Impaired Bone Formation in Pdia3 Deficient Mice

Abstract

1α,25-dihydroxyvitamin D3 [1α,25(OH)2D3] is crucial for normal skeletal development and bone homeostasis. Protein disulfide isomerase family A, member 3 (PDIA3) mediates 1α,25(OH)2D3 initiated-rapid membrane signaling in several cell types. To understand its role in regulating skeletal development, we generated Pdia3-deficient mice and examined the physiologic consequence of Pdia3-disruption in embryos and Pdia3 +/− heterozygotes at different ages. No mice homozygous for the Pdia3-deletion were found at birth nor were there embryos after E12.5, indicating that targeted disruption of the Pdia3 gene resulted in early embryonic lethality. Pdia3-deficiency also resulted in skeletal manifestations as revealed by µCT analysis of the tibias. In comparison to wild type mice, Pdia3 heterozygous mice displayed expanded growth plates associated with decreased tether formation. Histomorphometry also showed that the hypertrophic zone in Pdia3 +/− mice was more cellular than seen in wild type growth plates. Metaphyseal trabecular bone in Pdia3 +/− mice exhibited an age-dependent phenotype with lower BV/TV and trabecular numbers, which was most pronounced at 15 weeks of age. Bone marrow cells from Pdia3 +/− mice exhibited impaired osteoblastic differentiation, based on reduced expression of osteoblast markers and mineral deposition compared to cells from wild type animals. Collectively, our findings provide in vivo evidence that PDIA3 is essential for normal skeletal development. The fact that the Pdia3 +/− heterozygous mice share a similar growth plate and bone phenotype to nVdr knockout mice, suggests that PDIA3-mediated rapid membrane signaling might be an alternative mechanism responsible for 1α,25(OH)2D3’s actions in regulating skeletal development.

Mineral Homeostasis in Murine Fetuses Is Sensitive to Maternal Calcitriol but Not to Absence of Fetal Calcitriol

“…(12,13) But in Cyp27b1 null fetuses, in which calcitriol is absent, placental calcium transport and placental expression of Pthrp and Trpv6 are all normal, while expression of Pdia3 is reduced. These comparative results imply that the high levels of calcitriol in Vdr null fetuses are able to act on an alternative receptor, possibly Pdia3, (54)(55)(56)(57) to stimulate placental calcium transport and gene expression. This may explain why prior animal studies found that pharmacological treatment with calcitriol increased the rate of placental calcium transport.…”

“…Recent data support that protein disulfide isomerase family A, member 3 (PDIA3) may be a rapidly responsive plasma membrane receptor for calcitriol in certain tissues . Pdia3 null mice are embryonically lethal, but the heterozygous state has a skeletal phenotype similar to Vdr nulls …”

“…Recent data support that protein disulfide isomerase family A, member 3 (PDIA3) may be a rapidly responsive plasma membrane receptor for calcitriol in certain tissues. (28,(54)(55)(56)(57) Pdia3 null mice are embryonically lethal, but the heterozygous state has a skeletal phenotype similar to Vdr nulls. (55,56) The purpose of the studies presented here was to determine if loss of calcitriol, in Cyp27b1 null fetal mice, causes a similar phenotype as in Vdr null fetuses.…”

“…(28,(54)(55)(56)(57) Pdia3 null mice are embryonically lethal, but the heterozygous state has a skeletal phenotype similar to Vdr nulls. (55,56) The purpose of the studies presented here was to determine if loss of calcitriol, in Cyp27b1 null fetal mice, causes a similar phenotype as in Vdr null fetuses. If loss of calcitriol in Cyp27b1 null fetuses results in a different phenotype from Vdr null fetuses, that may imply that the Vdr null phenotype is partly due to high levels of calcitriol acting on non-classical receptors.…”

An integrated genome-wide multi-omics analysis of gene expression dynamics in the preimplantation mouse embryo

“…We selected three genes among those exhibiting strong negative correlations during preimplantation development and particularly from the 1-to the 4cell stage: Pdia3, Top1, and DNAjb11. These genes are particularly appropriate in the context of our study, because mutations of Pdia3, Top1 and Dnajb11 interfere with development and prove lethal in homozygosis(Morham et al 1996;Francisco et al 2010;Li et al 2014a;Wang et al 2014). The results of the TaqMan assay for Pdia3, Top1 and Dnajb11 correlated positively with those of RNA-seq, as did the results of the immunofluorescence with those of LC-MS/MS (see SupplementalFigure S19), confirming the existence of genes with strongly anticorrelated protein and transcript expression profiles.Finally, with the comfort of the validation data, we moved on to analyze the features of the genes at the extremes of the distribution of Spearman's rank correlation coefficients.…”