Examination of the Transcriptional Response to LaMIR166a Overexpression in Larix kaempferi (Lamb.) Carr

SIMPLE SUMMARY: The study of somatic embryogenesis can provide insights into early plant development. To elucidate the molecular mechanisms associated with miR166 in Larix kaempferi (Lamb.) Carr, the transcriptional profiles of wild-type (WT) and LaMIR166a-overexpressing embryonic cells were subjected to RNA sequencing. In total, 2467 differentially expressed genes were obtained. The cleaved degree of LaHDZ31–34 was higher in transgenic lines than in WT. The genes related to LaHDZ31–34 were screened by transcriptome analysis, and a yeast one-hybrid and dual-luciferase report assay revealed that LaHDZ31–34 could bind to the promoters of LaPAP, LaPP1, LaZFP5, and LaPHO1. This study provides insights into the regulatory mechanisms of miR166. ABSTRACT: The study of somatic embryogenesis can provide insight into early plant development. We previously obtained LaMIR166a-overexpressing embryonic cell lines of Larix kaempferi (Lamb.) Carr. To further elucidate the molecular mechanisms associated with miR166 in this species, the transcriptional profiles of wild-type (WT) and three LaMIR166a-overexpressing transgenic cell lines were subjected to RNA sequencing using the Illumina NovaSeq 6000 system. In total, 203,256 unigenes were generated using Trinity de novo assembly, and 2467 differentially expressed genes were obtained by comparing transgenic and WT lines. In addition, we analyzed the cleaved degree of LaMIR166a target genes LaHDZ31–34 in different transgenic cell lines by detecting the expression pattern of LaHdZ31–34, and their cleaved degree in transgenic cell lines was higher than that in WT. The downstream genes of LaHDZ31–34 were identified using Pearson correlation coefficients. Yeast one-hybrid and dual-luciferase report assays revealed that the transcription factors LaHDZ31–34 could bind to the promoters of LaPAP, LaPP1, LaZFP5, and LaPHO1. This is the first report of gene expression changes caused by LaMIR166a overexpression in Japanese larch. These findings lay a foundation for future studies on the regulatory mechanism of miR166.