Characterization of tumor suppressor SMAD4 as a novel regulator of genome instability in human cancer

by Atmika Paul
Doctoral thesis
Date of Examination:2024-11-20
Date of issue:2025-09-24
Advisor:Prof. Dr. Holger Bastians
Referee:Prof. Dr. Holger Bastians
Referee:Prof. Dr. Jürgen Wienands
Referee:Sergio Pérez Acebrón
Persistent Address: http://dx.doi.org/10.53846/goediss-11516

 

 

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Abstract

English

Chromosomal instability (CIN) is characterized by an increased rate of generation of chromosomal alterations and is a hallmark of human cancer. CIN is strongly associated with genomic heterogeneity, therapeutic resistance, and poor clinical outcomes. CIN manifests in two distinct forms: whole-chromosome instability (W-CIN) and structural chromosomal instability (S-CIN). W-CIN is marked by numerical chromosomal alterations and can be driven by mitotic defects, particularly by abnormal microtubule dynamics in mitosis. In contrast, S-CIN involves structural rearrangements that can result from DNA damage, DNA repair impairment or S phase-related replication stress. Recent studies have reported that replication stress not only causes structural chromosome abnormalities but also increases mitotic microtubule dynamics, thereby contributing to the development of numerical chromosome aberrations and W-CIN. Despite the widespread prevalence of CIN, the cancer-specific drivers of CIN and replication stress remain little understood. In this work, I identified the tumor suppressor SMAD4 as a key molecular trigger for both W-CIN and S-CIN. My results show that SMAD4 loss in colorectal cancer cells inactivates the BMP/SMAD4 signaling pathway, downregulating its transcriptional target gene, ID3. Loss of SMAD4 or ID3 impairs replication fork progression, indicative of replication stress, resulting subsequently in elevated mitotic microtubule polymerization rates and mis-segregation of chromosomes during anaphase, ultimately driving both structural and numerical chromosomal aberrations. Furthermore, SMAD4 or ID3 inactivation reduces homologous recombination DNA repair efficiency, associated with increased DNA damage. Notably, re-expressing ID3 in SMAD4-deficient cancer cells restores proper DNA repair, DNA replication, mitotic abnormalities and suppresses genomic instability, indicating that SMAD4 inactivation drives CIN through loss of ID3. Interestingly, depletion of key homologous recombination DNA repair components, such as BRCA1 and RAD51, also causes replication stress, abnormal microtubule growth rates, and chromosome segregation defects, suggesting a principal causal relationship between impaired homologous recombination machinery, replication stress and mitotic errors. In conclusion, this study revealed that inactivation of the key tumor suppressor gene SMAD4 triggers replication stress and chromosomal instability. The results presented here provide important new insights into the mechanisms of generation of genome instability in human cancer and offer potential therapeutic strategies for targeting chromosomally unstable SMAD4-deficient cancers.
Keywords: Cancer; Chromosomal instability; Replication stress; DNA damage repair; SMAD4