Stat5a Promotes Gene Expression During Chondrocyte Hypertrophic Differentiation

Overview

Journal Am J Transl Res

Specialty General Medicine

Date 2023 Jul 12

PMID 37434818

Authors

Xuan Wu

Chen Chen

Tiaotiao Han

Huiqin Bian

Jinnan Chen

Ruoxuan Hei

Ye Meng

Chenjing Wu

Yaojuan Lu

Junxia Gu

Longwei Qiao

Qiping Zheng

Affiliations

Soon will be listed here.

Abstract

Objectives: Multiple transcription factors (TFs) have previously been shown to control hypertrophic chondrocyte-specific mouse type X collagen gene () expression via interaction with promoters. This study aims to investigate the role and mechanism of the potential binding factor signal transduction and transcription activator 5a (Stat5a) of cis-enhancer, in controlling gene expression and chondrocyte hypertrophic differentiation.

Methods: The potential regulator was predicted by the transcription factor affinity prediction (TRAP) analysis of the 150-bp cis enhancer. Stat5a was screened and verified by qRT-PCR, western blot and IHC analyses. Transfection of Stat5a siRNA or expression plasmid into MCT and ATDC5 cells was performed to either knockdown or over-express Stat5a and to investigate the influence of Stat5a on gene expression during the chondrocyte hypertrophy. Dual-luciferase reporter assay was performed to explore the mechanism of Stat5a affecting transcription. Alcian blue, alkaline phosphatase, and alizarin red staining, as well as qRT-PCR analyses of related marker genes were performed to investigate the effect and possible mechanism of Stat5a on chondrocyte differentiation.

Results: The potential binding factor of cis-enhancer Stat5a and Col10a1 were both highly expressed and positively correlated within hypertrophic chondrocytes and . Knockdown of Stat5a reduced Col10a1 expression, while overexpression of Stat5a enhanced Col10a1 expression in hypertrophic chondrocytes, suggesting Stat5a as a positive Col10a1 regulator. Mechanistically, Stat5a was shown to potentiate the reporter activity mediated by promoter/enhancer. In addition, Stat5a increased the intensity of alkaline phosphatase staining of ATDC5 cells and the expression of relevant hypertrophic marker genes including Runx2, which was consistent with the expression of Stat5a and Col10a1.

Conclusions: Our results support that Stat5a promoted Col10a1 expression and chondrocyte hypertrophic differentiation, possibly via interaction with the 150-bp cis-enhancer.

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