Abstract
Variants within Histone H4 genes have been identified that underlie a developmental disorder within a cohort of 29 patients. All individuals with these variants are affected by developmental delay and intellectual disability, as well as more global developmental anomalies. Osteogenic Growth Peptide (OGP) is a circulating regulatory peptide that regulates bone development and homeostasis and is transcribed from an internal start codon in histone H4 mRNA. Skeletal anomalies including vertebral and digit anomalies, as well as craniosynostosis are present in patients within this cohort which may indicate dysregulation of skeletal development. This project sought to investigate the effects of patient variants on the regulation and activity of OGP, to further the understanding of both this disorder and OGP.
OGP regulates bone homeostasis through regulation of proliferation and activity of fibroblastic and osteoblastic cell lines and is required for basal proliferation. Low concentrations of OGP upregulate proliferation, while high concentrations inhibit proliferation to maintain bone homeostasis. OGP also promotes differentiation of mesenchymal stem cells into osteoblasts. As dysregulation of osteoblast and mesenchymal stem cells has been previously associated with the development of skeletal anomalies, this project aimed to address the hypothesis that patient variants disrupted the function of OGP.
Previous studies have suggested the existence of regulatory elements within the H4 transcript that alter translation of OGP. This project investigated the effects of patient variants on OGP through investigation of OGP translation. A mutant H4C3 was produced that could only translate OGP, (p.M1A), and patient variants were added. HEK293FT cells transfected with variant H4C3 p.M1A did not show any effect of patient variants on the translation of OGP as measured on a quantitative western blot.
This project also studied the effects of OGP on osteogenic cell proliferation, and MSC differentiation. The results of these experiments were inconclusive; however optimization of protocols will allow for further investigation into this peptide in the future.
Investigation into the regulation and activity of OGP did not implicate this peptide in the development of skeletal anomalies in these patients. This does however indicate that variants that disrupt the histone H4 protein underlie the development of skeletal anomalies and the other global developmental disruptions present in these patients.