ATF4 is a mediator of the nutrient-sensing response pathway that activates the human asparagine synthetase gene.
Transcription from the asparagine synthetase (A.S.) gene is increased in response to either amino acid (amino acid response) or glucose (endoplasmic reticulum stress response) deprivation. These two independent pathways converge on the same set of genomic cis-elements within the A.S. promoter referred to as nutrient-sensing response elements (NSRE) 1 and 2, both of which are necessary for gene activation. The NSRE-1 sequence was used to screen ATF/CREB family members by electrophoresis mobility shift assays and supershift by specific antibodies. The results indicated that ATF4 binds to the NSRE-1 sequence and that the amount of the ATF4 complex was increased when extracts from amino acid-deprived or glucose-deprived cells were tested. Using electrophoresis mobility shift assay experiments and a probe that contained both NSRE-1 and NSRE-2, mutation of the NSRE-1 sequence completely prevented formation of the ATF4-containing complexes, whereas mutation of the NSRE-2 sequence did not. Overexpression of ATF4 increased A.S. promoter-driven transcription, whereas an inhibitory dominant negative ATF4 mutant blocked both basal and starvation-enhanced transcription. Collectively, the results provide both in vitro and in vivo evidence for a role of ATF4 in the transcriptional activation of the A.S. gene in response to nutrient deprivation.
Siu
Fai
F
Department of Biochemistry and Molecular Biology, Center for Mammalian Genetics, and Center for Nutritional Sciences, University of Florida College of Medicine, Gainesville, Florida 32610-0245, USA.
Bain
Perry J
PJ
LeBlanc-Chaffin
Rene
R
Chen
Hong
H
Kilberg
Michael S
MS
D051701:Activating Transcription Factor 4; D001217:Aspartate-Ammonia Ligase / Q000096:biosynthesis / Q000235:genetics*; D001483:Base Sequence; D004789:Enzyme Activation; D015971:Gene Expression Regulation, Enzymologic; D006639:Histidine; D006801:Humans; D007700:Kinetics; D016350:Leucine Zippers; D008969:Molecular Sequence Data; D011401:Promoter Regions, Genetic; D012333:RNA, Messenger / Q000235:genetics; D011994:Recombinant Proteins / Q000378:metabolism; D017384:Sequence Deletion; D014157:Transcription Factors / Q000378:metabolism*; D015533:Transcriptional Activation; D014162:Transfection; D014407:Tumor Cells, Cultured
C497109:ATF4 protein, human; D012333:RNA, Messenger; D011994:Recombinant Proteins; D014157:Transcription Factors; D051701:Activating Transcription Factor 4; D006639:Histidine; D001217:Aspartate-Ammonia Ligase