Cytokinins are mobile phytohormones that regulate plant growth, development, and environmental adaptability. The major cytokinin species include isopentenyl adenine (IP), trans-zeatin (tZ), cis-zeatin (cZ), and dihydrozeatin (DZ). These cytokinin species are distributed in different organelles, cells, tissues, and organs, primarily shaped by biosynthesis via isopentenyltransferases (IPT), cytochrome P450 monooxygenase, and 5′-ribonucleotide phosphohydrolase, as well as conjugation or catabolism via glycosyltransferase or cytokinin oxidase/dehydrogenase. Cytokinins bind to histidine receptor kinases in the endoplasmic reticulum or plasma membrane and relay signals to response regulators in the nucleus via histidine phosphotransfer proteins. The movement of cytokinins from sites of biosynthesis to sites of signal perception requires long-distance, intercellular, and intracellular transport. ATP-binding cassette (ABC) transporters, purine permeases (PUP), AZA-GUANINE RESISTANT (AZG) transporters, equilibrative nucleoside transporters (ENT), and Sugars Will Eventually Be Exported transporters (SWEET) have been characterized as involved in cytokinin transport processes. This review introduces the spatial distributions of various cytokinins and the subcellular localizations of the proteins involved in their metabolism and signaling. It highlights an inventory of characterized transporters involved in cytokinin compartmentalization and the regulation of spatial distributions by environmental cues. Future directions for cytokinin research are also discussed.Cytokinins are mobile phytohormones that regulate plant growth, development, and environmental adaptability. The major cytokinin species include isopentenyl adenine (IP), trans-zeatin (tZ), cis-zeatin (cZ), and dihydrozeatin (DZ). These cytokinin species are distributed in different organelles, cells, tissues, and organs, primarily shaped by biosynthesis via isopentenyltransferases (IPT), cytochrome P450 monooxygenase, and 5′-ribonucleotide phosphohydrolase, as well as conjugation or catabolism via glycosyltransferase or cytokinin oxidase/dehydrogenase. Cytokinins bind to histidine receptor kinases in the endoplasmic reticulum or plasma membrane and relay signals to response regulators in the nucleus via histidine phosphotransfer proteins. The movement of cytokinins from sites of biosynthesis to sites of signal perception requires long-distance, intercellular, and intracellular transport. ATP-binding cassette (ABC) transporters, purine permeases (PUP), AZA-GUANINE RESISTANT (AZG) transporters, equilibrative nucleoside transporters (ENT), and Sugars Will Eventually Be Exported transporters (SWEET) have been characterized as involved in cytokinin transport processes. This review introduces the spatial distributions of various cytokinins and the subcellular localizations of the proteins involved in their metabolism and signaling. It highlights an inventory of characterized transporters involved in cytokinin compartmentalization and the regulation of spatial distributions by environmental cues. Future directions for cytokinin research are also discussed.