Granzyme G is expressed in the two-cell stage mouse embryo and is required for the maternal-zygotic transition.
BACKGROUND
Detailed knowledge of the molecular and cellular mechanisms that direct spatial and temporal gene expression in pre-implantation embryos is critical for understanding the control of the maternal-zygotic transition and cell differentiation in early embryonic development. In this study, twenty-three clones, expressed at different stages of early mouse development, were identified using differential display reverse transcription polymerase chain reaction (DDRT-PCR). One of these clones, which is expressed in 2-cell stage embryos at 48 hr post-hCG injection, shows a perfect sequence homology to the gene encoding the granzyme G protein. The granzyme family members are serine proteases that are present in the secretory granules of cytolytic T lymphocytes. However, the pattern of granzyme G expression and its function in early mouse embryos are entirely unknown.


RESULTS
Upon the introduction of an antisense morpholino (2 mM) against granzyme G to knock-down endogenous gene function, all embryos were arrested at the 2- to 4-cell stages of egg cleavage, and the de novo synthesis of zygotic RNAs was decreased. The embryonic survival rate was dramatically decreased at the late 2-cell stage when serine protease-specific inhibitors, 0.1 mM 3,4-dichloroisocoumarin (3,4-DCI), and 2 mM phenyl methanesulphonyl fluoride (PMSF), were added to the in vitro embryonic culture medium. Survival was not affected by the addition of 0.5 mM EDTA, a metalloproteinase inhibitor.


CONCLUSIONS
We characterized for the first time the expression and function of granzyme G during early stage embryogenesis. Our data suggest that granzyme G is an important factor in early mouse embryonic development and may play a novel role in the elimination of maternal proteins and the triggering of zygotic gene expression during the maternal-zygotic transition.
Tsai
Tung-Chou
TC
Department of Life Sciences, National Chung Hsing University, Taichung 402, Taiwan.
Lin
William
W
Yang
Shang-Hsun
SH
Cheng
Winston T K
WT
Cheng
En-Hui
EH
Lee
Maw-Sheng
MS
Chong
Kowit-Yu
KY
Chen
Chuan-Mu
CM
D000818:Animals; D004622:Embryo, Mammalian / Q000378:metabolism; D047108:Embryonic Development; D005260:Female; D018507:Gene Expression Regulation, Developmental; D055785:Gene Knockdown Techniques; D053804:Granzymes / Q000037:antagonists & inhibitors / Q000235:genetics* / Q000378:metabolism*; D051379:Mice; D016372:RNA, Antisense / Q000235:genetics; D020133:Reverse Transcriptase Polymerase Chain Reaction / Q000379:methods; D015842:Serine Proteinase Inhibitors / Q000378:metabolism; D015053:Zygote / Q000378:metabolism*
D016372:RNA, Antisense; D015842:Serine Proteinase Inhibitors; D053804:Granzymes; C066158:Gzmg protein, mouse