Biology subjects

Belmonte-Fernandez, A.

Publications and source records attributed to Belmonte-Fernandez, A..

3 recordsLinked to original sources

bioRxiv2025

hSpindly's dynamic controls SAC activity independently of the KBB pathway at unattached kinetochores

The Spindle Assembly Checkpoint (SAC) ensures that sister chromatids do not separate until all chromosomes are properly attached to spindle microtubules and correctly bi-oriented. SAC controls the metaphase-to-anaphase transition by inhibiting the Anaphase Promoting Complex/Cyclosome (APC/C) through the formation of the Mitotic Checkpoint Complex (MCC). A critical step in this process is the recruitment of the Mad1-Mad2 complex to unattached kinetochores. Two pathways are known to mediate this recruitment: the KBB pathway (KNL1-Bub3-Bub1) and the RZZ pathway (Rod- Zw10-Zwilch). Here, we demonstrate that hSpindly plays a central role in controlling the recruitment of the Mad1-Mad2 complex through the RZZ pathway, independently of the KBB pathway. We show that hSpindly is a dynamic protein that oligomerizes at unattached kinetochores. Importantly, we identify a specific residue, threonine 552, as critical for hSpindlys function and mobility. A non-phosphorylatable mutant (T552A) stabilizes hSpindly at kinetochores, impairs SAC signaling, and increases cellular resistance to antimitotic drugs. Altogether, our findings identify hSpindly as a novel dynamic modulator of SAC functionality via the RZZ pathway and highlight it as a potential therapeutic target for overcoming resistance to mitotic inhibitors.

cell biology↗

bioRxiv2025

βTrCP is involved in the localization of the MRN complex on chromatin to enhance DNA damage repair

Genomic instability underlies various diseases, including cancer. This instability arises from defects in critical cellular processes, particularly those involved in DNA damage repair. Therefore, a detailed understanding of these repair mechanisms is essential for developing strategies to prevent or diagnose such diseases. The MRN complex, composed of MRE11, NBS1, and RAD50, is among the earliest elements involved in detecting DNA damage. Upon detecting DNA breaks, this complex triggers a cascade of signaling events that regulate both cell cycle arrest and DNA repair. These signaling pathways are tightly controlled by various post-translational modifications, notably ubiquitination. Although several ubiquitin ligases have been implicated in different stages of the DNA damage response, our knowledge remains limited. In this study, we reveal that {beta}TrCP, a substrate-recognizing subunit of the SCF (SKP1/CUL1/F-box protein) ubiquitin ligase, interacts in vivo with the proteins of the MRN complex. These interactions occur in normally proliferating cells and are dependent on the GSK3 kinase. Moreover, we show that {beta}TrCP enhances the recruitment of the MRN complex to chromatin through MRE11, thereby promoting the efficient DNA damage repair. Hence, alterations in {beta}TrCP function affecting MRN dynamics could have severe consequences for the cell homeostasis.

cell biology↗

bioRxiv2024

Overexpression of betaTrCP1 elicits cell death in cisplatin-induced senescent cells

Senescence is a non-proliferative cellular state derived from aging or in response to exogenous insults, such as those that cause DNA damage. As a result of cancer treatments like cisplatin, certain tumor cells may undergo senescence. However, rather than being beneficial for patients, this is detrimental because these cells might proliferate again under specific conditions and, more importantly, because they synthesize and secrete molecules that promote the proliferation of nearby cells. Therefore, to achieve complete tumor remission, it is necessary to develop senolytic compounds to eliminate senescent cells. Here, we studied the role of {beta}TrCP1 in cell proliferation and senescence and found that lentiviral overexpression of {beta}TrCP1 induces the death of senescent cells obtained after cisplatin treatment in both two-dimensional cell cultures and tumorspheres. Mechanistically, we demonstrated that overexpression of {beta}TrCP1 triggers proteasome- dependent degradation of p21 CIP1, allowing damaged cells to progress through the cell cycle and consequently die. Furthermore, we identified nucleophosmin 1 (NPM1) as the intermediary molecule involved in the effect of {beta}TrCP1 on p21 CIP1. We determined that increased amounts of {beta}TrCP1 partially retains NPM1 in the nucleoli, preventing it from associating with p21 CIP1, thus leaving it unprotected from degradation by the proteasome. These results have allowed us to discover a potential new target for senolytic drugs, as retaining NPM1 in the nucleoli under senescent conditions induces cell death.

cell biology↗