HORMA domain-containing proteins play important roles in cell cycle regulation and DNA repair, acting as adaptors to recruit other proteins. Although containing the conserved HORMA domain in structure, different types of HORMA proteins have diverged significantly in function. The mechanisms underlying the evolution and functional diversification of HORMA proteins remain unclear. Here we conduct an integrative approach, combining sequence, structural, gene coexpression and protein-protein interaction data, to trace the structural and functional evolution of HORMA proteins. Comparative sequence and structure analysis revealed that variations in both amino acid sequence and domain composition contributed to the functional diversification of different HORMA domain-containing proteins. Multiple amino acid substitutions at the C-terminal region promoted the functional divergence between MAD2 and REV7 by facilitating interaction with different partners. The emergence of the HOP1CTD domain contributed to the function of HOP1 as a meiosis-specific structural component of the lateral elements of the synaptonemal complex. Additionally, different types of HORMA proteins were recruited into different functional modules in the genetic network while functioning distinctly.