Araujo-Abad Salomé, García-Peñarrubia Borja, Giudici A Marcela, Neira José L, Rizzuti Bruno, de Juan Romero Camino, Poveda José A
Cancer Research Group, Faculty of Engineering and Applied Sciences, Universidad de Las Américas, 170124 Quito, Ecuador.
IDIBE, Universidad Miguel Hernández, 03202 Elche, Alicante, Spain; Unidad de Investigación, Fundación para el Fomento de la Investigación Sanitaria y Biomédica de la Comunidad Valenciana (FISABIO), Hospital General Universitario de Elche, Camí de l'Almazara 11, 03203 Elche, Alicante, Spain.
J Mol Biol. 2025 Sep 1;437(17):169297. doi: 10.1016/j.jmb.2025.169297. Epub 2025 Jun 16.
Cancer cells need to implement and maintain molecular mechanisms relying on an energy trade-off between resistance and key functions to survive. Among them, changes in lipid metabolism are crucial. Phosphatidylserine (PS), phosphatidylcholine (PC) and phosphatidic acid (PA) are some of the lipids forming cell membranes and having key functions in lipid metabolism. PADI4 is an enzyme implicated in the conversion of arginine to citrulline (citrullination), that has been related with the development of several types of cancers. In this work, we report the lipid binding properties of PADI4. Such binding was assayed in vitro against zwitterionic (PC) and anionic (PA and PS) lipids, and it was monitored by several biophysical techniques. Furthermore, results in silico showed that the binding of PADI4 with PA, PC and PS occurred at the active site of the enzyme. This binding was confirmed in cellulo by using PS as the target lipid, employing immunofluorescence (IF) and proximity ligation assay (PLA) in different cancer cell lines. The use in cellulo of a specific enzymatic inhibitor of PADI4, GSK484, abolished the binding between PADI4 and PS in cancer cells, further indicating that their interaction occurred at the protein active site. Altogether, this work shows that PADI4 was capable of binding to lipids, and opens the venue to study the role that it could be playing in deimination processes and cancer development. Moreover, this study lays the foundation for developing novel cancer therapies from new perspectives, based on the interaction of lipids with citrullinating enzymes.
癌细胞需要实施并维持依赖于抗性与关键功能之间能量权衡的分子机制才能存活。其中，脂质代谢的变化至关重要。磷脂酰丝氨酸（PS）、磷脂酰胆碱（PC）和磷脂酸（PA）是构成细胞膜且在脂质代谢中具有关键功能的一些脂质。肽基精氨酸脱亚氨酶4（PADI4）是一种参与将精氨酸转化为瓜氨酸（瓜氨酸化）的酶，它与多种癌症的发展有关。在这项研究中，我们报告了PADI4的脂质结合特性。这种结合在体外针对两性离子脂质（PC）和阴离子脂质（PA和PS）进行了测定，并通过多种生物物理技术进行监测。此外，计算机模拟结果表明，PADI4与PA、PC和PS的结合发生在该酶的活性位点。通过使用PS作为靶脂质，在不同癌细胞系中采用免疫荧光（IF）和邻近连接分析（PLA），在细胞内证实了这种结合。在细胞内使用PADI4的特异性酶抑制剂GSK484，消除了癌细胞中PADI4与PS之间的结合，进一步表明它们的相互作用发生在蛋白质活性位点。总之，这项研究表明PADI4能够结合脂质，并为研究其在脱亚氨基过程和癌症发展中可能发挥的作用开辟了道路。此外，这项研究为基于脂质与瓜氨酸化酶的相互作用从新的角度开发新型癌症治疗方法奠定了基础。
