Identification of ZAG1, a novel protein expressed in mouse preimplantation, and its putative roles in zygotic genome activation.
We isolated a mouse cDNA, zag1 (zygotic gene activation-associated gene 1), that has an open reading frame of 1,728-bp encoding a protein of 66.2 kDa including both a bipartite nuclear targeting sequence and a P-loop motif containing nucleoside triphosphate hydrolase motifs. Northern blot analysis of mouse tissues showed that zag1 was widely expressed but was especially prominent in the ovary and testis. RT-PCR analysis of in vitro fertilized embryos showed that the abundance of zag1 transcripts in oocytes decreased after fertilization, and zag1 mRNA was detected at 15 h post insemination (hpi) in fertilized embryos indicating that the gene was expressed at the start of zygotic gene activation at the mouse 1-cell stage. The nuclear-localization of ZAG1 protein in mouse preimplantation embryos at 15 hpi was confirmed by both subcellular analysis of enhanced green fluorescent protein (EGFP)-tagged ZAG1 and immunocytochemical analysis with anti-ZAG1 antibody. Subsequently, using yeast two-hybrid screening, we identified U2 small nuclear ribonucleoprotein B (U2B"), which is associated with pre-mRNA splicing, as a putative interacting partner of ZAG1 protein. Furthermore, knockdown of zag1 expression by an antisense DNA plasmid induced arrest and/or delay of embryonic development in injected 1-cell embryos. These results suggest that ZAG1 may be closely associated with zygotic gene expression in mouse preimplantation embryos.
Matsuoka
Toshiki
T
Division of Biological Science, Graduate School of Biology-Oriented Science and Technology, Kinki University, Wakayama, Japan.
Sato
Manabu
M
Tokoro
Mikiko
M
Shin
Seung-Wook
SW
Uenoyama
Atsuto
A
Ito
Kazunari
K
Hitomi
Syuji
S
Amano
Tomoko
T
Anzai
Masayuki
M
Kato
Hiromi
H
Mitani
Tasuku
T
Saeki
Kazuhiro
K
Hosoi
Yoshihiko
Y
Iritani
Akira
A
Matsumoto
Kazuya
K
D000595:Amino Acid Sequence; D000818:Animals; D001755:Blastocyst / Q000502:physiology*; D005260:Female; D018507:Gene Expression Regulation, Developmental / Q000235:genetics*; D016678:Genome; D008297:Male; D051379:Mice; D008813:Mice, Inbred ICR; D008969:Molecular Sequence Data; D009687:Nuclear Proteins / Q000235:genetics* / Q000502:physiology*; D009865:Oocytes / Q000502:physiology; D010053:Ovary / Q000502:physiology; D011247:Pregnancy; D013737:Testis / Q000502:physiology; D015533:Transcriptional Activation; D020798:Two-Hybrid System Techniques; D015003:Yeasts; D015053:Zygote / Q000502:physiology*
D009687:Nuclear Proteins; C531060:Zag1 protein, mouse