Section of Infectious Disease, Department of Medicine, Baylor College of Medicine, Houston, TX 77030, USA.
Department of Molecular & Cellular Biology, Baylor College of Medicine, Houston, TX 77030, USA.
Cancers (Basel). 2014 Sep 29;6(4):2012-34. doi: 10.3390/cancers6042012.
Since its discovery in mice and humans 19 years ago, the contribution of alternatively spliced Stat3, Stat3β, to the overall functions of Stat3 has been controversial. Tyrosine-phosphorylated (p) Stat3β homodimers are more stable, bind DNA more avidly, are less susceptible to dephosphorylation, and exhibit distinct intracellular dynamics, most notably markedly prolonged nuclear retention, compared to pStat3α homodimers. Overexpression of one or the other isoform in cell lines demonstrated that Stat3β acted as a dominant-negative of Stat3α in transformation assays; however, studies with mouse strains deficient in one or the other isoform indicated distinct contributions of Stat3 isoforms to inflammation. Current immunological reagents cannot differentiate Stat3β proteins derived from alternative splicing vs. proteolytic cleavage of Stat3α. We developed monoclonal antibodies that recognize the 7 C-terminal amino acids unique to Stat3β (CT7) and do not cross-react with Stat3α. Immunoblotting studies revealed that levels of Stat3β protein, but not Stat3α, in breast cancer cell lines positively correlated with overall pStat3 levels, suggesting that Stat3β may contribute to constitutive Stat3 activation in this tumor system. The ability to unambiguously discriminate splice alternative Stat3β from proteolytic Stat3β and Stat3α will provide new insights into the contribution of Stat3β vs. Stat3α to oncogenesis, as well as other biological and pathological processes.
19 年前在小鼠和人类中发现的交替剪接 Stat3、Stat3β以来，其对 Stat3 整体功能的贡献一直存在争议。磷酸化酪氨酸（p）Stat3β同源二聚体更稳定，与 DNA 结合更紧密，不易去磷酸化，并且表现出不同的细胞内动力学，尤其是核保留时间明显延长，与 pStat3α同源二聚体相比。在细胞系中过表达一种或另一种异构体表明，Stat3β在转化实验中作为 Stat3α的显性负调控因子；然而，用一种或另一种异构体缺失的小鼠品系进行的研究表明，Stat3 异构体对炎症有不同的贡献。目前的免疫试剂无法区分源自替代剪接与 Stat3α蛋白水解切割的 Stat3β 蛋白。我们开发了能够识别 Stat3β 独特的 7 个 C 末端氨基酸（CT7）的单克隆抗体，并且与 Stat3α不发生交叉反应。免疫印迹研究表明，乳腺癌细胞系中 Stat3β 蛋白的水平，而不是 Stat3α，与总 pStat3 水平呈正相关，这表明 Stat3β 可能有助于该肿瘤系统中组成型 Stat3 激活。能够明确区分剪接替代 Stat3β与蛋白水解 Stat3β和 Stat3α的能力将为 Stat3β 与 Stat3α 对致癌作用以及其他生物学和病理过程的贡献提供新的见解。
