Biology subjects

Manning, A. L.

Publications and source records attributed to Manning, A. L..

4 recordsLinked to original sources

bioRxiv2025

RB dependent transcriptional regulation at mitotic centromeres preserves genome stability

Transcripts derived from centromere repeats play a critical role in the localization and activity of kinetochore components during mitosis such that disruption of RNA polymerase II-dependent transcription compromises the fidelity of chromosome segregation. Here, we show that the retinoblastoma tumor suppressor protein (RB), a critical regulator of the G1/S cell cycle transition, additionally plays an important role in the regulation of centromere transcription during mitosis. We find that cells lacking RB experience increased RNA Polymerase II activity at mitotic centromeres and a corresponding increase in nascent RNA transcripts derived from centromere sequences. Together with high levels of centromere transcription and corresponding R-loop formation, RB-deficient cells exhibit centromere DNA breaks and local activation of ATR that correspond with increased centromere localization of Aurora B, destabilization of kinetochore-microtubule attachments, and an increase in anaphase defects. Importantly, reduction of DNA damage, ATR activity, and mitotic defects following inhibition of RNA Pol II, or targeted repression of centromere transcription through centromere tethering of Suv420H2, support that mitotic defects in RB-deficient cells are linked to centromere transcription.

cell biology↗

bioRxiv2023

RB loss sensitizes cells to replication-associated DNA damage by PARP inhibition

The retinoblastoma tumor suppressor protein (RB) interacts physically and functionally with a number of epigenetic modifying enzymes to control transcriptional regulation, respond to replication stress, promote DNA damage response and repair pathways, and regulate genome stability. To better understand how disruption of RB function impacts epigenetic regulation of genome stability and determine whether such changes may represent exploitable weaknesses of RB-deficient cancer cells, we performed an imaging-based screen to identify epigenetic inhibitors that promote DNA damage and compromise viability of RB-deficient cells. We found that loss of RB alone leads to high levels of replication-dependent poly-ADP ribosylation (PARylation) and that preventing PARylation through inhibition of PARP enzymes enables RB-deficient cells to progress to mitosis with unresolved replication stress and under-replicated DNA. These defects contribute to high levels of DNA damage, decreased proliferation, and compromised cell viability. We demonstrate this sensitivity is conserved across a panel of inhibitors that target both PARP1 and PARP2 and can be suppressed by re-expression of the RB protein. Together, these data indicate that inhibitors of PARP1 and PARP2 may be clinically relevant for RB-deficient cancers.

cell biology↗

bioRxiv2020

Whole genome doubling confers unique genetic vulnerabilities on tumor cells

Whole genome doubling (WGD) occurs early in tumorigenesis and generates genetically unstable tetraploid cells that fuel tumor development. Cells that undergo WGD (WGD+) must adapt to accommodate their abnormal tetraploid state; however, the nature of these adaptations, and whether they confer vulnerabilities that can subsequently be exploited therapeutically, is unclear. Using sequencing data from [~]10,000 primary human cancer samples and essentiality data from [~]600 cancer cell lines, we show that WGD gives rise to common genetic traits that are accompanied by unique vulnerabilities. We reveal that WGD+ cells are more dependent on spindle assembly checkpoint signaling, DNA replication factors, and proteasome function than WGD- cells. We also identify KIF18A, which encodes for a mitotic kinesin, as being specifically required for the viability of WGD+ cells. While loss of KIF18A is largely dispensable for accurate chromosome segregation during mitosis in WGD- cells, its loss induces dramatic mitotic errors in WGD+ cells, ultimately impairing cell viability. Collectively, our results reveal new strategies to specifically target WGD+ cancer cells while sparing the normal, non-transformed WGD- cells that comprise human tissue.

cancer biology↗

bioRxiv2018

Aurora A inhibition limits centrosome clustering and promotes mitotic catastrophe in cells with supernumerary centrosomes

The presence of supernumerary centrosomes is prevalent in cancer, where they promote the formation of transient multipolar mitotic spindles. Active clustering of supernumerary centrosomes enables the formation of a functional bipolar spindle that is competent to complete a bipolar division. Disruption of spindle pole clustering in cancer cells promotes multipolar division and generation of non-proliferative daughter cells with compromised viability. Hence molecular pathways required for spindle pole clustering in cells with supernumerary centrosomes, but dispensable in normal cells, are promising therapeutic targets. Here we demonstrate that Aurora A kinase activity is required for spindle pole clustering in cells with extra centrosomes. While cells with two centrosomes are ultimately able to build a bipolar spindle and proceed through a normal cell division in the presence of Aurora A inhibition, cells with supernumerary centrosomes form multipolar and disorganized spindles that are not competent for chromosome segregation. Instead, following a prolonged mitosis, these cells experience catastrophic divisions that result in grossly aneuploid, and non-proliferative daughter cells. Aurora A inhibition in a panel of Acute Myeloid Leukemia cancer cells has a similarly disparate impact on cells with supernumerary centrosomes, suggesting that centrosome number and spindle polarity may serve as predictive biomarkers for response to therapeutic approaches that target Aurora A kinase function.

cancer biology↗