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Article Open Access

STAT1 accelerates cutaneous melanoma progression through TUBB4A expression regulation

  • Authors:
    • Rongxin Zhao
    • Kexin Fang
    • Xiaofei Zhang
    • Hongchao Li
  • View Affiliations / Copyright

    Affiliations: Department of Dermatology, Pudong New Area People's Hospital, Shanghai 201200; P.R. China, Department of Dermatology, 921st Hospital of Joint Logistics Support Force People's Liberation Army of China, The Second Affiliated Hospital of Hunan Normal University, Changsha, Hunan 410073, P.R. China, Shanghai Xinmei Medical Beauty Outpatient Department, Shanghai 200031, P.R. China, Department of Head and Neck Surgery, The Second Hospital of Zhuzhou City, Zhuzhou, Hunan 412000, P.R. China
    Copyright: © Zhao et al. This is an open access article distributed under the terms of Creative Commons Attribution License.
  • Article Number: 118
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    Published online on: February 13, 2026
       https://doi.org/10.3892/mmr.2026.13828
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Abstract

The present research explored the contributions of signal transducer and activator of transcription 1 (STAT1) and tubulin β4A (TUBB4A) in melanoma pathogenesis, focusing on their roles in modulating cellular proliferation, motility and apoptotic pathways. The goal of the study was to establish foundational evidence of the role of these proteins in melanoma for the development of precision therapeutic interventions. Gene silencing approaches were utilized to suppress STAT1 expression, while TUBB4A overexpression was achieved both in vitro and in a murine xenograft model. Cellular proliferation was evaluated via Cell Counting Kit‑8 and colony formation assay, whereas migration capacity was assessed using Transwell migration assays. Apoptotic activity was quantified by flow cytometry using Annexin V‑FITC and PI staining. Western blot analysis was performed to measure the protein expression levels of STAT1 and TUBB4A. STAT1 downregulation led to impaired proliferation and motility in A375 and RPMI‑7951 melanoma cell lines, concomitant with increased apoptotic rates. These phenotypic changes were partially reversed following TUBB4A overexpression. In vivo experiments demonstrated significantly smaller tumor volumes in STAT1 knockdown xenografts, although TUBB4A overexpression partially restored neoplastic growth. STAT1 drove melanoma progression by upregulating TUBB4A, which acted as a downstream signaling mediator. The ability of TUBB4A to counteract STAT1 inhibition effects suggested that targeting this regulatory axis represents a potential therapeutic strategy. The findings of the present study contributed novel mechanistic insights that may facilitate the development of innovative melanoma treatment modalities.
View Figures

Figure 1

Bioinformatic analysis demonstrates
elevated TUBB4A expression in melanoma, correlating with poor
prognosis. (A) Data from the Gene Expression Profiling Interactive
Analysis 2 database revealed that TUBB4A was markedly altered
across various cancers, with a marked upregulation in SKCM. (B)
Further comparison of TUBB4A expression in melanoma tissues vs.
control tissues highlighted a significant increase in TUBB4A levels
in melanoma samples. (C) Analysis of TUBB4A expression across
different melanoma stages revealed no significant variation in
expression levels. Kaplan-Meier survival analysis measuring TUBB4A
expression against (D) overall survival and (E) disease-free
survival showed that high TUBB4A expression was significantly
associated with poorer overall survival and disease-free survival
outcomes, compared with patients demonstrating low TUBB4A
expression. *P<0.05. SKCM, skin cutaneous melanoma; TUBB4A,
tubulin β4A; TPM, transcripts per million; HR, hazard ratio.

Figure 2

STAT1 and TUBB4A expression levels
are significantly correlated and upregulated in melanoma. (A)
Analysis of data in the Gene Expression Profiling Interactive
Analysis 2 database revealed a significant elevation of STAT1
expression in melanoma tissues. Quantification of (B) STAT1 and (C)
TUBB4A mRNA levels in 31 paired melanoma and normal tissue samples
from patients with SKCM demonstrated a significant upregulation of
both STAT1 and TUBB4A expression in melanoma tissues. (D)
Correlation analysis of STAT1 and TUBB4A mRNA levels in melanoma
samples showed a significant positive correlation. Comparative
analysis of (E) STAT1 and (F) TUBB4A mRNA expression in normal HEM
and melanoma cell lines revealed significantly higher expression in
the melanoma cell lines. *P<0.05 between groups; **P<0.01
compared with HEM cells. HEM, human epidermal melanocytes; STAT1,
signal transducer and activator of transcription 1; TUBB4A, tubulin
β4A; TPM, transcripts per million; SKCM, skin cutaneous
melanoma.

Figure 3

STAT1 knockdown significantly reduces
melanoma cell viability and proliferation. STAT1 mRNA expression
was analyzed in (A) A375 and (B) RPMI-7951 melanoma cells
post-STAT1 knockdown using siRNA. (C) Western blotting was used to
assess STAT1 protein levels in melanoma cell lines following
siRNA-mediated knockdown. Cell viability was evaluated after STAT1
knockdown in (D) A375 and (E) RPMI-7951 cells using Cell Counting
Kit-8 assays. (F) Colony formation assays were performed to assess
the proliferative capacity of cells after STAT1 knockdown. (G)
Quantification of colony formation assay results. **P<0.01 vs.
si-NC group. siRNA, small interfering RNA; STAT1, signal transducer
and activator of transcription 1; si-STAT1, siRNA targeting STAT1;
si-NC, negative control siRNA.

Figure 4

STAT1 knockdown significantly
promotes melanoma cell apoptosis and inhibits migration. (A and B)
Apoptosis was assessed and quantified in A375 and RPMI-7951 cells
following STAT1 knockdown using flow cytometry. (C) Transwell
migration assays were conducted to evaluate cell migration
following STAT1 knockdown. Magnification, ×200. (D) Quantification
of migration capacity in A375 and RPMI-7951 cells following STAT1
knockdown. *P<0.05 vs. si-NC. STAT1, signal transducer and
activator of transcription 1; si-STAT1, small interfering RNA
targeting STAT1; si-NC, negative control small interfering RNA.

Figure 5

STAT1 regulates TUBB4A expression at
the transcription level. (A) STAT1 mRNA levels were measured in
A375 and RPMI-7951 cells after STAT1 knockdown via transfection
with different siRNA sequences. (B) TUBB4A mRNA levels were
measured in A375 and RPMI-7951 cells after STAT1 knockdown via
transfection with different siRNA sequences. (C) Specific fragments
of the TUBB4A promoter region were cloned into the luciferase
reporter plasmids upstream of the firefly luciferase gene. (D)
Transcriptional activity of various TUBB4A promoter fragments was
analyzed by luciferase reporter assay in 293T cells, with the
−1,783 and −1,771 fragments exhibiting the highest activity. (E)
STAT1 siRNA-mediated knockdown significantly reduced STAT1 mRNA
levels in A375 cells. (F) STAT1 knockdown significantly reduced the
luciferase activity of the −1,783 fragment of the TUBB4A promoter,
but not the −1,771 fragment. (G) Chromatin immunoprecipitation
assays were performed in A375 and RPMI-7951 cells targeting the
−1,783 binding site in the TUBB4A promoter region. Quantitative PCR
provided evidence of STAT1 binding to this region. Genomic DNA
input was set to 100%. **P<0.01 vs. si-NC;
##P<0.01 vs. PGL3; &&P<0.01 vs.
IgG. STAT1, signal transducer and activator of transcription 1;
siRNA, small interfering RNA; si-NC, negative control siRNA;
si-STAT1, siRNA targeting STAT1; si-STAT1-1, siRNA targeting STAT1
sequence 1; si-STAT1-2, siRNA targeting STAT1 sequence 2; TUBB4A,
tubulin β4A; PGL3, promoter-gluc luciferase 3; LUC, firefly
luciferase gene.

Figure 6

TUBB4A overexpression mitigates the
effects of STAT1 knockdown on cell viability and proliferation.
TUBB4A mRNA levels were measured in (A) A375 and (B) RPMI-7951
cells following TUBB4A overexpression mediated by a lentiviral
vector. (C) TUBB4A protein expression was analyzed after its
overexpression. Combined STAT1 knockdown and TUBB4A overexpression
transfections were performed, followed by a western blot analysis
of TUBB4A protein levels in (D) A375 and (E) RPMI-7951 cells. Cell
viability was assessed via Cell Counting Kit-8 assays following
combined STAT1 knockdown and TUBB4A overexpression in (F) A375 and
(G) RPMI-7951 cells. (H) Colony formation assays were performed to
assess the proliferative capacity of cells after STAT1 knockdown
and TUBB4A overexpression. (I) Quantification of colony formation
assay results. **P<0.01 vs. Ov-NC; ##P<0.01 vs.
si-STAT1. TUBB4A, tubulin β4A; STAT1, signal transducer and
activator of transcription 1; si-NC, negative control small
interfering RNA; si-STAT1, small interfering RNA targeting STAT1;
Ov-NC, negative control lentiviral overexpression vector;
Ov-TUBB4A, lentiviral vector for TUBB4A overexpression.

Figure 7

TUBB4A overexpression reverses the
effects of STAT1 knockdown on apoptosis, migration and tumor
growth. (A-D) Apoptosis and migration were evaluated in A375 and
RPMI-7951 cells following STAT1 knockdown and TUBB4A
overexpression. Magnification, ×200. (E) Representative images of
isolated xenograft tumors in mice. Tumor volumes were measured in
nude mice injected subcutaneously with 2×106 A375 cells
that had been subject to STAT1 knockdown and TUBB4A overexpression.
(F) Quantification of mouse tumor volumes showed that STAT1
knockdown significantly suppressed tumor growth, whereas TUBB4A
overexpression reversed this inhibitory effect.
##P<0.01 vs. si-STAT1. TUBB4A, tubulin β4A; si-NC,
negative control small interfering RNA; Ov-TUBB4A, lentiviral
vector for TUBB4A overexpression.
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Copy and paste a formatted citation
Spandidos Publications style
Zhao R, Fang K, Zhang X and Li H: STAT1 accelerates cutaneous melanoma progression through TUBB4A expression regulation. Mol Med Rep 33: 118, 2026.
APA
Zhao, R., Fang, K., Zhang, X., & Li, H. (2026). STAT1 accelerates cutaneous melanoma progression through TUBB4A expression regulation. Molecular Medicine Reports, 33, 118. https://doi.org/10.3892/mmr.2026.13828
MLA
Zhao, R., Fang, K., Zhang, X., Li, H."STAT1 accelerates cutaneous melanoma progression through TUBB4A expression regulation". Molecular Medicine Reports 33.4 (2026): 118.
Chicago
Zhao, R., Fang, K., Zhang, X., Li, H."STAT1 accelerates cutaneous melanoma progression through TUBB4A expression regulation". Molecular Medicine Reports 33, no. 4 (2026): 118. https://doi.org/10.3892/mmr.2026.13828
Copy and paste a formatted citation
x
Spandidos Publications style
Zhao R, Fang K, Zhang X and Li H: STAT1 accelerates cutaneous melanoma progression through TUBB4A expression regulation. Mol Med Rep 33: 118, 2026.
APA
Zhao, R., Fang, K., Zhang, X., & Li, H. (2026). STAT1 accelerates cutaneous melanoma progression through TUBB4A expression regulation. Molecular Medicine Reports, 33, 118. https://doi.org/10.3892/mmr.2026.13828
MLA
Zhao, R., Fang, K., Zhang, X., Li, H."STAT1 accelerates cutaneous melanoma progression through TUBB4A expression regulation". Molecular Medicine Reports 33.4 (2026): 118.
Chicago
Zhao, R., Fang, K., Zhang, X., Li, H."STAT1 accelerates cutaneous melanoma progression through TUBB4A expression regulation". Molecular Medicine Reports 33, no. 4 (2026): 118. https://doi.org/10.3892/mmr.2026.13828
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