Mechanisms of transcriptional regulation of CAT-1 gene expression by endoplasmic reticulum (ER) stress

As a member of cationic amino acid transporter (CAT) family proteins, CAT-1 mediates the bidirectional transport of essential amino acids arginine and lysine by facilitated diffusion. Cat-1 gene expression is modulated by stimuli including endoplasmic reticulum (ER) stress, availability of nutrients, growth factors, and hormones. In mammalian cells, ER stress triggers 3 distinct signaling pathways initiated by the 3 ER membrane-associated sensor proteins PERK, IRE1, and ATF6. It is shown here that cat-1 mRNA level is induced to about 5 fold at the highest level in C6 cells during ER stress. Transcriptional induction of cat-1 by ER stress is mediated by two transcription factors ATF4 and XBP1s. ATF4 interacts with the cis-acting element AARE and XBP1s interacts with the putative cis-acting element ERSE-II to induce cat-1 gene transcription at early stage of ER stress. The sequential binding of XBP1s after ATF4 to the cat-1 promoter during ER stress may be responsible for sustained induction of cat-1 mRNA levels during ER stress. eIF2alpha phosphorylation is required for induction of cat-1 transcription by ER stress due to induction of ATF4 entirely and induction of XBP1s largely requires eIF2alpha phosphorylation. Because eIF2alpha phosphorylation is required for ATF4 mRNA translation as described in the literature and sufficient XBP1 mRNA splicing showing here. This indicates that there is a crosstalk between the PERK-eIF2alpha-ATF4 and the IRE1-XBP1 pathways. At the late stage of ER stress, C/EBPbeta LIP isoform down-regulates cat-1 gene transcription by specifically suppressing the induction mediated by AARE via binding of ATF4. Transcriptional induction of other two ATF4 induced genes SNAT2 and AS is also suppressed by C/EBPbeta LIP during ER stress. The down-regulation of C/EBPbeta LIP level at the early stage of ER stress is mediated by proteasome-mediated degradation and eIF2alpha phosphorylation. The dramatic increase of C/EBPbeta LIP at late staged of ER stress is caused by increased mRNA translation and stability of the protein. Physiologically, LIP induction may help cells to survive during ER stress. In our knowledge, the crosstalk between the PERK-eIF2alpha-ATF4 and the IRE1-XBP1 pathways of the 3 UPR signaling pathways is being demonstrated here at the first time.