Original Article Activin-like signaling activates Notch signaling during mesodermal induction TAKANORI ABE 1 , MIHO FURUE 2 , YASUFUMI MYOISHI 3 , TETSUJI OKAMOTO 3 , AKIKO KONDOW 4 and MAKOTO ASASHIMA* ,1, 4, 5 1 Department of Biological Science, Graduate School of Science, The University of Tokyo, Japan 2 Department of Biochemistry and Molecu- lar Biology, Kanagawa Dental College, Yokosuka, Japan 3 Department of Molecular Oral Medicine and Maxillofacial Surgery, Division of Frontier Medical Science, Graduate School of Biomedical Sciences, Hiroshima University, Japan, 4 Department of Life Sciences (Biology), Graduate School of Arts and Sciences, Japan and 5 SORST/Japan Science and Technology Corporation, The University of Tokyo, Japan ABSTRACT Both activin-like signaling and Notch signaling play fundamental roles during early development. Activin-like signaling is involved in mesodermal induction and can induce a broad range of mesodermal genes and tissues from prospective ectodermal cells (animal caps). On the other hand, Notch signaling plays important roles when multipotent precursor cells achieve a specific cell fate. However, the relationship between these two signal pathways is not well understood. Here, we show that activin A induces Delta-1, Delta-2 and Notch expression and then activates Notch signaling in animal caps. Also, in vivo, ectopic activin-like signaling induced the ectopic expression of Delta-1 and Delta-2, whereas inhibition of activin-like signaling abolished the expression of Delta-1 and Delta-2. Furthermore, we show that MyoD, which is myogenic gene induced by activin A, can induce Delta-1 expression. However, MyoD had no effect on Notch expression, and inhibited Delta-2 expression. These results indicated that activin A induces Delta- 1, Delta-2 and Notch by different cascades. We conclude that Notch signaling is activated when activin-like signaling induces various tissues from homogenous undifferentiated cells. KEY WORDS: Xenopus laevis, activin, Notch signaling, MyoD, mesoderm induction Int. J. Dev. Biol. 48: 327-332 (2004) 0214-6282/2004/$25.00 © UBC Press Printed in Spain www.ijdb.ehu.es *Address correspondence to: Dr. Makoto Asashima. Department of Life Sciences (Biology), Graduate School of Arts and Sciences, The University of Tokyo, 3- 8-1 Komaba, Meguro-ku, Tokyo, 153-8902, Japan. Fax: +81-3-5454-4330. e-mail: asashi@bio.c.u-tokyo.ac.jp Abbreviations used in this paper: ALK4 dn, dominant negative activin receptor- like kinase 4; AR1, activin type II receptor; DSL, Delta, Serrate and Lag2 ligand family; ODC, ornithine decarboxylase. Introduction Activin A, a member of the TGF-β family, which has a strong mesoderm-inducing activity (Asashima et al., 1990), can induce a broad range of mesodermal genes and tissues from amphib- ian prospective ectodermal cells (animal caps) in a concentra- tion-dependent manner (Ariizumi et al., 1991a; Ariizumi et al., 1991b; Green and Smith, 1990). At low concentrations, activin A induces the formation of ventral and posterior mesodermal tissues such as blood cells, coelomic epithelium and mesen- chyme. At intermediate concentrations, activin A induces muscle, and at high concentrations, it induces dorsal and anterior mesoderm tissues such as notochord, and yolk-loaded endo- derm. In addition, activin A in combination with retinoic acid induces pronephros (Brennan et al., 1999; Moriya et al., 1993; Osafune et al., 2002) and anterior endoderm tissues such as pancreas (Moriya et al., 2000), liver, and intestine. Further- more, activin A can also induce beating heart muscle (Ariizumi et al., 1996) and jaw cartilage (Furue et al., 2002) by the sandwich-culture method. Notch encodes a large transmem- brane protein that serves as a receptor for the Delta, Serrate, and Lag-2 (DSL) family of ligands. Binding of a DSL ligand to the extracellular domain of Notch causes cleavage of the receptor's intracellular domain, which is then released from the cell mem- brane (De Strooper et al., 1999; Mumm et al., 2000). The intracellular domain of Notch translocates into the nucleus, where it can interact with members of the CBF-1, Suppressor of Hairless, Lag-1 (CSL) family. The complex of DSL and the intracellular domain of Notch subsequently activates the ex- pression of target genes, such as ESR, Hairy, and Hey (Bailey and Posakony, 1995; Lecourtois and Schweisguth, 1995; Leimeister et al., 1999; Tannahill et al., 1995). Notch signaling is involved in multiple developmental processes and controls the cell-fate decision by regulating local cell-cell communica- tion (Artavanis-Tsakonas et al., 1999). Notch signaling main-