Fig. 1: SNORD88B is highly expressed in liver CSCs.

A 5 × 10⁶ liver CSCs (CD13⁺CD133⁺) cells and non-CSCs (CD13⁻CD133⁻) were respectively enriched from human HCC tumor tissues via flow cytometry, followed by snoRNA sequencing (left panel). Differentially expressed snoRNAs (fold change >2) in CSCs were shown in the right panel, and the top 10 upregulated snoRNAs were denoted. B 5 upregulated snoRNAs with high abundance were depleted in HCC primary cells via shRNA. Spheres more than 100 μm in diameter were counted. HCC #1 and HCC #2 denoted HCC sample numbers. sh#1 and sh#2 indicate two effective shRNAs targeting snoRNAs. n = 3 independent experiments. Data are shown as means ± SD. Exact P values from left to right: 4.9E−08, 2.4E−07. C Schematic representation of human SNORD88B. The position of the arrowheads indicates SNORD88A/B/C position in linear gene locus. SNORD88B was denoted in red arrowheads. E1, exon #1. D, E Northern blot of SNORD88B expression levels in oncospheres (S) and non-spheres (N), or in CD13⁺CD133⁺ CSCs (C) and CD13⁻CD133⁻ non-CSCs (N). 18S rRNA served as a loading control. F Total RNA from HCC oncospheres was extracted from the cytoplasm, nucleoplasm, nucleolus, and followed by qRT-PCR (left panel) and Western blot (right panel). U1 RNA served as a positive control for nuclear location. β-actin, NUP62, and Nucleolin are protein markers for cytoplasm, nucleoplasm, and nucleolus, respectively. n = 3 biologically independent experiments. Data are shown as means ± SD. G Representative FISH staining of SNORD88B and nucleolar marker Nucleolin (NCL) in liver CSCs or non-CSCs originated from primary HCC cells. Nuclei are annotated by white dotted lines. Scale bar, 15 μm. H Violin Plot of SNORD88B expression in HCC tumor tissues compared with normal tissues using TCGA database. Mapping by R software. Exact P value: 1.7E−08. I Kaplan–Meier survival analysis of HCC patients, which were from the TCGA database and grouped into two subsets according to SNORD88B expression levels. Kaplan–Meier method with the log-rank test was used. Mapping by R software. Survival curves with a 95% confidence interval were shown. J Violin plot of SNORD88B expression in HCC primary tumors with different sizes using TCGA database. Exact P values from upper to lower: 0.0163, 0.036, 0.0018. T1 (n = 92), T2 (n = 57), T3 (n = 28), T4 (n = 11). K Box plot of SNORD88B expression in different stages of HCC using TCGA database. Boxes represent the interquartile range (IQR); upper and lower lines correspond to the 75th and 25th percentiles, respectively. Whiskers above and below boxes extend to the 5th and 95th percentiles, respectively. Horizontal lines within boxes represent median values. Stage I (n = 84), Stage II (n = 51), Stage III (n = 48). Exact P value: 0.0225 *P < 0.05; **P < 0.01; ***P < 0.001 by two-tailed Student’s T-test. Data are representative of at least three independent experiments.