klf2ash317 Mutant XXXX Do Not Recapitulate Morpholino-Induced Vascular and Haematopoietic Phenotypes.

@entity712 @entity71892 @entity219 @entity71890 @entity71891 @entity71893 @entity280 @entity3392
INTRODUCTION AND OBJECTIVES: The @entity219 -finger transcription factor Kr  ppel-like factor 2 (KLF2) transduces blood flow into molecular signals responsible for a wide range of responses within the vasculature. KLF2 maintains a healthy, quiescent endothelial phenotype. Previous studies report a range of phenotypes following morpholino antisense oligonucleotide-induced @entity71890 knockdown in @entity3392 . Targeted genome editing is an increasingly applied method for functional assessment of candidate genes. We therefore generated a stable @entity71890 mutant @entity3392 and characterised its cardiovascular and haematopoietic development. METHODS AND RESULTS: Using Transcription Activator-Like Effector Nucleases (TALEN) we generated a @entity71890 mutant (klf2ash317) with a 14bp deletion leading to a premature stop codon in exon 2. Western blotting confirmed loss of wild type @entity71890 protein and the presence of a truncated protein in klf2ash317 mutants. Homozygous klf2ash317 mutants exhibit no defects in vascular patterning, survive to adulthood and are fertile, without displaying previously described morphant phenotypes such as high-output @entity712 , reduced haematopoetic stem cell (HSC) development or impaired formation of the 5th accessory aortic arch. Homozygous klf2ash317 mutation did not reduce angiogenesis in @entity3392 with homozygous mutations in @entity280 ( @entity280 ), a form of angiogenesis that is dependent on blood flow. We examined expression of three klf family members in wildtype and klf2ash317 @entity3392 . We detected vascular expression of @entity71891 (but not @entity71892 or @entity71893 / @entity71893 / @entity71893 ) in wildtypes but found no differences in expression that might account for the lack of phenotype in klf2ash317 mutants. @entity71891 morpholino knockdown did not affect heart rate or impair formation of the 5th accessory aortic arch in either wildtypes or klf2ash317 mutants. CONCLUSIONS: The klf2ash317 mutation produces a truncated @entity71890 protein but, unlike morpholino induced @entity71890 knockdown, does not affect cardiovascular development.
