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Gene & Protein in Disease m1A-mediated ESCCAL-1 promotes ESCA stemness

A D

B
E F I

C G H

Figure 2. ESCCAL-1 is essential for maintaining ESCA stemness. (A) The transcription levels of ESCCAL-1 in Het-1A, TE1, KYSE70, and EC1 cells were
detected by qRT-PCR, n = 4. (B and C) The knockdown (B) and overexpression (C) effects of ESCCAL-1 were tested by qRT-PCR, n = 3. **P < 0.01,
***P < 0.001. (D and F) The effects of deletion or overexpression of ESCCAL-1 on the stemness of ESCA cells were evaluated by tumor sphere formation
assay, n = 3. *P < 0.05, **P < 0.01. Scale bar = 100 μm. In panel (D), TE1 cells were transfected with sh-AL1, and KYSE70 were transfected with OE-AL1.
(G–I) A colony formation assay was used to observe the effects of knockdown (n = 2) or upregulation (n = 3) of ESCCAL-1 on ESCA growth. *P < 0.05. In
panel (G), TE1 cells were transfected with sh-AL1, and KYSE70 were transfected with OE-AL1.
ESCCAL-1: Esophageal squamous cell carcinoma associated long non-coding RNA 1; ESCA: Esophageal cancer; qRT-PCR: quantitative reverse
transcription polymerase chain reaction.

(Figure 3D). These results suggest that ESCCAL-1 may tumor types, such as liver cancer and cervical cancer [13-15] .
be involved in the stemness maintenance of ESCA by To explore whether m A may regulate ESCCAL-1, we first
1
regulating the expression of CD44 and KLF4. used the GEPIA database to analyze the expression of m A
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demethylase ALKBH3 in ESCA and its correlation with
3.4. RNA m A demethylase ALKBH3 mediates ESCCAL-1. Surprisingly, we found that ALKBH3 and
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ESCCAL-1 expression in ESCA ESCCAL-1 showed increased expression and significant
Given that ESCCAL-1 is upregulated in ESCA, we positive correlation in 182 ESCA samples (Figure 4A and B),
intended to investigate the upstream mechanism that implying that ALKBH3 may be involved in the regulation
causes its dysregulation. As a novel RNA modification of ESCCAL-1 and may play a role in ESCA.
mode, ALKBH3-mediated m A modification has been Data from qRT-PCR showed that the expression level
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reported recently to play a vital regulatory role in some of ALKBH3 in three ESCA cell lines (TE1, KYSE70,

Volume 2 Issue 1 (2023) 5 https://doi.org/10.36922/gpd.305