TCP transcription factors interact with AS2 in the repression of class-I KNOX genes in Arabidopsis thaliana

Leaf organogenesis occurs within the peripheral zone of the shoot apical meristem (SAM). The initiation and subsequent development of a leaf requires the stable repression of a highly conserved class of plant genes, namely class-I KNOTTED 1-like homeobox (KNOX) genes. In Arabidopsis, this class comprises four members: SHOOT MERISTEMLESS (STM); BREVIPEDICELLUS (BP); KNAT2 and KNAT6. Two transcription factors, ASYMMETRIC LEAVES 1 (AS1) and AS2, are known to form a protein complex to repress BP, KNAT2 and KNAT6. Here, we show that AS2 physically interacts with the microRNA319 (miR319)-regulated CINCINNATA-like TEOSINTE BRANCHED 1-CYCLOIDEA-PCF (TCP) transcription factors in vitro and in vivo. By chromatin immunoprecipitation, we demonstrated that AS2 and TCPs bind to similar regions of the BP and KNAT2 promoters. In addition, DNA-binding activities of the TCP proteins rely on the presence of AS2, as the activities were dramatically reduced in the as2 mutant. The jaw-D mutant, which overexpresses MIR319a to downregulate several target TCP genes, strongly enhanced the as2 phenotypes and caused more severe ectopic expression of BP, KNAT2 and KNAT6. Our results reveal that KNOX repression requires different types of transcription factors that function together to ensure normal leaf development.