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Background <p>Despite the emerging role of cuproptosis in cancer biology, the regulatory mechanism by which ubiquitination modulates this process through MKRN1 and SLC31A1 in colorectal cancer (CRC) progression remains unclear. This study aimed to elucidate the functional relationship between MKRN1, SLC31A1, and cuproptosis in CRC.</p> Methods <p>Stable MKRN1-knockdown and MKRN1-overexpressing colorectal cancer (CRC) cell lines were established.&#xa0;Cuproptosis was induced using elesclomol and copper sulfate, and assessed by cell viability assays,, measurement of intracellular copper levels, and western blot analysis of hallmark proteins (HSP70 and DLAT). The MKRN1-SLC31A1 interaction was examined by co-immunofluorescence, ubiquitination assays (employing MG132, CHX, and linkage-specific ubiquitin mutants), and quantitative real-time PCR (q-PCR). Mitochondrial function was evaluated via JC-1 staining and transmission electron microscopy. Rescue experiments were conducted using dual genetic manipulation of MKRN1 and SLC31A1. Finally, in vivo validation was performed using a xenograft model in NOD-SCID mice.</p> Results <p>MKRN1 overexpression significantly attenuated cuproptosis agonist-induced cytotoxicity and reduced intracellular copper accumulation. Mechanistically, MKRN1 directly interacts with SLC31A1 and promotes its polyubiquitination and subsequent proteasomal degradation. This degradation prevents copper overload, helping to maintain mitochondrial integrity.(as evidenced by maintained mitochondrial membrane potential and Tomm20 expression) and suppressing key cuproptosis hallmarks, including HSP70 induction and DLAT oligomerization. Genetic rescue experiments confirmed that SLC31A1 knockdown reversed the cuproptosis sensitivity conferred by MKRN1 depletion. Consistently, MKRN1 overexpression promoted tumor growth in vivo while concurrently suppressing cuproptosis.</p> Conclusion <p>In summary, MKRN1 suppresses copper-induced cell death in colorectal cancer by promoting the ubiquitination and proteasomal degradation of the copper transporter SLC31A1, thus limiting copper accumulation and mitochondrial toxicity. This mechanism confers resistance to copper-induced cell death and facilitates tumor progression, establishing the MKRN1/SLC31A1 axis as a promising therapeutic target.</p>

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MKRN1 suppresses cuproptosis in colorectal cancer through ubiquitin-dependent regulation of SLC31A1

  • 期刊论文

  • 出版模式：

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  • 发表日期：

    2026年7月7日

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  • Huali Luo,
  • Jiaxin Hu,
  • Xiaojing Lin,
  • Shu Zhao,
  • Mengxing Li,
  • Qinshan Li

摘要

Background

Despite the emerging role of cuproptosis in cancer biology, the regulatory mechanism by which ubiquitination modulates this process through MKRN1 and SLC31A1 in colorectal cancer (CRC) progression remains unclear. This study aimed to elucidate the functional relationship between MKRN1, SLC31A1, and cuproptosis in CRC.

Methods

Stable MKRN1-knockdown and MKRN1-overexpressing colorectal cancer (CRC) cell lines were established. Cuproptosis was induced using elesclomol and copper sulfate, and assessed by cell viability assays,, measurement of intracellular copper levels, and western blot analysis of hallmark proteins (HSP70 and DLAT). The MKRN1-SLC31A1 interaction was examined by co-immunofluorescence, ubiquitination assays (employing MG132, CHX, and linkage-specific ubiquitin mutants), and quantitative real-time PCR (q-PCR). Mitochondrial function was evaluated via JC-1 staining and transmission electron microscopy. Rescue experiments were conducted using dual genetic manipulation of MKRN1 and SLC31A1. Finally, in vivo validation was performed using a xenograft model in NOD-SCID mice.

Results

MKRN1 overexpression significantly attenuated cuproptosis agonist-induced cytotoxicity and reduced intracellular copper accumulation. Mechanistically, MKRN1 directly interacts with SLC31A1 and promotes its polyubiquitination and subsequent proteasomal degradation. This degradation prevents copper overload, helping to maintain mitochondrial integrity.(as evidenced by maintained mitochondrial membrane potential and Tomm20 expression) and suppressing key cuproptosis hallmarks, including HSP70 induction and DLAT oligomerization. Genetic rescue experiments confirmed that SLC31A1 knockdown reversed the cuproptosis sensitivity conferred by MKRN1 depletion. Consistently, MKRN1 overexpression promoted tumor growth in vivo while concurrently suppressing cuproptosis.

Conclusion

In summary, MKRN1 suppresses copper-induced cell death in colorectal cancer by promoting the ubiquitination and proteasomal degradation of the copper transporter SLC31A1, thus limiting copper accumulation and mitochondrial toxicity. This mechanism confers resistance to copper-induced cell death and facilitates tumor progression, establishing the MKRN1/SLC31A1 axis as a promising therapeutic target.