Abstract
BackgroundMitochondria-localized glutamic acid-rich protein (MGARP) is a novel mitochondrial transmembrane protein expressed mainly in steroidogenic tissues and in the visual system. Previous studies showed that MGARP functions in hormone biosynthesis and its expression is modulated by the HPG axis.Methodology/Principal FindingsBy bioinformatics, we identified two characteristic GC-rich motifs that are located proximal to the transcription start site (TSS) of MGARP, and each contains two Specificity protein 1 (Sp1) binding elements. We then determined that the −3 kb proximal MGARP promoter is activated in a Sp1-dependent manner using reporter assays and knockdown of Sp1 led to decreased expression of endogenous MGARP messages. We also demonstrated that one of the two GC-rich motifs, GC-Box1, harbors prominent promoter activity mediated by Sp1, and that it requires both GC boxes for full transcriptional activation. These findings suggest a dominant role for these GC boxes and Sp1 in activating the MGARP promoter through a synergistic mechanism. Consistently, the results of an Electrophoretic Mobility Gel Shift Assay (EMSA) and Chromatin Immunoprecipitation (ChIP) confirmed that Sp1 specifically interacts with the GC-rich region. We further found that estrogen receptor α (ERα), a known Sp1 co-activator, could potentiate GC-boxes containing MGARP promoter activity and this effect is mediated by Sp1. Knockdown of Sp1 significantly diminished the MGARP promoter transactivation and the expression of endogenous MGARP mediated by both Sp1 and ERα.Conclusions/SignificanceThe present study identified a proximal core sequence in the MGARP promoter that is composed of two enriched Sp1 binding motifs and established Sp1 as one major MGARP transactivator whose functions are synergistic with ERα, providing a novel understanding of the mechanisms of MGARP gene transcriptional regulation.
Highlights
Mitochondria-localized glutamic acid-rich protein (MGARP) was first identified in the ovary as the ovary-specific acidic protein (OSAP) [1] and, thereafter, it was identified in the cornea [2] and retina [3]
We compared the 23 kb promoter regions of three species and, as shown in Figure 1A, there are some key features of the promoter those are highly conserved among Homo sapiens (Human), Pan troglodytes (Chimpanzee) and Macaca mulatta (Macaque)
Transcription is regulated by chromatin-interacting factors, which bind to their specific DNA recognition sequences [29]
Summary
Mitochondria-localized glutamic acid-rich protein (MGARP) was first identified in the ovary as the ovary-specific acidic protein (OSAP) [1] and, thereafter, it was identified in the cornea [2] and retina [3]. Our previous studies have demonstrated that MGARP is highly expressed in the inner segment of the photoreceptor (IS), outer plexiform layer (OPL) and ganglion cell layer (GCL) of the retina, which are enriched with mitochondria [3]. Additional studies have indicated that MGARP is involved in steroidogenesis through its ability to maintain mitochondrial abundance and morphology, and importantly, it is highly expressed in the organs involved in steroidogenesis, such as the ovary, testis, adrenal gland and brain [4,5]. Mitochondria-localized glutamic acid-rich protein (MGARP) is a novel mitochondrial transmembrane protein expressed mainly in steroidogenic tissues and in the visual system. Previous studies showed that MGARP functions in hormone biosynthesis and its expression is modulated by the HPG axis
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