Abstract:
Sensing the lowering of endoplasmic reticulum (ER) calcium (Ca2+), STIM1 mediates a ubiquitous Ca2+ influx process called the store-operated Ca2+ entry (SOCE). Dysregulated STIM1 function or abnormal SOCE is strongly associated with autoimmune disorders, atherosclerosis, and various forms of cancers. Therefore, uncovering the molecular intricacies of post-translational modifications, such as oxidation, on STIM1 function is of paramount importance. In a recent proteomic screening, we identified three protein disulfide isomerases (PDIs)-Prolyl 4-hydroxylase subunit beta (P4HB), protein disulfide-isomerase A3 (PDIA3), and thioredoxin domain-containing protein 5 (TXNDC5)-as the ER-luminal interactors of STIM1. Here, we demonstrated that these PDIs dynamically associate with STIM1 and STIM2. The mutation of the two conserved cysteine residues of STIM1 (STIM1-2CA) decreased its Ca2+ affinity both in cellulo and in situ. Knockdown of PDIA3 or P4HB increased the Ca2+ affinity of wild-type STIM1 while showing no impact on the STIM1-2CA mutant, indicating that PDIA3 and P4HB regulate STIM1\'s Ca2+ affinity by acting on ER-luminal cysteine residues. This modulation of STIM1\'s Ca2+ sensitivity was further confirmed by Ca2+ imaging experiments, which showed that knockdown of these two PDIs does not affect STIM1-mediated SOCE upon full store depletion but leads to enhanced SOCE amplitudes upon partial store depletion. Thus, P4HB and PDIA3 dynamically modulate STIM1 activation by fine-tuning its Ca2+ binding affinity, adjusting the level of activated STIM1 in response to physiological cues. The coordination between STIM1-mediated Ca2+ signaling and redox responses reported herein may have implications for cell physiology and pathology.
摘要:
感觉到内质网(ER)钙(Ca2+)的降低,STIM1介导了一种普遍存在的Ca2流入过程,称为存储操作的Ca2进入(SOCE)。STIM1功能失调或SOCE异常与自身免疫性疾病密切相关。动脉粥样硬化,和各种形式的癌症。因此,揭示翻译后修饰的分子复杂性,如氧化,STIM1的功能至关重要。在最近的蛋白质组学筛查中,我们鉴定了三种蛋白质二硫键异构酶(PDIs)-脯氨酸4-羟化酶亚基β(P4HB),蛋白质二硫键异构酶A3(PDIA3),和含硫氧还蛋白结构域的蛋白5(TXNDC5)-作为STIM1的ER腔相互作用物。这里,我们证明了这些PDI与STIM1和STIM2动态关联。STIM1(STIM1-2CA)的两个保守半胱氨酸残基的突变降低了其在细胞和原位的Ca2亲和力。PDIA3或P4HB的敲除增加了野生型STIM1的Ca2亲和力,同时对STIM1-2CA突变体没有影响,表明PDIA3和P4HB通过作用于ER-管腔半胱氨酸残基来调节STIM1的Ca2+亲和力。这种STIM1的Ca2+敏感性的调制通过Ca2+成像实验进一步证实,这表明,这两个PDI的敲低不会影响STIM1介导的SOCE在完全存储耗尽时,但会导致SOCE振幅在部分存储耗尽时增强。因此,P4HB和PDIA3通过微调其Ca2结合亲和力动态调节STIM1激活,调节激活的STIM1水平以响应生理线索。本文报道的STIM1介导的Ca2+信号传导和氧化还原反应之间的协调可能对细胞生理学和病理学有影响。