PMID- 15115889
OWN - NLM
STAT- MEDLINE
DCOM- 20050204
LR  - 20151119
IS  - 1096-6080 (Print)
IS  - 1096-0929 (Linking)
VI  - 81
IP  - 2
DP  - 2004 Oct
TI  - Spatial activities and induction of glutamate-cysteine ligase (GCL) in the 
      postimplantation rat embryo and visceral yolk sac.
PG  - 371-8
AB  - Glutathione (GSH) synthesis is differentially regulated in the embryo and 
      visceral yolk sac (VYS) of the developing rat conceptus. The innate capacity to 
      respond to environmental insult and chemical exposure by inducing de novo GSH 
      synthesis may help to determine overall cell sensitivity and/or resistance to 
      chemically induced malformation. Specific activities of glutamate-cysteine ligase 
      (GCL), the rate limiting enzyme in GSH synthesis, were determined by measuring 
      the formation of gamma-glutamylcysteine (GC) in homogenates prepared from rat 
      embryos and VYSs. GC formation increased linearly with time and with relative 
      protein concentration. Specific activities were found to be 60.5 +/- 3.2 and 
      118.9 +/- 4.2 pmol GC/mg protein/min in the gestational day (GD) 10 embryo and 
      VYS, respectively, and 22.7 +/- 0.4 and 71.3 +/- 0.6 pmoles GC/mg protein/min in 
      the respective GD 11 embryo and VYS. Apparent kinetic constants determined from 
      embryo and VYS homogenates gave respective apparent K(m) values for glutamate of 
      0.75 and 1.38 mM and for cysteine 0.03 mM in both tissues. Apparent V(max) values 
      were higher in the VYS in each case, corresponding with a lower apparent K(m) and 
      higher GCL activity. GCL specific activities increased significantly following a 
      24 h in vitro exposure to diethyl maleate (DEM) and diamide, but remained 
      unchanged following exposure to prostaglandin A(2) (PGA(2)) and t-butylated 
      hydroxytoluene (BHT). Basal expression of GCL catalytic subunit (GCL(C)) and 
      regulatory subunit (GCL(R)) was 59- and 25-fold higher in VYS, respectively, 
      compared to the embryo. Quantitative real-time fluorescence reverse transcriptase 
      polymerase chain reaction (RT-PCR) showed that following DEM and diamide 
      treatment, GCL(C) expression increased up to 19-fold in embryonic tissues but was 
      not induced in the VYS. Only DEM increased the expression of the light/regulatory 
      subunit GCL(R) in the embryo (8-fold). Densitometry of immunoblots revealed 
      approximately 75% more GCL(C) in the VYS than in the embryo. Following 
      treatments, a marked increase was induced in embryonic GCL(C) content with both 
      DEM (85%) and diamide (19%), but in the VYS, only DEM caused an increase in 
      GCL(C) protein (38%).
FAU - Hansen, Jason M
AU  - Hansen JM
AD  - Department of Biochemistry, Emory University, Atlanta, Georgia 30322, USA.
FAU - Lee, Eunyong
AU  - Lee E
FAU - Harris, Craig
AU  - Harris C
LA  - eng
GR  - DK11457/DK/NIDDK NIH HHS/United States
GR  - ES05235/ES/NIEHS NIH HHS/United States
GR  - ES11457/ES/NIEHS NIH HHS/United States
PT  - Journal Article
PT  - Research Support, U.S. Gov't, P.H.S.
DEP - 20040428
PL  - United States
TA  - Toxicol Sci
JT  - Toxicological sciences : an official journal of the Society of Toxicology
JID - 9805461
RN  - 0 (DNA Primers)
RN  - 0 (Dipeptides)
RN  - 0 (RNA, Messenger)
RN  - EC 6.3.2.2 (Glutamate-Cysteine Ligase)
RN  - GAN16C9B8O (Glutathione)
RN  - M984VJS48P (gamma-glutamylcysteine)
SB  - IM
MH  - Animals
MH  - Blastocyst/*enzymology/pathology
MH  - Chromatography, High Pressure Liquid
MH  - DNA Primers
MH  - Dipeptides/metabolism
MH  - Enzyme Induction/drug effects
MH  - Female
MH  - Glutamate-Cysteine Ligase/*biosynthesis/genetics
MH  - Glutathione/metabolism
MH  - Immunoblotting
MH  - Oxidative Stress/drug effects/physiology
MH  - Pregnancy
MH  - RNA, Messenger/biosynthesis/genetics
MH  - Rats
MH  - Rats, Sprague-Dawley
MH  - Reverse Transcriptase Polymerase Chain Reaction
MH  - Yolk Sac/*enzymology/pathology
EDAT- 2004/04/30 05:00
MHDA- 2005/02/05 09:00
CRDT- 2004/04/30 05:00
PHST- 2004/04/30 05:00 [pubmed]
PHST- 2005/02/05 09:00 [medline]
PHST- 2004/04/30 05:00 [entrez]
AID - kfh154 [pii]
AID - 10.1093/toxsci/kfh154 [doi]
PST - ppublish
SO  - Toxicol Sci. 2004 Oct;81(2):371-8. doi: 10.1093/toxsci/kfh154. Epub 2004 Apr 28.