Cell suspensions of the model legume Medicago truncatula accumulated the isoflavonoid phytoalexin medicarpin in response to yeast elicitor or methyl jasmonate (MJ), accompanied by decreased levels of isoflavone glycosides in MJ-treated cells. DNA microarray analysis revealed rapid, massive induction of early (iso)flavonoid pathway gene transcripts in response to yeast elicitor, but not MJ, and differential induction by the two elicitors of sets of genes encoding transcription factors, ABC transporters, and beta-glucosidases. In contrast, both elicitors induced genes encoding enzymes for conversion of the isoflavone formononetin to medicarpin. Four MJ-induced beta-glucosidases were expressed as recombinant enzymes in yeast, and three were active with isoflavone glucosides. The most highly induced beta-glucosidase was nuclear localized and preferred flavones to isoflavones. The results indicate that the genetic and biochemical mechanisms underlying accumulation of medicarpin differ depending on the nature of the stimulus and suggest a role for MJ as a signal for rapid hydrolysis of preformed, conjugated intermediates for antimicrobial biosynthesis during wound responses.