# MIB2‐Mediated SUZ12 Ubiquitination Regulates Clonal Proliferation in Patients With Paroxysmal Nocturnal Haemoglobinuria

## TL;DR

This study shows that MIB2 regulates PNH clone proliferation by ubiquitinating SUZ12, affecting PRC2 stability and cell growth.

## Contribution

The novel finding is that MIB2 mediates SUZ12 ubiquitination, influencing PNH clonal expansion through PRC2 regulation.

## Key findings

- MIB2 physically interacts with SUZ12 in PIGA-KO K562 cells.
- MIB2 knockdown reduces SUZ12 protein levels and enhances its ubiquitination.
- MIB2 knockdown decreases PNH cell proliferation and increases apoptosis.

## Abstract

Secondary gene mutations are one of the main mechanisms underlying paroxysmal nocturnal haemoglobinuria (PNH) clone proliferation. Our previous studies showed that SUZ12 participates in PNH clone proliferation by regulating H3K27me3. However, the mechanism underlying the SUZ12 function remains unclear. Immunoprecipitation and mass spectrometry were used to identify SUZ12 interacting proteins in PIGA‐KO K562 cells. Furthermore, RNA‐seq was used to explore the signalling pathways. Finally, colony formation assays, western blotting, and flow cytometry were performed to determine the proliferative ability of the cells. We identified 59 potential proteins that interact with SUZ12 and revealed a physical interaction between MIB2 and SUZ12 exclusively in the K562‐KO cell line. Furthermore, we emphasise the vital involvement of the MIB/HERC and ZZ‐type domains in the physical association between MIB2 and SUZ12. After MIB2 knockdown, SUZ12 protein decreased while the ubiquitination levels of SUZ12 were enhanced. Additionally, PRC2‐related target genes were upregulated in the siMIB2 group. SUZ12 and H3K27me3 expression levels and cell proliferation significantly decreased after MIB2 knockdown, whereas cell apoptosis significantly increased. MIB2 protein levels are also elevated in patients with PNH. In conclusion, MIB2 affects the stability of the PRC2 complex by mediating SUZ12 ubiquitination, which in turn regulates PNH clone proliferation.