Dataset Information

Six ALPL gene variants in five children with hypophosphatasia.

ABSTRACT:

Background

Hypophosphatasia (HPP) is a rare hereditary disorder characterized by defective bone and tooth mineralization caused by mutations in the alkaline phosphatase (ALPL) gene encoding tissue-nonspecific alkaline phosphatase (TNSALP). Here we performed clinical and molecular studies on 5 HPP children to investigate the pathogenic mechanisms of the ALPL gene variants.

Methods

Clinical and genetic analyses were performed on 5 HPP children, and the loci where ALPL variants were identified. Plasmids containing the relevant loci were constructed. The molecular and cellular mechanisms of the pathogenic ALPL variants were investigated by cellular immunofluorescence, enzyme activity assay, and protein expression assay.

Results

A total of 6 ALPL variants were identified in 5 HPP children: proband 1: c.346G>A (p.A116T); proband 2: c.346G>A (p.A116T)/deletions from c.1097 to c.1099 CCT (p.T366_S367deli) compound heterozygous variant; proband 3: insertion of G from c.1014 to c.1015 (p.H338fs)/c.1446C>A (p.H482Q) compound heterozygous variant; proband 4: c.920C>T (p.P307L); and proband 5: c.883A>G (p.M295V). Twenty-four hours after the HEK-293T was transfected with different variant plasmids, its alkaline phosphatase activity and enzyme protein content were reduced compared with the wild type, and there were differences among different variants. Except for 1014-G-1015+C1446A, the degree of reduction in enzyme activity was negatively correlated with the severity of clinical manifestations. Immunofluorescence revealed that the variants (especially c.883A>G and c.920C>T) caused a decrease in alkaline phosphatase expression in the cellular membrane.

Conclusions

In total, 3 novel variants were identified in these 5 HPP children, the discovery of which will enrich the human ALPL gene mutation database. Different variants in the ALPL gene can downregulate the activity of TNSALP enzyme (and thus affect its function) by affecting protein expression and translational modifications. The same variant may cause clinical manifestations of different severities in different individuals due to the presence of dominant negative effects, alterations in noncoding sequences, blind area of intron regulatory region sequencing, and variations in environmental and individual factors. The molecular mechanisms via which the ALPL gene exerts its expression effect in vivo are highly variable and warrant further investigation.

SUBMITTER: Su N

PROVIDER: S-EPMC8184488 | biostudies-literature |

REPOSITORIES: biostudies-literature

ACCESS DATA

Similar Datasets

Project description:Loss-of-function mutations in ALPL result in hypophosphatasia (HPP), an inborn error of metabolism that causes defective skeletal and dental mineralization. ALPL encodes tissue-nonspecific alkaline phosphatase, an enzyme expressed in bone, teeth, liver, and kidney that hydrolyzes the mineralization inhibitor inorganic pyrophosphate. As Alpl-null mice die before weaning, we aimed to generate mouse models of late-onset HPP with extended life spans by engineering a floxed Alpl allele, allowing for conditional gene ablation (conditional knockout [cKO]) when crossed with Cre recombinase transgenic mice. The authors hypothesized that targeted deletion of Alpl in osteoblasts and selected dental cells ( Col1a1-cKO) or deletion in chondrocytes, osteoblasts, and craniofacial mesenchyme ( Prx1-cKO) would phenocopy skeletal and dental manifestations of late-onset HPP. Col1a1-cKO and Prx1-cKO mice were viable and fertile, and they did not manifest the epileptic seizures characteristic of the Alpl-/- model of severe infantile HPP. Both cKO models featured normal postnatal body weight but significant reduction as compared with wild type mice by 8 to 12 wk. Plasma alkaline phosphatase for both cKO models at 24 wk was reduced by approximately 75% as compared with controls. Radiography revealed profound skeletal defects in cKO mice, including rachitic changes, hypomineralized long bones, deformations, and signs of fractures. Microcomputed tomography confirmed quantitative differences in cortical and trabecular bone, including decreased cortical thickness and mineral density. Col1a1-cKO mice exhibited classic signs of HPP dentoalveolar disease, including short molar roots with thin dentin, lack of acellular cementum, and osteoid accumulation in alveolar bone. Prx1-cKO mice exhibited the same array of periodontal defects but featured less affected molar dentin. Both cKO models exhibited reduced alveolar bone height and 4-fold increased numbers of osteoclast-like cells versus wild type at 24 wk, consistent with HPP-associated periodontal disease. These novel models of late-onset HPP can inform on long-term skeletal and dental manifestations and will provide essential tools to further studies of etiopathologies and therapeutic interventions.