Human SPRY2 peptide

Tooth agenesis affects 20% of the world population, and maxillary lateral incisors agenesis (MLIA) is one of the most frequent subtypes, characterized by the absence of formation of deciduous or permanent lateral incisors. Odontogenesis is a complex mechanism regulated by sequential and reciprocal epithelial-mesenchymal interactions, controlled by activators and inhibitors involved in several pathways. Disturbances in these signaling cascades can lead to abnormalities in odontogenesis, resulting in alterations in the formation of the normal teeth number. Our aim was to study a large number of genes encoding either transcription factors or key components in signaling pathways shown to be involved in tooth odontogenesis. We selected 8 genes-MSX1, PAX9, AXIN2, EDA, SPRY2, TGFA, SPRY4, and WNT10A-and performed one of the largest case-control studies taking into account the number of genes and variants assessed, aiming at the identification of MLIA susceptibility factors. We show the involvement of PAX9, EDA, SPRY2, SPRY4, and WNT10A as risk factors for MLIA. Additionally, we uncovered 3 strong synergistic interactions between MLIA liability and MSX1-TGFA, AXIN2-TGFA, and SPRY2-SPRY4 gene pairs. We report the first evidence of the involvement of sprouty genes in MLIA susceptibility. This large study results in a better understanding of the genetic components and mechanisms underlying this trait.

Receptor Tyrosine Kinase (RTK) signaling plays a major role in tumorigenesis and normal development. Sprouty2 (Spry2) attenuates RTK signaling and inhibits processes such as angiogenesis, cell proliferation, migration and survival, which are all upregulated in tumors. Indeed in cancers of the liver, lung, prostate and breast, Spry2 protein levels are markedly decreased correlating with poor patient prognosis and shorter survival. Thus, it is important to understand how expression of Spry2 is regulated. While prior studies have focused on the post-translation regulation of Spry2, very few studies have focused on the transcriptional regulation of SPRY2 gene. Here, we demonstrate that in the human hepatoma cell line, Hep3B, the transcription of SPRY2 is inhibited by the transcription regulating hypoxia inducible factors (HIFs). HIFs are composed of an oxygen regulated alpha subunit (HIF1 alpha or HIF2 alpha) and a beta subunit (HIF1 beta). Intriguingly, silencing of HIF1 alpha and HIF2 alpha elevates SPRY2 mRNA and protein levels suggesting HIFs reduce the transcription of the SPRY2 promoter. In silico analysis identified ten hypoxia response elements (HREs) in the proximal promoter and first intron of SPRY2. Using chromatin immunoprecipitation (ChIP), we show that HIF1 alpha/2 alpha bind near the putative HREs in the proximal promoter and intron of SPRY2. Our studies demonstrated that not only is the SPRY2 promoter methylated, but silencing HIF1 alpha/2 alpha reduced the methylation. ChIP assays also showed DNA methyltransferase1 (DNMT1) binding to the proximal promoter and first intron of SPRY2 and silencing HIF1 alpha/2 alpha decreased this association. Additionally, silencing of DNMT1 mimicked the HIF1 alpha/2 alpha silencing-mediated increase in SPRY2 mRNA and protein. While simultaneous silencing of HIF1 alpha/2 alpha and DNMT1 increased SPRY2 mRNA a little more, the increase was not additive suggesting a common mechanism by which DNMT1 and HIF1 alpha/2 alpha regulate SPRY2 transcription. Together these data suggest that the transcription of SPRY2 is inhibited by HIFs, in part, via DNMT1-mediated methylation.