** AWARDS ISSUED PRIOR TO JANUARY 20, 2025, WERE FUNDED UNDER PREVIOUS ADMINISTRATIONS AND MAY NOT REFLECT THE PRIORITIES AND POLICIES OF THE CURRENT ADMINISTRATION.** CORNIS THE MOST SIGNIFICANT CEREAL CROP GROWN IN THE US FOR FOOD, FEED, AND BIOFUEL PRODUCTION. DUE TO ITS TROPICALORIGINS CORN IS SUSCEPTIBLE TO COLD STRESS ESPECIALLY DURING GERMINATION AND AS A SEEDLING WHEN IT'S PLANTING TIME LEAVES IT VULNERABLE TO SPRING TEMPERATURE DROPS. PLANTING CORN LATER IN THE SEASON IS NOT A VIABLE OPTION, AS 1) TEMPERATURE SPIKES LATER IN THE GROWING SEASON DURING REPRODUCTIVE GROWTH NEGATIVELY AND SEVERELY IMPACT CROP YIELD AND 2) CLIMATE CHANGE IS CAUSING MORE EXTREME AND FREQUENT TEMPERATURE FLUCTUATIONS TO OCCUR THROUGHOUT THE GROWING SEASON. THUS,IMPROVING CORN TEMPERATURE TOLERANCES IS A REQUIREMENT FOR MAXIMIZING CROP YIELD THROUGH FUTURE BREEDING OR GENETIC ENGINEERING PROGRAMS. WHILE COLD RESPONSE AND TOLERANCE IS WELL STUDIED ESPECIALLY AT THE TRANSCRIPTIONAL LEVEL WHICH OCCURS IN PART THROUGH CALCIUM AND REACTIVE OXYGEN SPECIES (ROS) SIGNALING, TRANSLATIONAL REGULATORY MECHANISMS WHICH AFFECT ONLY PROTEIN PRODUCTION HAVE NOT BEEN WELL EXPLORED ESPECIALLY IN COLD-STRESSED CORN. PRELIMINARY ANALYSES IDENTIFIED CYTOSOL AND MITOCHONDRIAL CALCIUM TRANSPORTATION GENES AS BEING BOTH TRANSCRIPTIONALLY AND TRANSLATIONALLY UPREGULATED BY COLD STRESS. WE PROPOSE TO EXPLORE CORN COLD TOLERANCE THROUGH THE LENS OF TRANSLATION REGULATION TO REVEAL REGULATORY PROCESSES THAT CAN BE TARGETED AND IMPROVED THROUGH BREEDING OR GENETIC ENGINEERING TO GENERATE COLD-RESISTANT CORN VARIETIES, AND TO EXAMINE THE FUNCTION AND IMPACT OF GENERAL AND MITOCHONDRIAL CALCIUM TRANSPORTATION GENES ON PLANT COLD TOLERANCE.TO CONFIRM THAT CALCIUM TRANSPORTATION GENES PLAY A ROLE IN COLD TOLERANCE, CANDIDATE GENE EXPRESSION WILL BE REDUCED OR ELIMINATED IN CORN AND THE MODEL PLANT ARABIDOPSIS, AND THESE PLATS WILL SUBSEQUENTLY BE GROWN UNDER CHILLING AND FREEZING STRESS TO DETERMINE IF LOSS OF THESE GENES REDUCES COLD TOLERANCE. AS CALCIUM AND ROS SIGNALING IN THE CYTOSOL ARE KNOWN TO INDUCE THE EXPRESSION OF COLD RESPONSE GENES, WE WILL MEASURE MITOCHONDRIAL AND CYTOSOLIC CALCIUM AND ROS LEVELS IN WILD-TYPE AND MUTANT PLANTS TO1) SEE IF THESE MUTATIONS INTERFERE WITH KNOWN COLD SIGNALING PATHWAYS AND 2) DETERMINE IF DISRUPTION OF ONLY MITOCHONDRIAL CALCIUM AND ROS BALANCE IS SUFFICIENT TO INHIBIT COLD TOLERANCE. TO IDENTIFY TRANSLATIONAL REGULATION MACHINERY, THE GENETIC SEQUENCES OF TRANSLATIONALLY-REGULATED GENES WILL BE ANALYZED TO IDENTIFY SHARED SEQUENCES OR STRUCTURES THAT MAY IMPACT TRANSLATIONAL REGULATION, WHICH WILL THEN BE TESTED TO SEE IF THEY AFFECT TRANSCRIPTION OR TRANSLATION OF REPORTER GENES UNDER CONTROL AND COLD STRESS CONDITIONS.THE ULTIMATE GOALS OF THIS PROJECT ARE 1) TODETERMINE IF AND HOW MITOCHONDRIAL CALCIUM TRANSPORTATION PARTICIPATES IN PLANT COLD RESPONSE OR TOLERANCE AND 2) TO IDENTIFY GENETIC SEQUENCES OR MOTIFS THAT PROMOTE GENE TRANSLATION UNDER COLD STRESS IN CORN. COMPLETION OF THESE GOALS WILL PROVIDE NOVEL TARGETS FOR BREEDING OR ENGINEERING IMPROVED COLD TOLERANCE IN CORN, PARTICULARLY IN THE CASE O,F THE TRANSLATIONAL REGULATORS WHICH WOULD BEE EXPECTED TO ACTIVATE ONLY UNDER COLD STRESS WITHOUT HAVING A SEVERE IMPACT ON NORMAL GROWTH. UTILIZATION OF THESE FINDINGS FOR BREEDING AND ENGINEERING PURPOSES IS EXPECTED TO RESULT IN THE CREATION OF COLD-RESILIENTCROPS WHICH COULD MORE RELIABLE BE PLANTED EARLIER IN THE GROWING SEASON TO IMPROVE CROP YIELDS. ADDITIONALLY, THIS WORKFLOW COULD BE APPLIED TO OTHER STRESS CONDITIONS TO IDENTIFY OTHER TRANSLATIONAL REGULATORY ELEMENTS THAT ARE ACTIVATED BY VARIOUS STRESSORS SUCH AS DROUGHT, HEAT OR VARIOUS PATHOGENS TO IMPROVE CORN TOLERANCE TO THESE ENVIRONMENTAL STRESSES.