Genome-wide characterization of the TALE homeodomain family and the KNOX-BLH interaction network in tomato.

Abstract

Comprehensive yeast and protoplast two-hybrid analyses illustrated the protein-protein interaction network of the TALE homeodomain protein family, KNOX and BLH proteins, in tomato leaf and fruit development. KNOTTED-like (KNOX, KN) proteins and BELL1-like (BLH) proteins, which belong to the same TALE homeodomain family, act together by forming KNOX-BLH heterodimer modules. These modules play crucial roles in regulating multiple developmental processes in plants, like organ differentiation. However, despite the increasing knowledge about individual KNOX and BLH functions, a comprehensive view of their functional protein-protein interaction (PPI) network remains elusive in most plants, including tomato (Solanum lycopersicum), an important model plant to study fruit and leaf development. Here, we characterized eight tomato KNOX genes (SlKN1 to SlKN8) and fourteen tomato BLH genes (SlBLH1 to SlBLH14) by expression profiling, co-expression analysis, and PPI network analysis using two-hybrid techniques in yeasts (Y2H) and protoplasts (P2H). We identified 75 pairwise KNOX-BLH interactions, including ten novel interactors of SlKN2/TKN2, a primary class I KNOX protein, and nine novel interactors of SlKN5, a primary class II KNOX protein. Based on these data, we classified KNOX-BLH modules into several categories, which made us infer the order and combination of the KNOX-BLH modules involved in differentiation processes in leaf and fruit. Notably, the co-expression and interaction of SlKN5 and fruit preferentially expressing BLH1-clade paralogs (SlBLH5/SlBEL11 and SlBLH7) suggest their important roles in regulating fruit differentiation. Furthermore, in silico modeling of the KNOX-BLH modules, sequence analysis, and P2H assay identified several residues and a linker region potentially influencing the affinity of BLHs to KNOXs within their conserved dimerization domains. Together, these findings provide insights into the regulatory mechanism of KNOX-BLH modules underlying tomato organ differentiation.