Int. J. Dev. Biol. 58: 873 - 879 (2014)

https://doi.org/10.1387/ijdb.140193aw

Vol 58, Issue 10-11-12

Special Issue: Developmental Herpetology

In silico identification of the genes for sperm-egg interaction in the internal fertilization of the newt Cynops pyrrhogaster

Published: 2 July 2015

Akihiko Watanabe^1,* and Eriko Takayama-Watanabe²

¹Department of Biology, Faculty of Science and ²Institute of Arts and Sciences, Yamagata University, Kojirakawa, Yamagata, Japan

Abstract

A specific sperm-egg interaction in the oviductal matrix is crucial for internal fertilization of the red-bellied newt, Cynops pyrrhogaster. An understanding of the molecular basis of this interaction is expected to elucidate the evolutionary history of internal fertilization in amphibians. Recently, deep sequencing technology has provided global gene information even in nonmodel animals, allowing us to understand specific features of the molecular mechanisms underlying fertilization in C. pyrrhogaster. In the present study, we screened de novo assembled RNAseq from ovary, testis, and oviduct samples in C. pyrrhogaster and identified the base sequences encoding zona pellucida (ZP) proteins, voltage-dependent Ca²⁺ channels, and cysteine-rich secretory proteins (CRISPs), which respectively are sperm receptors for egg envelopes, major mediators of sperm intracellular signaling, and expected extracellular modulators for sperm function in the female reproductive tract. In the ovary, ZP homologues of all six subgroups were found, including a ZP1 homologue that was newly found in amphibians, a ZP4 homologue, and six ZPC homologues. The unique combination of ZP proteins suggests a new mechanism for sperm binding to egg envelopes in the internal fertilization of C. pyrrhogaster. In the testis, CaV1.1, 1.2, and 3.2, which are L- and T-type voltage-dependent Ca²⁺ channels, were found as potential mediators for the internal fertilization-specific sperm-egg interaction. We also found CRISP 2 in the oviduct, which is speculated to participate in the sperm-egg interaction. These results indicate that the de novo assembled RNAseq is a powerful tool allowing analysis of the specific sperm-egg interactions in C. pyrrhogaster.

Keywords

zp proteins, ca²⁺ channel, crisp, rnaseq, internal fertilization, urodele