Vaisid Taly, Kosower Nechama S
Department of Human Molecular Genetics and Biochemistry, Sackler School of Medicine, Tel-Aviv University, Ramat-Aviv, Tel-Aviv, Israel.
Biochim Biophys Acta. 2013 Oct;1833(10):2369-77. doi: 10.1016/j.bbamcr.2013.06.006. Epub 2013 Jun 14.
Calpain (intracellular Ca(2+)-dependent protease) and calpastatin (calpain specific endogenous inhibitor) are widely distributed in biological systems, and have been implicated in many cellular physiological and pathological processes. Calpastatin level is of central importance to the control of calpain activity. We demonstrated for the first time that calpastatin is overexpressed in mycoplasma-contaminated cultured cells (SH-SY5Y cells that are infected by a strain of Mycoplasma hyorhinis (NDMh)). We have found that the calpastatin-upregulating activity resides in the mycoplasmal membrane lipoproteins, and is associated with NF-κB activation. Calpain-promoted proteolysis is attenuated in the NDMh lipoprotein-treated cells. Here we show that the NDMh lipoproteins promoted an increase in calpastatin in SH-SY5Y cells via the TLR2/TAK1/NF-κB pathway. The synthetic mycoplasmal lipopeptide MALP-2 and the bacterial lipopeptide PAM3CSK4 (TLR2 agonists) also promoted calpastatin upregulation. LPS (TLR4 agonist) activated NF-κB without calpastatin increase in the cell. In contrast, lipoteichoic acid (TLR2 agonist) upregulated calpastatin not via NF-κB activation, but via the MEK1/ELK1 pathway. Zymosan and peptidoglycan, TLR2 agonists that lack lipids, did not induce calpastatin upregulation. Cell treatment with a calpastatin-upregulating agonist (lipoteichoic acid) led to the attenuation of Ca(2+)-promoted calpain activity, whereas agonists that do not upregulate calpastatin (LPS, Zymosan) were ineffective. Overall, the results indicate that in these non-immune cells, calpastatin is upregulated by TLR2-agonists containing lipids, with more than one downstream pathway involved. Such agonists may be useful for studying mechanisms and factors involved in calpastatin regulation. In addition, suitable TLR2 agonists may be of interest in devising treatments for pathological processes involving excessive calpain activation.
钙蛋白酶（细胞内钙离子依赖性蛋白酶）和钙蛋白酶抑制蛋白（钙蛋白酶特异性内源性抑制剂）广泛分布于生物系统中，并与许多细胞生理和病理过程有关。钙蛋白酶抑制蛋白水平对于控制钙蛋白酶活性至关重要。我们首次证明，在支原体污染的培养细胞（被猪鼻支原体菌株（NDMh）感染的SH-SY5Y细胞）中钙蛋白酶抑制蛋白过表达。我们发现，钙蛋白酶抑制蛋白上调活性存在于支原体膜脂蛋白中，并与NF-κB激活相关。在NDMh脂蛋白处理的细胞中，钙蛋白酶促进的蛋白水解作用减弱。在此我们表明，NDMh脂蛋白通过TLR2/TAK1/NF-κB途径促进SH-SY5Y细胞中钙蛋白酶抑制蛋白增加。合成支原体脂肽MALP-2和细菌脂肽PAM3CSK4（TLR2激动剂）也促进钙蛋白酶抑制蛋白上调。脂多糖（TLR4激动剂）激活NF-κB，但细胞中钙蛋白酶抑制蛋白未增加。相反，脂磷壁酸（TLR2激动剂）不是通过NF-κB激活上调钙蛋白酶抑制蛋白，而是通过MEK1/ELK1途径上调。缺乏脂质的TLR2激动剂酵母聚糖和肽聚糖未诱导钙蛋白酶抑制蛋白上调。用钙蛋白酶抑制蛋白上调激动剂（脂磷壁酸）处理细胞导致钙离子促进的钙蛋白酶活性减弱，而不上调钙蛋白酶抑制蛋白的激动剂（脂多糖、酵母聚糖）无效。总体而言，结果表明，在这些非免疫细胞中，含脂质的TLR2激动剂上调钙蛋白酶抑制蛋白，涉及不止一条下游途径。此类激动剂可能有助于研究参与钙蛋白酶抑制蛋白调节的机制和因素。此外，合适的TLR2激动剂可能在设计涉及过度钙蛋白酶激活的病理过程的治疗方法方面具有意义。
