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The commitment of differentiating cells to a specialized fate is fundamental to the correct assembly of tissues within a multicellular organism. Because commitment is often irreversible, entry into and progression through this phase of development must be tightly regulated. Under nitrogen-limiting conditions, the multicellular cyanobacterium Anabaena sp. strain PCC 7120 terminally commits ∼10% of its cells to become specialized nitrogen-fixing heterocysts. Although commitment is known to occur 9–14 h after the induction of differentiation, the factors that regulate the initiation and duration of this phase have yet to be elucidated. Here, we report the identification of four genes that share a functional domain and modulate heterocyst commitment: hetP (alr2818), asl1930, alr2902, and alr3234. Epistatic relationships between all four genes relating to commitment were revealed by deleting them individually and in combination; asl1930 and alr3234 acted most upstream to delay commitment, alr2902 acted next in the pathway to inhibit development, and hetP acted most downstream to drive commitment forward. Possible protein–protein interactions between HetP, its homologs, and the heterocyst master regulator, HetR, were assessed, and interaction partners were defined. Finally, patterns of gene expression for each homolog, as determined by promoter fusions to gfp and reverse transcription–quantitative PCR, were distinct from that of hetP in both spatiotemporal organization and regulation. We posit that a dynamic succession of protein–protein interactions modulates the timing and efficiency of the commitment phase of development and note that this work highlights the utility of a multicellular cyanobacterium as a model for the study of developmental processes.