Growth retardation in a mouse model of Kabuki syndrome 2 bears mechanistic similarities to Kabuki syndrome 1.
Growth deficiency is a characteristic feature of both Kabuki syndrome 1 (KS1) and Kabuki syndrome 2 (KS2), Mendelian disorders of the epigenetic machinery with similar phenotypes but distinct genetic etiologies. We previously described skeletal growth deficiency in a mouse model of KS1 and further established that a Kmt2d -/- chondrocyte model of KS1 exhibits precocious differentiation. Here we characterized growth deficiency in a mouse model of KS2, Kdm6a tm1d/+. We show that Kdm6a tm1d/+ mice have decreased femur and tibia length compared to controls and exhibit abnormalities in cortical and trabecular bone structure. Kdm6a tm1d/+ growth plates are also shorter, due to decreases in hypertrophic chondrocyte size and hypertrophic zone height. Given these disturbances in the growth plate, we generated Kdm6a -/- chondrogenic cell lines. Similar to our prior in vitro model of KS1, we found that Kdm6a -/- cells undergo premature, enhanced differentiation towards chondrocytes compared to Kdm6a +/+ controls. RNA-seq showed that Kdm6a -/- cells have a distinct transcriptomic profile that indicates dysregulation of cartilage development. Finally, we performed RNA-seq simultaneously on Kmt2d -/-, Kdm6a -/-, and control lines at Days 7 and 14 of differentiation. This revealed surprising resemblance in gene expression between Kmt2d -/- and Kdm6a -/- at both time points and indicates that the similarity in phenotype between KS1 and KS2 also exists at the transcriptional level. Overall design: We generated Kdm6a -/- and Kdm6a +/+ cell lines from the murine teratocarcinoma parental cell line ATDC5 using CRISPR-Cas9. For the first experiment, we differentiated Kdm6a -/- and Kdm6a +/+ cells towards chondrocytes for 14 days. We then harvested RNA and performed library prep for RNA-seq. For the second experiment, we performed chondrogenic differentiation on Kdm6a -/- and Kdm6a +/+ cells as well as Kmt2d -/- and Kmt2d +/+ cell lines, previously generated from the same parental ATDC5 cell line. We collected samples for RNA-seq at both Day 7 and Day 14 of differentiation for all cell lines.