Fig. 8 | Signal Transduction and Targeted Therapy

Fig. 8

From: LncRNA Wee1-AS coordinates oxidative fatty acid metabolism through the activation of mitochondrial CDK1/CYCLIN B1

Fig. 8The alternative text for this image may have been generated using AI.

Knockdown of LNC106435.1, a human analog of Wee1-AS, suppresses mitochondrial function. a Transcript of LNC106435.1 was annotated previously (NCBI Reference Sequence NONHSAG106435.1) (dark blue). In this study, LNC106435.1 transcripts were identified from Huh7 cells via RACE (light blue). The size of the transcript is shown in parenthesis (bottom). b Cytosol- (blue) and nucleus- (red) RNAs in the subcellular fractions of primary hepatocytes were purified. The RNA levels of LNC106435.1, RPS14, and snRNA U1 were analyzed via qRT‒PCR. The proper cellular fractionation was confirmed by localization of RPS14 and snRNA U1. The data are represented as the means ± SDs (n = 4). c Primary human hepatocytes were transfected with ASO-control (ctl) or ASO-LNC106435.1. The RNA levels of LNC106435.1 were measured by RT‒PCR. The RNA and protein levels of Wee1 were analyzed via qRT‒PCR and western blotting, respectively. d RNA pull-down assays were carried out with biotinylated LNC106435.1 (1-646 nt, within which the sequence is conserved in mouse Wee1-AS with 50.59% identity) (sense, S) or its antisense (AS) with protein lysates prepared from Huh7 cells. Pull-down proteins were analyzed via western blotting. e Primary human hepatocytes were transfected with si-control (ctl) or si-LNC106435.1. The cells were stained with 100 nM MitoTracker Red CMXRos, and examined via a confocal microscope. Scale bar, 20 μm. Representative images are shown. The fluorescence intensity in at least 100 cells was quantified via Image J software. The data were analyzed via the Mann‒Whitney test. **P < 0.01 vs si-control (ctl). f The expression levels of LNC106435.1 were analyzed via two publicly available NCBI GEO datasets, namely, GSE162694, which included 143 individuals categorized as healthy controls (HCs) or patients with NAFLD activity scores (NASs) ranging from 1 to 7. Patients were further classified as mild MASLD (NAS 1–2), borderline MASLD (NAS 3–4), and advanced MASH (NAS ≥ 5), and GSE126848 which included 57 individuals comprising HCs and patients with obesity, MASL, or MASH. The transcript abundance of COL3A1 (positive control) and LNC106435.1 was quantified via Salmon software and expressed as log(TPM + 1). The white circles represent the mean values. The data were analyzed via the t test. *P < 0.05, **P < 0.01, and ***P < 0.001. g Representative images of immunohistochemical staining of WEE1, p-CDK1 (Y15), and CYCLIN B1 in human liver tissue samples from normal, MASL, and MASH patients. Scale bar, 50 μm. h Schematic illustration of how the spatiotemporal lncRNA Wee1-AS controls mitochondrial function through the activation of CDK1/CYCLIN B1 (created with BioRender.com)

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