Choi Song-Yi, Hong Su-Hyung, Lee Heon-Jin
Department of Microbiology and Immunology, School of Dentistry, Kyungpook National University, Daegu, 41940, South Korea; Craniofacial Nerve-Bone Network Research Center, Kyungpook National University, Daegu, 41940, South Korea.
Department of Microbiology and Immunology, School of Dentistry, Kyungpook National University, Daegu, 41940, South Korea.
Biochem Biophys Res Commun. 2022 Jun 25;610:147-153. doi: 10.1016/j.bbrc.2022.04.048. Epub 2022 Apr 14.
Extracellular vesicles such as exosomes in eukaryotes have drawn scrutiny due to their various roles in intercellular communication. Small RNAs, including microRNAs (miRNAs), are more abundant among the cargo of exosomes than other RNA types. MiRNAs loaded in secreted exosomes (or extracellular microRNAs) can be transported to recipient cells and may play a regulatory role although the miRNA loading (or sorting) mechanism in exosomes has not been clearly elucidated. Therefore, this study analyzed exosomal miRNA sequencing data from human myeloid U937 cells treated with phorbol 12-myristate 13-acetate (PMA) and compared it with data from PMA-untreated U937 cells. MiR-24 was highly expressed in the cytoplasm and exosomes of PMA-treated U937 cells. Also, miRNA pull-down and mass spectrophotometry analysis of PMA-treated U937 cells revealed that miR-24 was specifically associated with α-tubulin and hnRNP-E1 proteins. Furthermore, exosomal miR-24 was dramatically reduced when those proteins were inactivated with siRNAs, whereas cellular miR-24 showed no significant effect. We conclude that miR-24 is transported into exosomes from activated macrophages with the support of α-tubulin and hnRNP-E1.
真核生物中的细胞外囊泡，如外泌体，因其在细胞间通讯中的多种作用而受到关注。小RNA，包括微小RNA（miRNA），在外泌体的货物中比其他RNA类型更为丰富。分泌型外泌体中装载的miRNA（或细胞外miRNA）可以转运到受体细胞，并可能发挥调节作用，尽管外泌体中的miRNA装载（或分选）机制尚未完全阐明。因此，本研究分析了用佛波酯12-肉豆蔻酸酯13-乙酸酯（PMA）处理的人髓样U937细胞的外泌体miRNA测序数据，并将其与未用PMA处理的U937细胞的数据进行比较。miR-24在PMA处理的U937细胞的细胞质和外泌体中高表达。此外，对PMA处理的U937细胞进行miRNA下拉和质谱分析发现，miR-24与α-微管蛋白和hnRNP-E1蛋白特异性相关。此外，当这些蛋白质被siRNA失活时，外泌体miR-24显著减少，而细胞内miR-24没有显著变化。我们得出结论，在α-微管蛋白和hnRNP-E1的支持下，miR-24从活化的巨噬细胞转运到外泌体中。
