Biology subjects

Flynn, R. L.

Publications and source records attributed to Flynn, R. L..

3 recordsLinked to original sources

bioRxiv2026

Bloom syndrome helicase is required for efficient HIV-1 reverse transcription in macrophages

The induction of DNA damage by HIV-1 prior to integration suggests a function for DNA damage responses (DDR) during early infection, however what this role is remains incompletely understood. Initial experiments, using specific inhibitors for DDR pathways demonstrate that both ATM and ATR are necessary for efficient HIV-1 infection of macrophages with ATM acting at the late reverse transcription step. To identify DDR factors associated with ATM/ATR pathways that influence HIV-1 infection, a CRISPR knockout screen using a DDR-focused sgRNA library was performed. Approximately 30 DDR genes that impacted HIV-1 infection were identified with 13 factors that facilitated HIV-1 infection and 17 DDR factors that restrict HIV-1 infection. Several hits were factors associated with the Fanconi anemia pathway, such as BTR complex proteins, including the RecQ helicase Bloom syndrome helicase. BLM was specifically demonstrated to enhance HIV-1 infection and replication with knockdown of BLM expression diminishing integration and the establishment of intact HIV-1 proviruses in macrophages by 50%. BLM is associated with HIV-1 late reverse transcription intermediates, the step that proceeds HIV-1 integration. These findings identify BLM as a DDR host factor that promotes early HIV-1 infection by facilitating completion of reverse transcription and subsequent integration. Significance StatementHIV-1 infection elicits cellular DNA damage responses although the role of DNA damage in HIV-1 infection has not been fully characterized. Our study identifies specific DNA Damage factors that facilitate or restrict HIV infection. In particular, we show the RecQ helicase Bloom syndrome helicase (BLM) is a mediator of HIV-1 reverse transcription and integration in macrophages. This work highlights a functional interface between DNA damage repair pathways and HIV-1 integration suggesting that targeting select host DNA damage response factors can limit HIV-1 infection and persistence.

microbiology↗

bioRxiv2026

Functional inactivation of the telomerase chaperone TCAB1 primes cells for the activation of ALT in osteosarcoma

Activation of the alternative lengthening of telomeres (ALT) pathway accounts for cellular immortalization in 75% of pediatric osteosarcoma. ALT does not rely on a single enzyme but instead, catalyzes telomere elongation via homologous recombination. There has been steady progress in defining the mechanisms that regulate the ALT pathway. However, the spectrum of genetic mutations that underlie activation of ALT remains unclear. Osteosarcomas, like many cancers, frequently harbor inactivating mutations in the tumor suppressor gene TP53. However, instead of single nucleotide variants that lead to expression of mutant TP53 protein, osteosarcoma tumors often acquire unique structural variations within the first intron of the TP53 gene leading to complete gene inactivation. TP53 is located on chromosome 17p13.1 in a head-to-head orientation and partially overlapping with the gene WRAP53 (WD repeat containing antisense to TP53). WRAP53, also known as TCAB1, is an RNA chaperone that is an essential component of the telomerase holoenzyme. TCAB1 functions to facilitate trafficking of the telomerase RNA (hTR) within the nucleus to ensure assembly and localization of the telomerase enzyme to telomere ends to promote telomere elongation. Loss of TCAB1 function abolishes telomerase activity, driving progressive telomere attrition. Here, using whole-genome sequencing of osteosarcoma samples we identified SVs within the TP53 gene that not only compromise TP53, but also inactivate TCAB1. These TCAB1 SVs were prevalent in approximately 40% of ALT positive osteosarcoma tumors suggesting that functional inactivation of the telomerase holoenzyme may be an early and previously unrecognized event contributing to the activation of the ALT pathway.

cancer 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↗