Stimulation of the Runx2 P1 promoter by collagen-derived dipeptide prolyl-hydroxyproline bound to Foxg1 and Foxo1 in osteoblasts.

Kaho Nomura,Aya Kataoka-Matsushita,Yoshifumi Kimira,Yoshihiro Osawa,Hiroshi Mano,Masahiro Wada,Yoshiaki Sugita,Jun Shimizu,Koichi Takao

doi:10.1042/bsr20210304

Abstract

Collagen-derived dipeptide prolyl-hydroxyproline (Pro-Hyp) directly binds to the forkhead box g1 (Foxg1) protein and causes it to undergo structural alteration. Pro-Hyp also promotes the production of a regulator of osteoblast differentiation, Runt-related transcription factor 2 (Runx2), through Foxg1, inducing osteoblast differentiation. In addition, Pro-Hyp disrupts the interaction between Foxg1 and Runx2, and Foxg1 appears to interact with Runx2 in the absence of Pro-Hyp. To elucidate the mechanism of Pro-Hyp that promotes osteoblast differentiation, we investigated whether Pro-Hyp regulates the Runx2 P1 promoter together with Foxg1. The present study revealed that Pro-Hyp is taken up by osteoblastic cells via the solute carrier family 15 member (Slc15a) 4. In the presence of Pro-Hyp, Runx2 is translocated from the nucleus to the cytoplasm and Foxg1 is translocated from the cytoplasm to the nucleus. We also found that Pro-Hyp promoted the interaction between Forkhead box o1 (Foxo1) and Runx2 and the dissociation of Foxg1 from Runx2. Moreover, we identified the Pro-Hyp response element in the Runx2 distal P1 promoter at nt −375 to −316, including the Runx2 binding sites and Fox core sequence. In the presence of Pro-Hyp, Runx2 is dissociated from the Pro-Hyp response element in the Runx2 distal P1 promoter. Subsequently, Foxg1 and Foxo1 activated the Runx2 promoter by binding to the Pro-Hyp response element. In summary, we delineated the mechanism by which Pro-Hyp stimulates the bone-related Runx2 distal P1 promoter activity in osteoblastic cells through Foxg1, Foxo1, and Runx2.

Highlights

Bones are composed of osteoblasts embedded in an extracellular matrix of organic compounds and inorganic compounds, mainly comprising type 1 collagen and hydroxyapatite, respectively
prolyl hydroxyproline (Pro-Hyp) binds to Forkhead box o1 (Foxo1) and promotes interaction between Foxo1 and Runt-related transcription factor 2 (Runx2)
Foxo1 did not bind to the control magnetic beads (Supplementary Data S2). These data suggest that Pro-Hyp binds to Foxo1

Summary

Introduction

Bones are composed of osteoblasts embedded in an extracellular matrix of organic compounds and inorganic compounds, mainly comprising type 1 collagen and hydroxyapatite, respectively. During bone formation, osteoblasts secrete type 1 collagen and osteocalcin, which play a central role [1]. Osteoclasts secrete both hydrochloric acid, which resorbs hydroxyapatite, and proteases, such as matrix metalloproteinase (MMP) and cathepsin K, which degrade collagen and other bone matrix proteins [2]. Bone formation and bone resorption are linked and regulated by coupling factors [3]. Balancing the two processes is essential for healthy bone formation. Osteoblast differentiation is necessary for bone formation because osteoblasts secrete collagen and facilitate the deposition of hydroxyapatite in the bone matrix

Methods

Results

Discussion

Conclusion