Gain-of-function suppressors restore microtubule dynamics and rescue dominant-negative tubulinopathies

Missense mutations in tubulin-coding genes often exert dominant-negative effects on microtubule (MT) assembly, leading to developmental disorders collectively known as tubulinopathies. Here we performed suppressor screens in Caenorhabditis elegans and identified three functionally distinct classes of missense suppressors that counteract toxic α- and β-tubulin mutants. Intergenic suppressors-missense variants in the reciprocal tubulin gene-restore MT architecture via two distinct mechanisms: assembly-defective variants competitively exclude mutant tubulins, whereas assembly-competent variants modulate MT dynamics. These mechanisms are conserved and recapitulated in human cells and murine oocytes, where selected intergenic suppressors rescue pathogenic tubulin-induced MT defects and outperform wild-type tubulin supplementation. A systematic mutational analysis of TUBA1A further defines a landscape of gain-of-function assembly-competent suppressors, supported by molecular simulations showing restoration of protofilament geometry. Together, our results provide a structural and functional framework for tubulin suppression and highlight intergenic suppressors as potential candidates for precision therapeutics targeting dominant tubulinopathies.