Perera, H. | Search bioRxiv

bioRxiv2026

Mechanism of K63-linked polyubiquitin recognition and cleavage by the BRCA1-A complex

Deubiquitylases modulate cellular processes by cleaving monoubiquitin or polyubiquitin chains. The ARISC-RAP80 complex partners with BRCA1-BARD1 to form the BRCA1-A super-complex, which recognizes K63-linked ubiquitin chains at DNA damage sites. ARISC-RAP80 contains multiple ubiquitin-binding sites, yet how these influence recognition and cleavage of K63-polyubiquitylated substrates remains unknown. We discover that a composite three-subunit interface allows ARISC-RAP80 to position K63-linked polyubiquitin chains in its catalytic site. Substrate recognition is further supported by RAP80 and non-catalytic ubiquitin-binding sites that impose a compact conformation to K63-polyubiquitylated substrates. This mechanism exploits the inherent flexibility of long ubiquitin chains and differs considerably from other deubiquitylases. Structure-guided mutagenesis validate ubiquitin chain interactions, and cell-based assays demonstrate a functional role of the observed interfaces in chromatin recruitment. Our findings define mechanisms of polyubiquitin chain decoding and cleavage by ARISC-RAP80, linking ubiquitin reading and erasing functions to BRCA1-A mediated DNA damage responses.

biochemistry↗

bioRxiv2025

Preclinical Study of Cannflavins A and B Action Against Glioblastoma Cells

AO_SCPLOWBSTRACTC_SCPLOWO_ST_ABSBackgroundC_ST_ABSFlavonoids represent a large group of naturally occurring polyphenolic compounds, many of which have been found to produce valuable biological outcomes, including action against cancer cells. Glioblastoma multiforme (GBM) is an aggressive type of brain tumor that is associated with a poor prognosis and has limited treatment options. Previous findings suggest that cannflavins can produce promising effects for pancreatic and bladder cancers, but the efficacy of these phytochemicals in attacking brain tumour cells remains unknown. Our study evaluates the potential of cannflavin A and cannflavin B against GBM cells using a range of in vitro approaches. ResultsWe conducted experiments using A-172 and U-87 GBM lines to assess the impact of cannflavins A and B on cell viability, cycle, migration, and invasion capacity. Our results revealed a consistent dose-dependent decrease in cell viability in both lines after the addition of cannflavin B to the culture media. Interestingly, we found that chrysoeriol (the non-prenylated synthesis precursor of cannflavins in the Cannabis sativa plant) only has a limited adverse effect on survival using the same approach, while techtochrysin (an O-methylated flavone) has none. Using time-lapse live-cell imaging and a scratch assay, we also show that cannflavins can inhibit tumor cell migration at concentrations below those that produce significant cell death. Finally, we found that cannflavin B exhibits anti-migratory and anti-invasive properties in transwell and tumorsphere assays, underscoring its multifaceted therapeutic potential. ConclusionThese findings suggest that cannflavin B holds promise as a therapeutic agent in the treatment of GBM. HO_SCPLOWIGHLIGHTSC_SCPLOWO_LICannflavin B but not cannflavin A limits the viability of A-172 and U-87 GBM cells in a concentration- and time-dependent manner. C_LIO_LIUsing time-lapse live-cell imaging and a scratch assay, we provide evidence that cannflavins A and B can inhibit GBM cell migration at concentrations below those that result in a significant reduction in cell viability. C_LIO_LICannflavin B exhibits robust anti-migratory and anti-invasive properties in transwell and tumorsphere assays, underscoring its multifaceted therapeutic potential. C_LIO_LIAlthough previous work had reported the capacity of cannflavins A and B to interfere with TrkB and downstream the MAPK/AKT signaling pathways in mouse primary neurons, the effect of those molecules is limited in GBMs, suggesting that other targets are engaged to produce the cellular effects. C_LI

pharmacology and toxicology↗