An essential role for REV3 in mammalian cell survival: absence of REV3 induces p53-independent embryonic death Jiyang O-Wang, a, * Kagemasa Kajiwara, b Kiyoko Kawamura, a Minoru Kimura, c Hiro Miyagishima, d Haruhiko Koseki, d and Masatoshi Tagawa a a Division of Pathology, Chiba Cancer Center Research Institute, 666-2 Nitona, Chuo-ku, Chiba 260-8717, Japan b Institute of Medical Sciences, School of Medicine, Tokai University, Isehara 259-1193, Japan c Department of Molecular Life Science, School of Medicine, Tokai University, Isehara 259-1193, Japan d Department of Molecular Embryology, Graduate School of Medicine, Chiba University, 1-8-1 Inohana, Chuo-ku, Chiba 260-8670, Japan Received 8 April 2002 Abstract The REV3 gene of budding yeast encodes the catalytic subunit of DNA polymerase f that carries out translesion DNA synthesis. While REV3-null yeast mutants are viable and exhibit normal growth, Rev3-deficient mice die around midgestation of embryo- genesis, which is accompanied by massive apoptosis of cells within the embryo proper. We have investigated whether REV3 is required for the survival of mouse cells and whether the embryonic lethality caused by REV3 deficiency can be rescued by intro- duction of a Rev3 transgene or by inactivation of p53, the cellular gatekeeper that regulates DNA damage-induced apoptosis. We show that Rev3 = blastocysts were unable to survive and grow in culture but expression of a Rev3 transgene restored their out- growth. Moreover, Rev3 transgene expression suppressed the apoptosis in E7.5 Rev3 = embryos. The Rev3 = embryonic lethality, however, was not rescued by either Rev3 transgene expression or p53 deficiency. These results reveal an essential role for REV3 in the survival and growth of mammalian cells and suggest that Rev3 = embryonic death occurs in a p53-independent pathway. Ó 2002 Elsevier Science (USA). All rights reserved. Keywords: REV3; Translesion DNA synthesis; Apoptosis; p53 DNA damage is continually induced by both exoge- nous sources such as UV light and chemical agent and endogenous sources such as oxygen-free radicals. While cells possess multiple repair pathways to remove various types of nucleotide damage [1], certain lesions may persist during DNA synthesis and block the replication fork [2,3]. To overcome this blockage and enable the completion of DNA replication, both prokaryotic and eukaryotic cells utilize specialized DNA polymerases that are capable of carrying out DNA synthesis across the unrepaired lesions [3,4]. Biochemical analyses re- vealed that these polymerases in general have relatively low fidelity and processivity, and lack 3 0 –5 0 exonuclease activity that can proofread replication errors [5–9]. Al- though error-prone or error-free bypass appears to de- pend on the type of lesions and the sequence context, these polymerases often incorporate incorrect nucleo- tides during bypass of damage and are implicated in mutagenesis. Their biological functions in mammalian cells, however, are still poorly understood. The REV3 gene of budding yeast, Saccharomyces cerevisiae, encodes the catalytic subunit of DNA poly- merase f [10–12]. REV3-null yeast mutants are viable and display normal growth, but have greatly reduced rates of both spontaneous and DNA damage-induced mutations [11]. Human and mouse homologues of the yeast REV3 gene have recently been described [13–17]. The mammalian REV3 has about twice the size of the yeast counterpart and contains regions with no signifi- cant homology to the yeast REV3. Inhibition of REV3 expression by an anti-sense RNA reportedly reduced the UV-induced mutagenesis in cultured human cells, sug- gesting that mammalian REV3 may have a function similar to that of yeast REV3 [13]. In contrast to REV3- Biochemical and Biophysical Research Communications 293 (2002) 1132–1137 www.academicpress.com BBRC * Corresponding author. Fax: +81-43-265-4459. E-mail address: oh@chiba-cc.pref.chiba.jp (J. O-Wang). 0006-291X/02/$ - see front matter Ó 2002 Elsevier Science (USA). All rights reserved. PII:S0006-291X(02)00341-8