Related Experiment Videos
Naturally occurring antisense transcripts are present in chick embryo chondrocytes simultaneously with the
1Molecular Biology and Biochemistry Department, Wesleyan University, Middletown, Connecticut 06459-0175.
Abstract:
It has previously been shown that very low steady state levels of alpha 1 (I) collagen mRNA are present in chick embryo sternal chondrocytes (Askew, G. R., Wang, S., and Lukens, L. N. (1991) J. Biol. Chem. 266, 16834-16841), yet nuclear run-on experiments with double-stranded cDNA probes indicated a high transcription rate at this locus. These findings were investigated in this study using single-stranded probes, where nuclear run-on experiments showed that antisense transcription of the alpha 1 (I) collagen gene was occurring in chondrocytes, while sense strand transcription was down-regulated. Treatment of these chondrocytes with 5-bromo-2'-deoxyuridine (BrdU), which causes the cells to resemble their mesenchymal precursors, resulted in an antiparallel situation, where antisense transcription was lost, and instead, sense strand transcription was acquired, suggesting that the reverse switch from sense to antisense transcription occurs during chondrogenesis. Very large (> 10 kilobases) and heterogeneous antisense transcripts of moderate stability were shown to span both ends of the gene in chondrocytes, while their absence was shown in BrdU-treated chondrocytes, chick embryo fibroblasts, and a variety of other tissues. The function of these antisense transcripts is so far unknown, but their unusual chondrocyte-specific appearance, concurrent with little or no sense strand transcription, suggests a possible functional role in the down-regulation of the alpha 1 (I) collagen gene.
Insights
Antisense transcription of the alpha 1 (I) collagen gene is active in chondrocytes, while sense transcription is low. This reverse switch during chondrogenesis may regulate collagen gene expression.
Area of Science:
- Molecular Biology
- Developmental Biology
- Gene Regulation
Background:
- Low steady-state levels of alpha 1 (I) collagen mRNA exist in chick embryo chondrocytes.
- Nuclear run-on experiments with double-stranded probes suggested high transcription rates at this locus.
Purpose of the Study:
- Investigate the discrepancy between low mRNA levels and high transcription rates of alpha 1 (I) collagen.
- Determine the role of sense and antisense transcription during chondrogenesis.
Main Methods:
- Nuclear run-on experiments using single-stranded probes.
- Treatment of chondrocytes with 5-bromo-2'-deoxyuridine (BrdU).
- Analysis of antisense and sense transcript size, stability, and tissue distribution.
Main Results:
- Antisense transcription of alpha 1 (I) collagen occurs in chondrocytes, with concurrent down-regulation of sense transcription.
- BrdU treatment reversed this pattern, showing loss of antisense and gain of sense transcription, suggesting a switch during chondrogenesis.
- Large, heterogeneous antisense transcripts spanning the gene were detected specifically in chondrocytes.
Conclusions:
- A reverse switch from sense to antisense transcription occurs during chondrogenesis.
- Chondrocyte-specific antisense transcripts may play a role in down-regulating alpha 1 (I) collagen gene expression.
- The precise function of these antisense transcripts remains to be elucidated.