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Sp1 upregulates the proximal promoter activity of the mouse collagen α1(XI) gene (Col11a1) in chondrocytes

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Abstract

Type XI collagen is a cartilage-specific extracellular matrix, and is important for collagen fibril formation and skeletal morphogenesis. We have previously reported that NF-Y regulated the proximal promoter activity of the mouse collagen α1(XI) gene (Col11a1) in chondrocytes (Hida et. al. In Vitro Cell. Dev. Biol. Anim. 2014). However, the mechanism of the Col11a1 gene regulation in chondrocytes has not been fully elucidated. In this study, we further characterized the proximal promoter activity of the mouse Col11a1 gene in chondrocytes. Cell transfection experiments with deletion and mutation constructs indicated that the downstream region of the NF-Y binding site (-116 to +1) is also necessary to regulate the proximal promoter activity of the mouse Col11a1 gene. This minimal promoter region has no TATA box and GC-rich sequence; we therefore examined whether the GC-rich sequence (-96 to -67) is necessary for the transcription regulation of the Col11a1 gene. Luciferase assays using a series of mutation constructs exhibited that the GC-rich sequence is a critical element of Col11a1 promoter activity in chondrocytes. Moreover, in silico analysis of this region suggested that one of the most effective candidates was transcription factor Sp1. Consistent with the prediction, overexpression of Sp1 significantly increased the promoter activity. Furthermore, knockdown of Sp1 expression by siRNA transfection suppressed the proximal promoter activity and the expression of endogenous transcript of the mouse Col11a1 gene. Taken together, these results indicate that the transcription factor Sp1 upregulates the proximal promoter activity of the mouse Col11a1 gene in chondrocytes.

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Acknowledgments

We thank Ms. H Satoh for the technical support, and the staff members of the Research Promotion Institute, Oita University. This work was supported by Grants-In-Aid for Scientific Research (No. 21591952 to N.M.) from the Ministry of Education, Culture, Sports, Science and Technology of Japan.

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Authors and Affiliations

  1. Department of Matrix medicine, Faculty of medicine, Oita University, 1-1 Idaigaoka, Hasama-machi, Yufu, Oita, 879-5593, Japan

    Keijirou Watanabe, Mariko Hida, Takako Sasaki, Hidekatsu Yoshioka & Noritaka Matsuo

  2. Department of Oral and Maxillo-Facial Surgery, Faculty of medicine, Oita University, 1-1, Idaigaoka, Hasama-machi, Yufu, Oita, 879-5593, Japan

    Keijirou Watanabe & Kenji Kawano

  3. Division of Radioisotope Research, Department of Research Support, Research Promotion Project, Faculty of medicine, Oita University, 1-1 Idaigaoka, Hasama-machi, Yufu, Oita, 879-5593, Japan

    Hiroyuki Yano

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  1. Keijirou Watanabe
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Correspondence to Noritaka Matsuo.

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Watanabe, K., Hida, M., Sasaki, T. et al. Sp1 upregulates the proximal promoter activity of the mouse collagen α1(XI) gene (Col11a1) in chondrocytes. In Vitro Cell.Dev.Biol.-Animal 52, 235–242 (2016). https://doi.org/10.1007/s11626-015-9959-y

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