Carboxy-Terminal Cementum Protein 1-Derived Peptide 4 (cemp1-p4) Promotes Mineralization through wnt/β-catenin Signaling in Human Oral Mucosa Stem Cells.

Rita Arroyo,Higinio Arzate,Sonia López,Yonathan Garfias,Enrique Romo,Claudia Pedraza,Gonzalo Montoya,Lía Hoz

doi:10.3390/ijms21041307

Rita Arroyo, Higinio Arzate + Show 6 more

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https://doi.org/10.3390/ijms21041307

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Abstract

Human cementum protein 1 (CEMP1) is known to induce cementoblast and osteoblast differentiation and alkaline phosphatase (ALP) activity in human periodontal ligament-derived cells in vitro and promotes bone regeneration in vivo. CEMP1′s secondary structure analysis shows that it has a random-coiled structure and is considered an Intrinsic Disordered Protein (IDP). CEMP1′s short peptide sequences mimic the biological capabilities of CEMP1. However, the role and mechanisms of CEMP1′s C-terminal-derived synthetic peptide (CEMP1-p4) in the canonical Wnt/β-catenin signaling pathway are yet to be described. Here we report that CEMP1-p4 promotes proliferation and differentiation of Human Oral Mucosa Stem Cells (HOMSCs) by activating the Wnt/β-catenin pathway. CEMP1-p4 stimulation upregulated the expression of β-catenin and glycogen synthase kinase 3 beta (GSK-3B) and activated the transcription factors TCF1/7 and Lymphoid Enhancer binding Factor 1 (LEF1) at the mRNA and protein levels. We found translocation of β-catenin to the nucleus in CEMP1-p4-treated cultures. The peptide also penetrates the cell membrane and aggregates around the cell nucleus. Analysis of CEMP1-p4 secondary structure revealed that it has a random-coiled structure. Its biological activities included the induction to nucleate hydroxyapatite crystals. In CEMP1-p4-treated HOMSCs, ALP activity and calcium deposits increased. Expression of Osterix (OSX), Runt-related transcription factor 2 (RUNX2), Integrin binding sialoproptein (IBSP) and osteocalcin (OCN) were upregulated. Altogether, these data show that CEMP1-p4 plays a direct role in the differentiation of HOMSCs to a “mineralizing-like” phenotype by activating the β-catenin signaling cascade.

Highlights

The periodontium is a unique, complex and functional biological system that provides support to the dental organs
Our results showed that cementum protein 1 (CEMP1)-p4 enhanced the expression of β-catenin, while expression of Gsk3b decreased and downstream transcriptional factors Tcf1 and Lef1 were activated by CEMP1-p4 treatment [30], revealing that CEMP1-p4 activated the Wnt/β-catenin signaling pathway, inducing Human Oral Mucosa Stem Cells (HOMSCs) differentiation to a “mineralizing-like” phenotype
To determine if CEMP1-p4 possessed a secondary structure, the peptide was analyzed by circular dichroism spectroscopy (CD) and its behavior in solution was analyzed by dynamic light scattering (DLS)

Summary

Introduction

The periodontium is a unique, complex and functional biological system that provides support to the dental organs It is composed of two connective mineralized tissues—alveolar bone and cementum and two soft connective tissues, gingiva and periodontal ligament. The cellular and molecular mechanisms that regulate the formation of cementum are poorly understood [2] This unique tissue covers the root surface and in conjunction with periodontal ligament and alveolar bone, provides a functional anchoring apparatus in the periodontium [3,4,5]. A few years ago, cementum markers such as cementum protein 1 (CEMP1) and cementum attachment protein (CAP) were cloned and characterized [7,8] These two proteins have shown to promote the proliferation and differentiation of periodontal ligament cells toward a “mineralizing-like” phenotype [9,10,11]. CEMP1 s short peptide sequences are possible regulators of stem cell differentiation toward a “mineralizing-like” cell phenotype [12]

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