Characterization of zebrafish IKK2 mutants generated by zinc finger nucleases

Abstract

IκB kinase β (IKK2/IKKβ) is known as a master regulator of NF-κB signaling, which mediates stress, immune and inflammatory responses. Unfortunately, embryonic lethality in IKK2-deficient mice hinders studies of zygotic IKK2 function beyond embryonic stages. Although conditional knockout mice have been used to circumvent this problem, studies using zebrafish as another vertebrate model offer an alternative approach. Recently, two zebrafish IKK2 mutants had been developed using zinc finger nuclease-mediated mutagenesis. In this work, phenotypic characterization of these two IKK2 mutants was performed. Zygotic mutants of a null allele of IKK2 (ikk2+2) showed survival rates comparable to wild-type from one-cell stage until three months post fertilization, but decreased body lengths at nine months of age. In contrast, zygotic mutants of a truncated allele of IKK2 (ikk2Δ7) containing only the kinase domain showed reduced survival rates as early as one month post fertilization with vascular defects such as blood pooling and abnormal vasculature development. In addition, Tol2 transposon-mediated knock-in was initiated to rescue embryonic lethality observed in homozygous crosses of IKK2 mutants. Taken together, results obtained establish zebrafish IKK2 mutants as valid and useful models for investigating the functional roles of IKK2 during development and dissecting the underlying molecular mechanisms.