Analysis of glutaminase activity and RNA expression in preimplantation mouse embryos.
Glutamine is utilized as an energy substrate in preimplantation mouse embryos. Glutaminase is the enzyme responsible for the conversion of glutamine to glutamic acid, which then enters the trichloro acetic acid (TCA) cycle as alpha-ketoglutarate. Glutaminase enzyme activity was assessed in preimplantation embryos that developed in vivo, and glutaminase RNA expression was examined in embryos that developed in vivo or were cultured in CZB medium to various preimplantation stages between 1-cell and blastocyst. Glutaminase activity in 1-8-cell-stage mouse embryos that developed in vivo ranged from 0.009-0.01 U/mg protein (2.39-2.95 x 10(-7) U per embryo) and increased 3-4 fold to 0.034 U/mg protein (8.13 x 10(-7) U per embryo) at the blastocyst stage. Relative stage-specific expression of glutaminase RNA was assessed by reverse transcription polymerase chain reaction (RT-PCR) in embryos that developed both in vivo and in CZB culture. In vivo, glutaminase RNA was expressed at the 1-cell stage, declined to 23% of 1-cell levels at the early 2-cell stage, and reaccumulated from late 2-cell through blastocyst stage, where it reached a high of 204% of 1-cell levels. CZB-cultured embryos exhibited a similar pattern of developmental RNA expression, declining to 30% of 1-cell levels at the early 2-cell stage, and increasing RNA expression at the blastocyst stage to 191% of the 1-cell level.
Chatot
C L
CL
Department of Biology, Ball State University, Muncie, Indiana, USA.
Lawry
J R
JR
Germain
B
B
Ziomek
C A
CA
D000818:Animals; D015139:Blotting, Southern; D002451:Cell Compartmentation; D046508:Culture Techniques; D004622:Embryo, Mammalian / Q000201:enzymology*; D047108:Embryonic Development; D005260:Female; D005972:Glutaminase / Q000235:genetics / Q000378:metabolism*; D051379:Mice; D016133:Polymerase Chain Reaction; D011247:Pregnancy; D012333:RNA, Messenger / Q000378:metabolism*
D012333:RNA, Messenger; D005972:Glutaminase