SUMMARY: The paths of transfer of phosphate groups during uptake of inorganic orthophosphate by Micrococcus pyogenes var. aureus (strain Duncan) have been studied by following the incorporation of 32P-labelled phosphate groups into the phospholipid (LP), acid-soluble inorganic (AI) and organic (AO), deoxyribonucleic acid (DNA), ribonucleic acid (RNA) and glycerophospho-protein complex (PGP) fractions of the cells. In resting cells a strictly reciprocal exchange of inorganic phosphate across the osmotic barrier is half complete in 70 min. During respiration or growth, phosphate moves inward through the osmotic barrier at the same rate as during rest, but the outward movement is abolished. Phosphate accumulates mainly in the AI, AO and LP during respiration, but in all the fractions during growth. Most of the phosphate of the organic fractions is drawn through the AI. The RNA is formed from the nucleotides of the AO, while the DNA phosphate is derived either directly from the AI or from intermediates in the AO. Little or no turnover of RNA or DNA phosphate occurs during growth, and phosphate is not transferred directly or indirectly from RNA to DNA or vice versa. The PGP, although the major constituent of the mechanically rigid cell envelope, does not participate as a whole in phosphate transfer reactions during rest or respiration; but during growth the amount of PGP increases in proportion to the expansion of cell envelope area. The LP phosphate exhibits rapid turnover during respiration and growth.