A retrotransposon gag-like-3 gene RTL3 and SOX-9 co-regulate the expression of COL2A1 in chondrocytes.

Title

A retrotransposon gag-like-3 gene RTL3 and SOX-9 co-regulate the expression of COL2A1 in chondrocytes.

Creator

Ball HC;Ansari M Y;Ahmad N;Novak K;Haqqi TM

Publisher

Connective Tissue Research

Date

2020
2020-10-12

Description

PURPOSE: Transposable elements are known to remodel gene structure and provide a known source of genetic variation. Retrotransposon gag-like-3 (RTL3) is a mammalian retrotransposon-derived transcript (MART) whose function in the skeletal tissue is unknown. This study aimed to elucidate the biological significance of RTL3 in chondrogenesis and type-II collagen (COL2A1) gene expression in chondrocytes. MATERIALS AND METHODS: Expression of RTL3, SOX-9 and COL2A1 mRNAs was determined by TaqMan assays and the protein expression by immunoblotting. RTL3 and Sox-9 depletion in human chondrocytes was achieved using validated siRNAs. An RTL3 mutant (∆RTL3) lacking the zinc finger domain was created using in vitro mutagenesis. Forced expression of RTL3, ∆RTL3, and SOX-9 was achieved using CMV promoter containing expression plasmids. CRISPR-Cas9 was utilized to delete Rtl3 and create a stable ATDC5(Rlt3-/-) cell line. Matrix deposition and Col2a1 quantification during chondrogenesis were determined by Alcian blue staining and Sircol™ Soluble Collagen Assay, respectively. RESULTS: RTL3 is not ubiquitously expressed but showed strong expression in cartilage, chondrocytes and synoviocytes but not in muscle, brain, or other tissues analyzed. Loss-of-function and gain-of-function studies demonstrated a critical role of RTL3 in the regulation of SOX-9 and COL2A1 expression and matrix synthesis during chondrogenesis. Both RTL3 and SOX-9 displayed co-regulated expression in chondrocytes. Gene regulatory activity of RTL3 requires the c-terminal CCHC zinc-finger binding domain. CONCLUSIONS: Our results identify a novel regulatory mechanism of COL2A1 expression in chondrocytes that may help to further understand the skeletal development and the pathogenesis of diseases with altered COL2A1 expression.

Subject

gene regulation; chondrogenesis; extracellular matrix; chondrocyte; ZCCHC5/RTL3; RTL3; ZCCHC5

Format

journalArticle

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Rights

Article information provided for research and reference use only. All rights are retained by the journal listed under publisher and/or the creator(s).

Pages

1-14

ISSN

1607-8438 0300-8207

NEOMED College

NEOMED College of Medicine
NEOMED College of Pharmacy

NEOMED Department

Department of Anatomy & Neurobiology
Department of Pharmacy Practice
NEOMED Student Publications

Update Year & Number

October 2020 List

Citation

Ball HC;Ansari M Y;Ahmad N;Novak K;Haqqi TM, “A retrotransposon gag-like-3 gene RTL3 and SOX-9 co-regulate the expression of COL2A1 in chondrocytes.,” NEOMED Bibliography Database, accessed January 26, 2021, https://neomed.omeka.net/items/show/11345.

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