The stability of vtRNA1-1 is influenced by the TRIM21-RBCK1-TRIM25 axis.
<p>(A) Immunoblot analysis using the indicated antibodies (left). After transfecting WT Huh7 cells with the siRNA targeting TRIM21, lysates were obtained and analyzed. NC denotes the negative control (mock transfection). GAPDH served as loading control. The intensity of indicated proteins was...
Saved in:
| Main Author: | |
|---|---|
| Other Authors: | |
| Published: |
2025
|
| Subjects: | |
| Tags: |
Add Tag
No Tags, Be the first to tag this record!
|
| Summary: | <p>(A) Immunoblot analysis using the indicated antibodies (left). After transfecting WT Huh7 cells with the siRNA targeting TRIM21, lysates were obtained and analyzed. NC denotes the negative control (mock transfection). GAPDH served as loading control. The intensity of indicated proteins was normalized to that of GAPDH (right, n=4 for TRIM25, and n=3 for RBCK1). Error bars indicate the standard deviation. *p < 0.05 (unpired <i>t t</i>est) (B) RT-qPCR analysis of <i>trim21</i> and <i>trim25</i> mRNA (n=4) after transfecting WT Huh7 cells with the siRNAs either targeting TRIM21 or TRIM25. Error bars indicate the standard deviation. *p < 0.05, ****p < 0.0001, ns: non-significant (two-way ANOVA test) (C) Immunoblot analysis using the indicated antibodies (left). After overexpressing TRIM21, lysates were obtained and analyzed. EV denotes the empty vector (mock transfection). GAPDH served as loading control. The intensity of RBCK1 was normalized to that of GAPDH (right, n=3). Error bars indicate the standard deviation. *p < 0.05 (unpaired test) (D) Immunoblot analysis using the indicated antibodies. After transfecting WT Huh7 cells with the siRNA targeting TRIM21 and treating with proteasomal inhibitor (MG132) or not (DMSO), lysates were obtained and analyzed. GAPDH served as a loading control. (E) Northern blot analysis using the indicated probes (top). After transfecting WT Huh7 cells with the siRNAs either targeting TRIM21, RBCK1, or both simultaneously (21&R), total RNA was extracted and analyzed. 5.8S rRNA served as loading control. The intensity of vtRNA1-1 was normalized to that of 5.8S rRNA (bottom, n=4). Error bars indicate the standard deviation. *p < 0.05, ****p < 0.0001 (one-way ANOVAtest) (F) Northern blot analysis using the indicated probes (left, top), and immunoblot analysis using the indicated antibodies (left, bottom). After transfecting WT Huh7 cells with the siRNA targeting TRIM21 and treating with proteasomal inhibitor (MG132) or not (DMSO), total RNA and lysate were obtained and analyzed. 5.8S rRNA served as loading control. The intensity of vtRNA1-1 was normalized to that of 5.8S rRNA (right, n=4). Error bars indicate the standard deviation. *p < 0.05, ***p < 0.001 (one-way ANOVA test) (G) Schematic model for the regulatory process of vtRNA1-1 stability mediated by the TRIM21-RBCK1-TRIM25 axis. Under normal conditions, TRIM21 mediates the proteasomal degradation of RBCK1, preventing TRIM25 degradation, thereby stabilizing vtRNA1-1 through the functions of TRIM21 and TRIM25 (left). When TRIM21 is depleted, undegraded RBCK1 induces TRIM25 degradation. This simultaneous lack of both TRIM21 and TRIM25 leads to a reduction in vtRNA1-1 stability (middle). Since TRIM25 knockdown shows no effect on TRIM21 levels, TRIM21 partially maintains vtRNA1-1 stability in the absence of TRIM25 (right).</p> |
|---|