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Published on: December 18, 2019
Differential gene expression by Osterix knockdown in mouse chondrogenic ATDC5 cells
Seung-Yoon Park1, Jung-Eun Kim
1Department of Biochemistry, School of Medicine, Dongguk University, and Medical Institute of Dongguk University, Gyeongju, 780-714, Republic of Korea.
Abstract:
Osterix (Osx) is a transcription factor required for osteoblast differentiation during intramembranous and endochondral ossification. Recently, several reports have described novel functions of Osx in chondrocyte differentiation. In an in vitro study, in which the effects of Osx gene silencing were examined in mouse chondrogenic ATDC5 cells, chondrocyte marker genes were found to be expressionally downregulated and chondrocyte differentiation reduced. On the other hand, in vivo studies based on chondrocyte-specific Osx knockouts demonstrated impaired endochondral bone formation with delayed chondrocyte differentiation and reduced cartilage matrix ossification. However, little is known about the mechanism or targets of Osx involved in the control of chondrocyte differentiation. Here, we attempted to high-density of Affymetrix GeneChip microarray to investigate global gene expression profile changes caused by Osx knockdown in ATDC5 chondrocytes. The mRNA expressions of 112 genes were significantly modified by Osx knockdown: 68 genes were upregulated and 44 genes downregulated. Functional categories of gene expression classified by gene ontology demonstrated that genes related to cell adhesion, development, and signal transduction were highly affected by Osx knockdown. The expressions of differential genes, such as Sfrp2, Sema3a, Nox4, Rgs4, Zfp521, Has2, Sox6, Scn2a1, Sirpa, and Thbs2, were validated by quantitative real-time PCR. This study shows that expression profiling can be used to identify genes that are transcriptionally modified following Osx knockdown and to reveal the molecular mechanism of chondrocyte differentiation regulated by Osx.
Insights
Osterix (Osx) is crucial for bone development. This study reveals Osx regulates chondrocyte differentiation by altering gene expression, identifying key molecular targets involved in cartilage formation and ossification.
Area of Science:
- Molecular Biology
- Developmental Biology
- Genetics
Background:
- Osterix (Osx) is a key transcription factor essential for osteoblast differentiation.
- Emerging evidence suggests novel roles for Osx in chondrocyte differentiation.
- The precise mechanisms and targets of Osx in chondrocytes remain largely unelucidated.
Purpose of the Study:
- To investigate the global gene expression profile changes induced by Osx knockdown in chondrocytes.
- To identify novel Osx target genes involved in chondrocyte differentiation.
- To elucidate the molecular mechanisms underlying Osx-mediated chondrogenesis.
Main Methods:
- Utilized high-density Affymetrix GeneChip microarray for gene expression profiling.
- Performed Osx gene silencing (knockdown) in mouse chondrogenic ATDC5 cells.
- Validated differential gene expression using quantitative real-time PCR.
Main Results:
- Osx knockdown significantly altered the mRNA expression of 112 genes in ATDC5 chondrocytes (68 upregulated, 44 downregulated).
- Functional analysis revealed Osx knockdown impacts genes involved in cell adhesion, development, and signal transduction.
- Specific genes like Sfrp2, Sema3a, Nox4, and Sox6 showed significant expression changes.
Conclusions:
- Gene expression profiling effectively identifies Osx-modulated genes in chondrocytes.
- Osx plays a critical role in regulating chondrocyte differentiation through transcriptional control of specific target genes.
- This study provides insights into the molecular pathways governing chondrogenesis regulated by Osx.
