Amino acid deprivation and endoplasmic reticulum stress induce expression of multiple activating transcription factor-3 mRNA species that, when overexpressed in HepG2 cells, modulate transcription by the human asparagine synthetase promoter

Transcription from the ASNS (asparagine synthetase) 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 ASNS promoter, referred to as nutrient-sensing response element-1 and -2. Chromatin immunoprecipitation analysis provides the first in vivo evidence for activating transcription factor (ATF)-3 binding to the proximal ASNS promoter containing the nutrient-sensing response element-1 sequence. Overexpression of the full-length ATF3 protein caused a concentration-dependent biphasic response in ASNS promoter-driven transcription. Both amino acid limitation and activation of the endoplasmic reticulum stress response by glucose deprivation caused an increase in ATF3 mRNA content. However, reverse transcriptase-PCR analysis revealed that the increase in the ATF3 mRNA species detected by Northern analysis actually encoded both full-length ATF3 and two predicted truncated ATF3 isoforms (ATF3ΔZip2c and ATF3ΔZip3). Based on sequence analysis, one of the predicted truncated proteins (ATF3ΔZip3) is likely incapable of binding DNA; and yet, exogenous expression of the cDNA enhanced starvation-induced or ATF4-activated ASNS transcription, possibly by sequestering corepressor proteins. Collectively, the results provide evidence for a potential role of multiple predicted ATF3 isoforms in the transcriptional regulation of the ASNS gene in response to nutrient deprivation.