Bhavsar Shefalee K, Gu Shuchen, Bobbala Diwakar, Lang Florian
Department of Physiology, University of Tübingen, Gmelinstrasse 5, Tübingen, Germany.
Cell Physiol Biochem. 2011;27(5):547-56. doi: 10.1159/000329956. Epub 2011 Jun 15.
Janus kinase 3, a tyrosine kinase expressed in haematopoetic tissues, plays a decisive role in T-lymphocyte survival. JAK3 deficiency leads to (Severe) Combined Immunodeficiency (SCID) resulting from enhanced lymphocyte apoptosis. JAK3 is activated by phosphorylation. Nothing is known about expression of JAK3 in erythrocytes, which may undergo apoptosis-like cell death (eryptosis) characterized by cell membrane scrambling with phosphatidylserine exposure and cell shrinkage. Triggers of eryptosis include energy depletion. The present study utilized immunohistochemistry and confocal microscopy to test for JAK3 expression and phosphorylation, and FACS analysis to determine phosphatidylserine exposure (annexin binding) and cell volume (forward scatter). As a result, JAK3 was expressed in erythrocytes and phosphorylated following 24h and 48h glucose depletion. Forward scatter was slightly but significantly smaller in erythrocytes from JAK3-deficient mice (jak3(-/-)) than in erythrocytes from wild type mice (jak3(+/+)). Annexin V binding was similarly low in both genotypes. The JAK3 inhibitors WHI-P131/JANEX-1 (4-(4'-Hydroxyphenyl)amino-6,7-dimethoxyquinazoline, 156μM) and WHI-P154 (4-[(3'-Bromo-4'-hydroxyphenyl)amino]-6,7-dimethoxyquinazoline, 11.2μM) did not significantly modify annexin V binding or forward scatter. Glucose depletion increased annexin V binding, an effect significantly blunted in jak3(-/-) erythrocytes and in the presence of the JAK3 inhibitors. The observations disclose a completely novel role of Janus kinase 3, i.e. the triggering of cell membrane scrambling in energy depleted erythrocytes.
Janus激酶3是一种在造血组织中表达的酪氨酸激酶，在T淋巴细胞存活中起决定性作用。JAK3缺陷导致淋巴细胞凋亡增强，从而引发严重联合免疫缺陷（SCID）。JAK3通过磷酸化被激活。关于JAK3在红细胞中的表达情况尚无定论，红细胞可能会经历类似凋亡的细胞死亡（红细胞凋亡），其特征是细胞膜磷脂酰丝氨酸暴露导致膜紊乱和细胞收缩。红细胞凋亡的触发因素包括能量耗竭。本研究利用免疫组织化学和共聚焦显微镜检测JAK3的表达和磷酸化情况，并通过流式细胞术分析确定磷脂酰丝氨酸暴露（膜联蛋白结合）和细胞体积（前向散射）。结果显示，JAK3在红细胞中表达，且在葡萄糖耗竭24小时和48小时后发生磷酸化。JAK3缺陷小鼠（jak3(-/-)）红细胞的前向散射略低于野生型小鼠（jak3(+/+)）红细胞，但差异具有统计学意义。两种基因型的膜联蛋白V结合率均较低。JAK3抑制剂WHI-P131/JANEX-1（4-(对羟基苯基)氨基-6,7-二甲氧基喹唑啉，156μM）和WHI-P154（4-[(3'-溴-4'-羟基苯基)氨基]-6,7-二甲氧基喹唑啉，11.2μM）对膜联蛋白V结合或前向散射无显著影响。葡萄糖耗竭增加了膜联蛋白V结合，这一效应在jak3(-/-)红细胞及JAK3抑制剂存在的情况下显著减弱。这些观察结果揭示了Janus激酶3的一个全新作用，即在能量耗竭的红细胞中引发细胞膜紊乱。
