宫颈癌是导致全球女性因癌症死亡的主要原因之一，在女性的恶性肿瘤中总体发病率和死亡率排名第四，严重威胁到女性的生命健康，因此对宫颈癌的机制研究迫在眉睫。CHIP是一种E3泛素连接酶，也是分子伴侣结合蛋白。近年来，越来越多的研究聚焦到CHIP在癌症中的作用，本实验室首次发现CHIP在宫颈癌中起到抑癌因子的作用。

为了探究CHIP抑制宫颈癌的分子机制，基于CHIP作为E3泛素连接酶的特性，我们寻找到一个在细胞水平上与CHIP蛋白表达呈负相关的潜在靶点TH1L。进一步数据库预测显示，在组织水平，与正常宫颈组织相比，在宫颈癌组织中，CHIP低表达、TH1L高表达，并呈负相关。随后，我们通过细胞增殖、克隆形成、划痕、Transwell和EMT相关标志物检测等一系列实验证实TH1L在宫颈癌中的作用是促进癌症细胞的生长和迁移。接下来，我们通过内外源IP及GST pull down实验确认了CHIP和TH1L具有相互作用，且CHIP参与相互作用的具体区域为TPR结构域。通过内外源 CHX 实验发现，CHIP能够降低TH1L蛋白稳定性。降解和泛素化实验证实，CHIP通过泛素蛋白酶体途径降解TH1L。数据库结合点突变实验显示，TH1L主要是K58和K116位点被泛素化。更进一步，利用构建好的CHIP功能缺失突变体，发现分子伴侣影响CHIP和TH1L的相互作用、降解和泛素化作用，说明CHIP对TH1L的调控是分子伴侣依赖的。最后，我们在过表达CHIP的基础上回补TH1L，细胞增殖、划痕以及克隆形成等实验显示，TH1L削弱了CHIP对宫颈癌的抑制作用。

此外，我们通过蛋白质双向电泳结合质谱的组学方法，在宫颈癌中寻找到CHIP另一个可能的底物蛋白S100A11。Co-IP实验显示，CHIP和S100A11存在明显的相互作用。进一步降解实验发现，CHIP可以通过蛋白酶体途径降解S100A11。

综上所述，本论文首次发现CHIP在宫颈癌中的两个新底物TH1L和S100A11，并证实TH1L是宫颈癌的促癌因子，而CHIP可以通过降解TH1L抑制宫颈癌的生长和转移。以上发现对于深入了解CHIP在宫颈癌治疗中的具体调节机制具有至关重要的作用，并为宫颈癌的靶向治疗开辟了新的研究途径。

Cervical cancer is one of the leading causes of cancer-related deaths in women worldwide, ranking fourth in overall incidence and mortality among female malignancies, posing a serious threat to women's lives and health. Therefore, there is an urgent need to study the mechanism of cervical cancer. CHIP is an E3 ubiquitin ligase and a chaperone-binding protein. In recent years, more and more studies have focused on the function  of CHIP in cancer.Our laboratory firstly found that CHIP's function as a tumor suppressor in cervical cancer.

In order to explore the molecular mechanism of CHIP inhibition of cervical cancer, based on the properties of CHIP as an E3 ubiquitin ligase, we identified a potential target TH1L that was negatively correlated with CHIP protein expression at the cellular level. Further database predictions showed that at the tissue level, there was a negative correlation between low expression of CHIP and high expression of TH1L in cervical cancer tissues compared with normal cervical tissues. Subsequently, we confirmed the role of TH1L in cervical cancer by promoting the growth and migration of cancer cells through a series of experiments such as cell proliferation, clone formation, scratching, Transwell and EMT-related marker detection. Next, we confirmed that CHIP and TH1L interacted with each other through endogenous and exogenous IP and GST pull down experiments, and the specific region of CHIP interaction was the TPR domain.Endogenous and exogenous CHX experiments showed that CHIP could reduce the stability of TH1L protein. Experiments on degradation and ubiquitination verified that CHIP degraded TH1L through the ubiquitin proteasome pathway. Database binding point mutation experiments showed that TH1L was mainly ubiquitinated at K58 and K116 sites. Furthermore, using the constructed CHIP loss-of-function mutants, it was found that chaperones affected the interaction, degradation and ubiquitination of CHIP and TH1L, indicating that the regulation of TH1L by CHIP  was chaperone-dependent. Finally, we supplemented TH1L on the basis of overexpression of CHIP, and experiments such as cell proliferation, scratching and clone formation indicated that TH1L weakened the inhibitory effect of CHIP on cervical cancer.

In addition, we used protein two-dimensional electrophoresis combined with mass spectrometry to find another possible substrate protein S100A11 for CHIP in cervical cancer. Co-IP experiments indicated that there was a significant interaction between CHIP and S100A11.Further degradation experiments revealed that CHIP could   degrade S100A11 by the ubiquitin proteasome pathway.

In summary, this paper firstly identifies two new substrates of CHIP in cervical cancer, TH1L and S100A11 and confirms that TH1L is a cancer-promoting factor in cervical cancer. CHIP can inhibit the growth and metastasis of cervical cancer by degrading TH1L.The above findings are crucial for understanding the specific regulatory mechanism of CHIP in the treatment of cervical cancer and open up a new research avenue for targeted therapy of cervical cancer.