Department of Biomolecular Chemistry, Radboud University Nijmegen, Nijmegen, The Netherlands.
Cell Stress Chaperones. 2012 Nov;17(6):743-55. doi: 10.1007/s12192-012-0347-1. Epub 2012 Jul 14.
Mammalian cells respond to a lack of amino acids by activating a transcriptional program with the transcription factor ATF4 as one of the main actors. When cells are faced with cytoplasmic proteotoxic stress, a quite different transcriptional response is mounted, the heat shock response, which is mediated by HSF1. Here, we show that amino acid deprivation results in the inactivation of HSF1. In amino acid deprived cells, active HSF1 loses its DNA binding activity as demonstrated by EMSA and ChIP. A sharp decrease in the transcript level of HSF1 target genes such as HSPA1A (Hsp70), DNAJB1 (Hsp40), and HSP90AA1 is also seen. HSPA1A mRNA, but not DNAJB1 mRNA, was also destabilized. In cells cultured with limiting leucine, HSF1 activity also declined. Lack of amino acids thus could lead to a lower chaperoning capacity and cellular frailty. We show that the nutrient sensing response unit of the ASNS gene contains an HSF1 binding site, but we could not detect binding of HSF1 to this site in vivo. Expression of either an HSF1 mutant lacking the activation domain (HSF379) or an HSF1 mutant unable to bind DNA (K80Q) had only a minor effect on the transcript levels of amino acid deprivation responsive genes.
哺乳动物细胞在缺乏氨基酸时会激活一个转录程序，其中转录因子 ATF4 是主要的执行者之一。当细胞面临细胞质蛋白毒性应激时，会引发截然不同的转录反应，即热休克反应，该反应由 HSF1 介导。在这里，我们发现氨基酸剥夺会导致 HSF1 的失活。在缺乏氨基酸的细胞中，如 EMSA 和 ChIP 所示，活性 HSF1 失去了其 DNA 结合活性。HSF1 靶基因如 HSPA1A（Hsp70）、DNAJB1（Hsp40）和 HSP90AA1 的转录水平也急剧下降。还观察到 HSPA1A mRNA 而非 DNAJB1 mRNA 的不稳定性。在培养有限制性亮氨酸的细胞中，HSF1 活性也下降。因此，缺乏氨基酸可能导致较低的伴侣蛋白能力和细胞脆弱性。我们发现 ASNS 基因的营养感应反应单元包含 HSF1 结合位点，但我们无法在体内检测到 HSF1 与该位点的结合。表达缺乏激活域的 HSF1 突变体（HSF379）或无法结合 DNA 的 HSF1 突变体（K80Q）仅对氨基酸剥夺反应基因的转录水平产生轻微影响。
