Sarkar A, Yang P, Fan Y H, Mu Z M, Hauptmann R, Adolf G R, Stass S A, Chang K S
Molecular Hematopathology Program, University of Texas M.D. Anderson Cancer Center, Houston 77030.
Blood. 1994 Jul 1;84(1):279-86.
Annexin VIII is a calcium-dependent phospholipid-binding protein previously identified as a blood anticoagulant based on in vitro studies. However, the physiologic function of annexin VIII remains unknown. In acute promyelocytic leukemia (APL) the annexin VIII gene is highly expressed, but its expression is undetectable in the blasts of other acute leukemias. In the present investigation, we showed using the APL-derived NB4 cell line that expression of the annexin VIII gene is regulated at the transcription level during induced differentiation by all-trans retinoic acid (ATRA). The half-life of the annexin VIII mRNA is about 5 to 6 hours, as determined by using actinomycin D as a transcription inhibitor. Analysis of the expression of annexin VIII protein in NB4 cells and in APL samples showed a consistent expression of a predominant 36-kD protein and a weak 72-kD protein. After ATRA-induced differentiation of NB4 cells, the annexin VIII protein level reduced gradually, but a detectable level persisted even after 4 days of induction. Because annexin VIII mRNA becomes undetectable after 48 hours of ATRA induction, this result indicates that annexin VIII is a relatively stable protein. A multiple tissue Northern blot analysis was performed, and we found that annexin VIII is normally expressed in the placenta and the lung. Cellular localization of the annexin VIII protein was determined by immunofluorescence staining and subcellular fractionation. These results indicated that annexin VIII is predominantly localized to the plasma membrane. The annexin VIII is neither an extracellular protein nor associated with the cell surface suggesting that it does not play a role in blood coagulation in vivo. The plasma membrane localization and its property as a phospholipase inhibitor suggests that annexin VIII may have a role in the signal transduction pathway in the APL cells.
膜联蛋白VIII是一种钙依赖性磷脂结合蛋白，基于体外研究，它先前被鉴定为血液抗凝剂。然而，膜联蛋白VIII的生理功能仍然未知。在急性早幼粒细胞白血病（APL）中，膜联蛋白VIII基因高度表达，但在其他急性白血病的原始细胞中未检测到其表达。在本研究中，我们使用源自APL的NB4细胞系表明，全反式维甲酸（ATRA）诱导分化过程中，膜联蛋白VIII基因的表达在转录水平受到调控。通过使用放线菌素D作为转录抑制剂测定，膜联蛋白VIII mRNA的半衰期约为5至6小时。对NB4细胞和APL样本中膜联蛋白VIII蛋白表达的分析显示，主要的36-kD蛋白和较弱的72-kD蛋白一致表达。ATRA诱导NB4细胞分化后，膜联蛋白VIII蛋白水平逐渐降低，但即使在诱导4天后仍保持可检测水平。由于ATRA诱导48小时后膜联蛋白VIII mRNA变得不可检测，该结果表明膜联蛋白VIII是一种相对稳定的蛋白。进行了多组织Northern印迹分析，我们发现膜联蛋白VIII在胎盘和肺中正常表达。通过免疫荧光染色和亚细胞分级分离确定膜联蛋白VIII蛋白的细胞定位。这些结果表明，膜联蛋白VIII主要定位于质膜。膜联蛋白VIII既不是细胞外蛋白，也不与细胞表面相关，这表明它在体内血液凝固中不起作用。质膜定位及其作为磷脂酶抑制剂的特性表明，膜联蛋白VIII可能在APL细胞的信号转导途径中起作用。
