Expression of two novel alternatively spliced COL2A1 isoforms during chondrocyte differentiation

Audrey McAlinden, Brian Johnstone, John Kollar, Najam Kazmi, Thomas M. Hering

Research output: Contribution to journalArticle

19 Citations (Scopus)

Abstract

Alternative splicing of the type II procollagen gene (COL2A1) is developmentally regulated during chondrogenesis. Type IIA procollagen (+ exon 2) is synthesized by chondroprogenitor cells while type IIB procollagen (- exon 2) is synthesized by differentiated chondrocytes. Here, we report expression of two additional alternatively spliced COL2A1 isoforms during chondrocyte differentiation of bone marrow-derived mesenchymal stem cells (MSCs). One isoform, named IIC, contains only the first 34 nucleotides of exon 2 by the use of an alternative 5′ splice site, resulting in a premature termination codon and possible nonsense-mediated decay of IIC mRNA. Low levels of the IIC isoform were detected by RT-PCR and Southern analysis of COL2A1 cDNA amplified from differentiating rabbit and human MSCs. A second novel transcript, named IID, arises by the use of another 5′ alternative splice site in intron 2. The IID isoform contains exon 2 plus 3 nucleotides, resulting in the insertion of an additional amino acid. The IID isoform was co-expressed with the IIA isoform during chondrogenesis, and was approximately one-third as abundant. Deletion of the IIC alternative 5′ splice site from a COL2A1 mini-gene construct resulted in a significant increase in the IIA:IIB ratio. A mutant mini-gene that inhibited production of the IID isoform, however, had differential effects on the production of the IIA and IIB isoforms: this was apparently related to the differentiation status of the cell type used. These data suggest that COL2A1 mRNA abundance and other aspects of chondrocyte differentiation may be regulated by the use of these previously undetermined alternative splice sites.

Original languageEnglish (US)
Pages (from-to)254-266
Number of pages13
JournalMatrix Biology
Volume27
Issue number3
DOIs
StatePublished - Apr 2008
Externally publishedYes

Fingerprint

RNA Splice Sites
Chondrocytes
Protein Isoforms
Exons
Chondrogenesis
Procollagen
Mesenchymal Stromal Cells
Nucleotides
Nonsense Mediated mRNA Decay
Genes
Collagen Type II
Nonsense Codon
Alternative Splicing
Introns
Cell Differentiation
Complementary DNA
Bone Marrow
Rabbits
Amino Acids
Polymerase Chain Reaction

Keywords

  • Alternative splicing
  • Cartilage
  • Chondrocyte
  • Chondrogenesis
  • COL2A1
  • Mesenchymal stem cell
  • Procollagen type II

ASJC Scopus subject areas

  • Molecular Biology

Cite this

Expression of two novel alternatively spliced COL2A1 isoforms during chondrocyte differentiation. / McAlinden, Audrey; Johnstone, Brian; Kollar, John; Kazmi, Najam; Hering, Thomas M.

In: Matrix Biology, Vol. 27, No. 3, 04.2008, p. 254-266.

Research output: Contribution to journalArticle

McAlinden, Audrey ; Johnstone, Brian ; Kollar, John ; Kazmi, Najam ; Hering, Thomas M. / Expression of two novel alternatively spliced COL2A1 isoforms during chondrocyte differentiation. In: Matrix Biology. 2008 ; Vol. 27, No. 3. pp. 254-266.
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