HOW FA METABOLISM BY TURICIBACTERS AND RELATED BACTERIA INFLUENCES HOST LIPID METABOLISM AND INFORM PRODUCTION OF MICROBIOTA-BASED THERAPEUTICS FOR METABOLIC DISORDERS. $50.1k 5/7/26 Not listed CHARACTERIZING FATTY ACID UPTAKE, METABOLISM, AND STORAGE BY GUT BACTERIA FROM THE GENUS TURICIBACTER - PROJECT SUMMARY/ABSTRACT THE GASTROINTESTINAL MICROBIOTA IS A POTENT MEDIATOR OF HOST LIPID INTERACTIONS AND METABOLISM. GRAM POSITIVE BACTERIA FROM THE UNDERSTUDIED TURICIBACTER GENUS LIVE IN THE GUTS OF MAMMALS, BIRDS, AND FISH AND ARE PREDICTED TO INTERACT WITH LIPIDS IN HUMAN AND ANIMAL HOSTS. MONOCOLONIZATION OF MICE WITH DIFFERENT STRAINS OF TURICIBACTER ALTER HOST SERUM LIPIDS WITH A NOTABLE DECREASE IN SERUM TRIGLYCERIDES, PARTICULARLY THOSE CONTAINING SATURATED AND MONOUNSATURATED FATTY ACIDS (FAS). TO UNDERSTAND HOW TURICIBACTERS DIRECTLY INTERACT WITH FAS, I PERFORMED RNASEQ ON TURICIBACTER CULTURES PREINCUBATED WITH FAS AND FOUND THAT SATURATED, MONOUNSATURATED, AND POLYUNSATURATED FAS ELICIT DISTINCT TRANSCRIPTIONAL RESPONSES IN THESE CELLS. THESE RESULTS LED ME TO IDENTIFY PUTATIVE TURICIBACTER GENE HOMOLOGS OF A RECENTLY DISCOVERED GRAM POSITIVE FA UPTAKE SYSTEM CALLED THE FATTY ACID KINASE (FAK) SYSTEM. MY RNASEQ DATA INDICATED THAT DIFFERENT FA SUBSTRATES PROMPTED A DIFFERENT REGULATION PATTERN FOR THE FOUR FA-BINDING (FAKB) HOMOLOGS POSSESSED BY TURICIBACTERS, AND LATER AMINO ACID SEQUENCE ANALYSIS REVEALED THAT THREE OF THESE HOMOLOGS ARE DISTINCT FROM ALL PREVIOUSLY STUDIED FAK PROTEINS. AIM 1 OF MY PROPOSAL IS TO DEFINE THE MECHANISM OF FA UPTAKE BY TURICIBACTERS AND CHARACTERIZE FA BINDING AND SPECIFICITY OF NOVEL FAKB PROTEINS. TO DO THIS, I WILL PURIFY FAK KINASE AND FAKB HOMOLOGS IDENTIFIED IN TURICIBACTERS, THEN PROBE THEIR ABILITY TO PHOSPHORYLATE FAS (A CRUCIAL FIRST STEP DURING METABOLISM) AND CONDUCT FA BINDING AFFINITY AND COMPETITION ASSAYS TO DETERMINE THEIR BINDING SPECIFICITY. THIS CHARACTERIZATION OF THE TURICIBACTER FAK SYSTEM WILL IMPROVE OUR UNDERSTANDING OF FA UPTAKE ACROSS GRAM POSITIVE BACTERIA. AIM 2 OF MY PROPOSAL WILL DEFINE THE CONSEQUENCES OF TURICIBACTER FA UPTAKE BY STUDYING LIPIDOMIC AND METABOLOMIC CHANGES IN TURICIBACTER CELLS AND SUBCELLULAR LIPID DROPLET (LD)-LIKE STRUCTURES DURING GROWTH WITH FAS. INITIAL EXPERIMENTS USING FLUORESCENT LABELED FAS INDICATE THAT TURICIBACTERS INCORPORATE EXOGENOUS FAS INTO POLAR AND NEUTRAL LIPIDS, AND IMAGING ANALYSIS REVEALED THE PRESENCE OF LIPID-LABELED STRUCTURES WITHIN TURICIBACTER CELLS THAT ARE NOT WELL STUDIED IN GUT BACTERIA. I WILL FIRST STUDY CHANGES IN WHOLE CELLS WHEN TURICIBACTERS ARE INCUBATED WITH DIFFERENT FAS, THEN ISOLATE LD-LIKE STRUCTURES AND IDENTIFY THEIR SPECIFIC LIPID AND PROTEIN COMPONENTS. FINALLY, I WILL USE FLUORESCENT LABELING AND IMMUNOFLUORESCENCE TO STUDY THE CO-OCCURRENCE OF FAK PROTEINS WITH LIPID ACCUMULATION AND CELL GROWTH. THESE EXPERIMENTS WILL ALLOW US TO IDENTIFY SPECIFIC PRODUCTS OF FAS TAKEN UP BY TURICIBACTERS AND PROVIDE CLUES TO THE FUNCTION OF LIPID STORAGE STRUCTURES IN ENVIRONMENTS WHERE NUTRIENTS ARE ABUNDANT. TOGETHER, THESE AIMS WILL CHARACTERIZE HOW TURICIBACTERS TAKE UP, METABOLIZE, AND STORE EXOGENOUS FAS. THE RESULTS OF THESE STUDIES WILL LAY A FOUNDATION FOR FUTURE RESEARCH INTO HOW FA METABOLISM BY TURICIBACTERS AND RELATED BACTERIA INFLUENCES HOST LIPID METABOLISM AND INFORM PRODUCTION OF MICROBIOTA-BASED THERAPEUTICS FOR METABOLIC DISORDERS. $0 5/7/26