The AT-HOOK MOTIF CONTAINING NUCLEAR LOCALIZED gene promotes somatic embryogenesis and polyploidy formation in...

This study reveals that LhAH15 is localized to the nucleus and functions as a transcription factor involved in somatic embryogenesis. LhAH15 enhances both somatic embryogenesis effi ciency and embryogenic callus induction efficiency in Liriodendron hybrids by regulating the expression of key genes in auxin and cytokinin signaling pathways. Furthermore, overexpression of LhAH15 also induces polyploidy formation. The artificially created Liriodendron hybrids, generated through the crossbreeding of L. chinense and L. tulipifera, exhibit enhanced stress resilience and superior growth characteristics. Somatic embryogenesis is a developmental process in which somatic cells undergo dedifferentiation to form embryogenic cells, which subsequently develop into entire plants under controlled conditions. In this study, the conserved transcription factor gene LhAHL15 was successfully cloned, and overexpression vectors as well as CRISPR/Cas9 vectors were constructed to investigate its role in somatic embryogenesis. Subcellular localization analysis revealed that the LhAHL15 protein is localized within the nucleus, suggesting its function as a transcriptional regulator involved in the regulation of plant growth and development. Further examination of the spatiotemporal expression profile of the LhAHL15 promoter demonstrated that transcription of LhAHL15 is initiated during the early stages of somatic embryogenesis and remains active throughout the formation of spherical, heart-shaped, torpedo-shaped, and cotyledonary embryos. These results underscore the importance of LhAHL15 as a key transcriptional regulator in somatic embryogenesis. Notably, overexpression of LhAHL15 in Liriodendron hybrids significantly upregulates the expression of critical genes associated with somatic embryogenesis, including BBM, LEC1, PIN1, PLT2, ARR7, and ARF12. This upregulation markedly improved the efficiency of somatic embryogenesis and the induction of embryogenic callus. Flow cytometry analysis revealed that the chromosome number of the AHL15-OE line was approximately twice that of the control, indicating a polyploid state. These findings suggest that overexpression of LhAHL15 may induce chromosome doubling in plants. In summary, our study reveals the pivotal role of LhAHL15 in promoting somatic embryogenesis and inducing polyploidy. This advancement effectively addresses the technical limitations of traditional polyploid breeding approaches, thereby establishing a solid theoretical foundation and providing technical support for the improvement and utilization of Liriodendron hybrids.