Luo Xiaomin, Liu Fang, Zhu Lijun, Liu Caizhi, Shen Ruhui, Ding Xiaoyin, Wang Yufan, Tang Xiaofang, Peng Yongde, Zhang Zhijian
Department of Endocrinology and Metabolism, Shanghai General Hospital, Shanghai Jiao Tong University School of Medicine, Shanghai, China.
Department of Endocrinology and Metabolism, Shanghai General Hospital, Shanghai Jiao Tong University School of Medicine, Shanghai, China.
Mol Metab. 2025 Jan;91:102075. doi: 10.1016/j.molmet.2024.102075. Epub 2024 Nov 26.
As the primary source of glucose during fasting, hepatic gluconeogenesis is rigorously regulated to maintain euglycemia. Abnormal gluconeogenesis in the liver can lead to hyperglycemia, a key diagnostic marker and the primary pathological contributor to type 2 diabetes (T2D) and metabolic disorders. Hepatic nuclear factor-4 (HNF4α) is an important regulator of gluconeogenesis. In this study, we identify leupaxin (LPXN) as a novel coactivator for HNF4α. Although previous studies have shown that LPXN is highly correlated with cancer types such as B-cell differentiation and hepatocellular carcinoma progression, the role of LPXN in gluconeogenesis remains unknown.
We initially used protein pull-down assays, mass spectrometry and luciferase assays to identify the coactivator that interacts with HNF4α in gluconeogenesis. We further leveraged cell cultures and mouse models to validate the functional importance of molecular pathway during gluconeogenesis by using adenovirus-mediated overexpression and adeno-associated virus shRNA-mediated knockdown both in vivo and ex vivo, such as in ob/db/DIO mice, HepG2 and primary hepatocytes. Following, we used CUT&Tag and chip qPCR to identify the LPXN-mediated mechanisms underlying the observed abnormal gluconeogenesis. Additionally, we assessed the translational relevance of our findings using human liver tissues from both healthy donors and patients with obesity/type 2 diabetes.
We found that LPXN interacts with HNF4α to participate in gluconeogenesis. Knockdown of LPXN expression in the liver effectively enhanced glucose metabolism, while its overexpression in the liver effectively inhibited it. Mechanistically, LPXN could translocate into the nucleus and was essential for regulating gluconeogenesis by binding to the PEPCK promoter, which controlled the expression of an enzyme involved in gluconeogenesis, mainly through the Gcg-cAMP-PKA pathway. Additionally, LPXN expression was found to be increased in the livers of patients with steatosis and diabetes, supporting a pathological role of LPXN.
Taken together, our study provides evidence that LPXN plays a critical role in modulating hepatic gluconeogenesis, thereby reinforcing the fact that targeting LPXN may be a potential approach for the treatment of diabetes and metabolic disorders.
作为空腹期间葡萄糖的主要来源，肝脏糖异生受到严格调控以维持血糖正常。肝脏中异常的糖异生会导致高血糖，这是2型糖尿病（T2D）和代谢紊乱的关键诊断标志物及主要病理促成因素。肝细胞核因子-4（HNF4α）是糖异生的重要调节因子。在本研究中，我们鉴定出leupaxin（LPXN）是HNF4α的一种新型共激活因子。尽管先前的研究表明LPXN与B细胞分化和肝细胞癌进展等癌症类型高度相关，但LPXN在糖异生中的作用仍不清楚。
我们最初使用蛋白质下拉测定、质谱和荧光素酶测定来鉴定在糖异生中与HNF4α相互作用的共激活因子。我们进一步利用细胞培养和小鼠模型，通过在体内和体外使用腺病毒介导的过表达和腺相关病毒shRNA介导的敲低，验证糖异生过程中分子途径的功能重要性，如在ob/db/DIO小鼠、HepG2细胞和原代肝细胞中。随后，我们使用CUT&Tag和芯片qPCR来确定LPXN介导的观察到的异常糖异生的潜在机制。此外，我们使用来自健康供体以及肥胖/2型糖尿病患者的人类肝脏组织评估了我们研究结果的转化相关性。
我们发现LPXN与HNF4α相互作用以参与糖异生。肝脏中LPXN表达的敲低有效地增强了葡萄糖代谢，而其在肝脏中的过表达则有效地抑制了葡萄糖代谢。从机制上讲，LPXN可以转运到细胞核中，并且通过与磷酸烯醇式丙酮酸羧激酶（PEPCK）启动子结合对调节糖异生至关重要，PEPCK启动子控制着一种参与糖异生的酶的表达，主要通过胰高血糖素-cAMP-蛋白激酶A（Gcg-cAMP-PKA）途径。此外，发现脂肪变性和糖尿病患者肝脏中LPXN的表达增加，这支持了LPXN的病理作用。
综上所述，我们的研究提供了证据表明LPXN在调节肝脏糖异生中起关键作用，从而强化了靶向LPXN可能是治疗糖尿病和代谢紊乱的一种潜在方法这一事实。
