Stomata,which consist of paired guard cells,are known to have played crucial roles in the colonization of land by plants.Turgor-driven stomatal movement controls transpiration and gas exchange between plants and the environment.Several key genes and regulatory networks underlying stomatal development have been uncovered by molecular genetic analysis,however,much less is known about how signals involved in stomatal development are transmitted to RNA polymerase Ⅱ (Pol Ⅱ),which plays a central role in the transcription of mRNA coding genes.Herein,we reported a partial loss-of-function mutant of the third largest subunit of nuclear DNA-dependent RNA polymerase Ⅱ (NRPB3).It exhibited an increased number of stomatal lineage cells and stomatal clusters.Similar stomatal phenotypes were observed in a weak allele of the second largest subunit of nuclear DNA-dependent RNA polymerase Ⅱ (NRPB2).These results suggested that Pol Ⅱ plays essential roles in stomatal development.Genetic analysis indicated that NRPB3 synergistically interacted with stomatal patterning and differentiation regulators.We also found physical associations of NRPB3 with two bHLH transcription factors,FAMA and INDUCER OF CBF EXPRESSION1 (ICE1),indicating that NRPB3 serves as an acceptor for signals from transcription factors involved in stomatal development,Our findings highlight the surprisingly conserved activating mechanisms mediated by the third largest subunit of Pol Ⅱ in eukaryotes.