Received: 18 October 2018 | Revised: 16 February 2019 | Accepted: 20 March 2019 DOI: 10.1002/jezb.22855 RESEARCH ARTICLE Tuftelinʼs involvement in embryonic development Dekel Shilo 1 | Anat Blumenfeld 1 | Amir Haze 1 | Shay Sharon 1 | Koby Goren 1 | Salem Hanhan 1 | Yael Gruenbaum‐Cohen 1 | Asher Ornoy 2 | Dan Deutsch 1 1 Dental Research Laboratory, Faculty of Dental Medicine, Institute of Dental Sciences, The Hebrew University of Jerusalem– Hadassah, Jerusalem, Israel 2 Laboratory of Teratology, Department of Medical Neurobiology, The Hebrew University of Jerusalem–Hadassah, Jerusalem, Israel Correspondence Dekel Shilo, Rambam Health Care Campus, 8 Ha’Aliyah Street, Haifa 35254, Israel. Email: dekelshi@yahoo.com Abstract Little is known about tuftelin expression in the developing embryo, previously it was thought to play a role in tooth enamel mineralization. In this study we show tuftelinʼs spatio‐temporal expression in mineralizing and nonmineralizing tissues of the craniofacial complex in the developing mouse embryo. Embryos aged E10.5-E18.5 and newborns aged P3 were used in this study. Polymerase chain reaction (PCR), Real‐time PCR, sequencing, and in‐situ hybridization were used to detect and quantify messenger RNA (mRNA) expression in different developmental stages. We applied indirect immunohistochemistry and western‐blot analyses to investigate protein expression. Two tuftelin mRNA transcripts and a single 64KDa protein were detected throughout embryonic development. Tuftelin was detected in tissues which develop from different embryonic origins; ectoderm, ectomesenchyme, and meso- derm. Tuftelin mRNA and protein were expressed already at E10.5, before the initiation of tooth formation and earlier than previously described. The expression pattern of tuftelin mRNA and protein exhibits dynamic spatio‐temporal changes in various tissues. Tuftelin is expressed in neuronal tissues, thus fitting with its described correlation to nerve growth factor. A shift between cytoplasmatic and perinuclear/nuclear expression implies a possible role in regulation of transcrip- tion. Recent studies showed tuftelin is induced under hypoxic conditions in‐vitro and in‐vivo, through the hypoxia‐inducible factor 1‐α pathway. These results led to the hypothesis that tuftelin is involved in adaptation to hypoxic conditions. The fact that much of mammalian embryogenesis occurs at O 2 concentrations of 1–5%, raises the possibility that tuftelin expression throughout development is due to its role in the adaptive mechanisms in response to hypoxia. KEYWORDS CNS, craniofacial complex, embryonic development, mouse, tuftelin 1 | INTRODUCTION Tuftelin is an acidic protein, first discovered, mapped, and cloned from a complementary DNA (cDNA) library of ameloblasts (Deutsch et al., 1991). It is expressed by epithelial cells at very early stages of odontogenesis (Deutsch et al., 1998; Deutsch et al., 2002; Zeichner‐ David et al., 1997), when dentin or enamel is not yet formed; tuftelin mRNA was detected at mouse embryonic Day 13 (E13), while the protein was detected at E17 (Zeichner‐David et al., 1997). At very early stages of odontogenesis, the dental papilla mesenchyme and the preodontoblasts that eventually form the underlying dentin express tuftelin in a transient manner (Diekwisch, Ware, Fincham, & Zeichner‐David, 1997; Zeichner‐David et al., 1997). Hence, it has been suggested that tuftelin might be involved in mesenchyme‐ ectoderm interactions during tooth development (Deutsch et al., 1998; Deutsch et al., 2002; Zeichner‐David et al., 1997). Later, when J Exp Zool (Mol Dev Evol). 2019;1–11. wileyonlinelibrary.com/journal/jezb © 2019 Wiley Periodicals, Inc. | 1