Fig 1: UASR1 promoted CR4 cell proliferation through the miR-107/CDK8 axis. (A) The effects of overexpression of UASR1, miR-107 and CDK8 on the proliferation of CR4 cells were analyzed by performing cell proliferation. (B) Reverse transcription-quantitative polymerase chain reaction was performed to confirm the overexpression of CDK8 in cells with the CDK8 expression vector and UASR1-overexpression and miR-107 in cells co-transfected with UASR1 expression vector and miR-107 mimic. *P<0.05. NC negative control.
Fig 2: CDK8 does not regulate HIV-1 infection, and the activity of dCA is independent of CDK8. HeLa CD4 cells were transduced with VLPs expressing shRNAs against CDK8, CDK19, or both and puromycin selected to stabilize knockdown (KD). These cells were then infected with the NL4-3 strain for 24 h, in the presence of DMSO or dCA (200 nM). After 24 hours, cells were washed and fresh medium with compounds was added. Seventy hours later, mRNA, DNA, and p24 capsid in supernatant were collected and measured. (A and B) CDK8 mRNA and CDK19 mRNA expression was assessed by qRT-PCR and normalized to GAPDH mRNA. Shown is the mean ± SEM from n = 3 independent experiments. (C) Protein expression of CDK8 and CDK19 was analyzed 72 hours postinfection by Western blotting. GAPDH was used as an internal control. The image shown is representative of n = 3 independent experiments. (D) Viral mRNA production measured by qRT-PCR using primers against the Nef region was measured as a function of total integrated proviral DNA. Data represent the mean ± SEM from n = 3 independent experiments. (E) p24 capsid production in the supernatant relative to total HIV DNA and normalized to the DMSO of each condition is represented. Data represent the mean ± SEM from n = 3 independent experiments. Statistical significance was determined using one-way ANOVA with post hoc Tukey’s test. ns, not significant; nsa, DMSO KD CDK19 versus dCA KD CDK19. *, P < 0.01; ***, P < 0.0001; **, P < 0.001.
Fig 3: UASR1 and CDK8 were upregulated in CRC and high expression levels of UASR1 in CRC predicted poor survival. The expression of (A) UASR1 and (B) CDK8 in paired CRC and non-tumor tissues from 62 patients with CRC were determined by reverse transcription-quantitative polymerase chain reaction. ***P<0.001. The 62 patients with CRC were divided into high and low UASR1 expression groups (n=31) with the median level of UASR1 in CRC tissues as the cut-off value. Survival curves were plotted for the two groups based on follow-up data. Survival curves were compared using the log-rank test. (C) It is worth noting that miR-107 expression was not significantly correlated with patient survival (P=0.488; HR=1.123; 95% CI: 0.598–2.117), while the expression of CDK8 was correlated with patient survival (P=0.0167; HR=2.376; 95% CI: 1.237–4.417). CRC, colorectal cancer; HR, hazards ratio; CI, confidence interval.
Fig 4: The expression of CDK8 was upregulated in CR4 cells with UASR1-overexpression. To test the possibility of UASR1 as an internal sponge of miR-107, the effects of overexpression of UASR1 and miR-107 on the expression of CDK8, a miR-107 target, were analyzed by (A) reverse transcription-quantitative polymerase chain reaction and (B) western blot analysis. All polymerase chain reaction reactions were repeated 3 times and mean values were presented and compared. *P<0.05. NC, negative control.
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