ABSTRACT The MADS box transcription factor SrfA is required for spore differentiation in Dictyostelium discoideum. srfA null strains form rounded spores that do not resist adverse environmental conditions. Five genes whose expression is dependent on SrfA have been isolated by differential hybridization. One of these genes, sigC , is identical to phg1b , previously characterized in mutants with altered adhesive properties and found to encode a nine-transmembrane-domain protein. This gene is transcribed into two mRNAs as the result of alternative splicing of two internal exons. The slower-migrating mRNA codes for a shorter protein that lacks the first transmembrane fragment and is not expressed in srfA null strains. The other four genes ( sigA , sigB , sigD , and 45D ) are expressed only during late developmental stages. In situ hybridization experiments showed that expression of sigA , sigB , and sigD is restricted to the sorus of developing structures. sigA codes for a homologue of malate dehydrogenase that converts pyruvate to malate to replenish the tricarboxylic acid cycle. sigB encodes a protein with significant similarity to the GP63 metalloproteinase of Leishmania , leishmanolysin. The sequence of SigD is highly similar to that of several spore coat proteins of D. discoideum , and it may play a role in that structure. The gene 45D codes for an RNA-binding protein homologue whose expression is also dependent on the GATA transcription factor stalky (StkA). The expression of sigB is also dependent on both SrfA and StkA. The expression of 45D , but not of sigA , sigB , sigC , and sigD , can be induced in srfA null cells by constitutive protein kinase A activation. Strains in which either sigA , sigB , or sigD is disrupted were isolated and found to form spores that are not detectably different from those of wild-type strains.