SPLIT HAND/FOOT VARIANTS FAIL TO RESCUE PRDM1A MUTANT CRANIOFACIAL DEFECTS
Abstract
Background
Split Hand/Foot Malformation (SHFM) is a congenital limb disorder presenting with limb anomalies, such as missing, hypoplastic, or fused digits, and often craniofacial defects, including a cleft lip/palate, microdontia, micrognathia, or maxillary hypoplasia. We previously identified three novel variants in the transcription factor,PRDM1, that are associated with SHFM phenotypes. One individual also presented with a high arch palate. Studies in vertebrates indicate that PRDM1 is important for development of the skull; however, prior to our study, human variants inPRDM1had not been associated with craniofacial anomalies.
Methods
Using transient mRNA overexpression assays inprdm1a-/-mutant zebrafish, we tested whether thePRDM1SHFM variants were functional and could lead to a rescue of the craniofacial defects observed inprdm1a-/-mutants. We also mined a CUT&RUN and RNA-seq dataset to examine Prdm1a binding and the effect of Prdm1a loss on craniofacial genes.
Results
prdm1a-/-mutants exhibit craniofacial defects including a hypoplastic neurocranium, a loss of posterior ceratobranchial arches, a shorter palatoquadrate, and an inverted ceratohyal. Injection of wildtypehPRDM1inprdm1a-/-mutants partially rescues these structures. However, injection of each of the three SHFM variants fails to rescue the skeletal defects. Loss ofprdm1aleads to a decreased expression of important craniofacial genes, such asdlx5a/dlx6a, hand2, sox9b, col2a1a, andhoxbgenes.
Conclusion
These data suggest that the three SHFM variants are not functional and may have led to the craniofacial defects observed in the humans. Finally, they demonstrate how Prdm1a can directly bind and regulate craniofacial gene expression.
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