Differential transactivation of the upstream aggrecan enhancer regulated by PAX1/9 depends on SOX9-driven transactivation
2019
A previously identified enhancer 10 kb upstream of the
Aggrecan(Acan) gene (UE) can drive cartilage specific reporter expression in vivo. Here, we report that the paralogous transcription factors PAX1 and
PAX9differentially drive UE, depending on the presence or absence of
SOX9-driven
transactivation. In the developing
vertebral column, PAX1/9 expression was inversely correlated with Acan expression. Moreover, PAX1/9 was co-expressed with
SOX9/5/6 in the intervertebral mesenchyme and the inner annulus fibrosus (AF), and with
SOX9in the outer AF. Significant Acan upregulation was observed during chondrification of Pax1-silenced AF cells, while, Acan was significantly downregulated by persistent expression of Pax1 in cartilage. Deletion of UE using CRISPR/
Cas9resulted in ~30% and ~40% reduction of Acan expression in cartilage and the AF, respectively. In the UE, PAX1/9 acts as weak
transactivatorsthrough a PAX1/9-binding site partially overlapped with a
SOX9-binding site. In the presence of
SOX9, which otherwise drives robust Acan expression along with SOX5/6, PAX1/9 competes with
SOX9for occupancy of the binding site, resulting in reduced
transactivationof Acan. Coimmunoprecipitation revealed the physical interaction of Pax1 with
SOX9. Thus,
transactivationof the UE is differentially regulated by concerted action of PAX1/9,
SOX9, and SOX5/6 in a context-dependent manner.
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