Biology subjects

Bracey, H.

Publications and source records attributed to Bracey, H..

3 recordsLinked to original sources

bioRxiv2024

Essential functions of Inositol hexakisphosphate (IP6) in Murine Leukemia Virus replication

We have investigated the function of inositol hexakisphosphate (IP6) and inositol pentakisphosphate (IP5) in the replication of murine leukemia virus (MLV). While IP6 is known to be critical for the life cycle of HIV-1, its significance in MLV remains unexplored. We find that IP6 is indeed important for MLV replication. It significantly enhances endogenous reverse transcription (ERT) in MLV. Additionally, a pelleting-based assay reveals that IP6 can stabilize MLV cores, thereby facilitating ERT. We find that IP5 and IP6 are packaged in MLV particles. However, unlike HIV-1, MLV depends upon the presence of IP6 and IP5 in target cells for successful infection. This IP6/5 requirement for infection is reflected in impaired reverse transcription observed in IP6/5-deficient cell lines. In summary, our findings demonstrate the importance of capsid stabilization by IP6/5 in the replication of diverse retroviruses; we suggest possible reasons for the differences from HIV-1 that we observed in MLV.

molecular biology↗

bioRxiv2023

The HIV-1 capsid serves as a nanoscale reaction vessel for reverse transcription

The viral capsid performs critical functions during HIV-1 infection and is a validated target for antiviral therapy. Previous studies have established that the proper structure and stability of the capsid are required for efficient HIV-1 reverse transcription in target cells. Moreover, it has recently been demonstrated that permeabilized virions and purified HIV-1 cores undergo efficient reverse transcription in vitro when the capsid is stabilized by addition of the host cell metabolite inositol hexakisphosphate (IP6). However, the molecular mechanism by which the capsid promotes reverse transcription is undefined. Here we show that wild type HIV-1 particles can undergo efficient reverse transcription in vitro in the absence of a membrane-permeabilizing agent. This activity, originally termed "natural endogenous reverse transcription" (NERT), depends on expression of the viral envelope glycoprotein during virus assembly and its incorporation into virions. Truncation of the gp41 cytoplasmic tail markedly reduced NERT activity, indicating that gp41 permits the entry of nucleotides into virions. Protease treatment of virions markedly reduced NERT suggesting the presence of a proteinaceous membrane channel. By contrast to reverse transcription in permeabilized virions, NERT required neither the addition of IP6 nor a mature capsid, indicating that an intact viral membrane can substitute for the function of the viral capsid during reverse transcription in vitro. Collectively, these results demonstrate that the viral capsid functions as a nanoscale container for reverse transcription during HIV-1 infection.

microbiology↗

bioRxiv2022

A small molecule that inhibits the evolution of antibiotic resistance

Antimicrobial resistance (AMR) rapidly develops against almost all available therapeutics. New antibiotics target essential processes in bacteria but fail to address the root of the problem: mutagenesis and evolution. We recently proposed that inhibiting the molecular mechanisms underlying bacterial evolution is the ultimate solution to preventing AMR development. Here, we describe the first compound that inhibits the occurrence and progression of AMR by directly targeting a highly conserved bacterial evolvability factor, Mfd. We previously found that this RNA polymerase-associated translocase is required for rapid AMR development across highly divergent pathogens. Through an in vivo screen, we identified 43 potential Mfd-inhibiting compounds. Here we present on target validation, biochemical characterization, and in vivo efficacy studies of a lead compound, referred to as ARM-1. ARM-1 binds Mfd and modulates its RNA polymerase interaction. Inhibition of Mfd activity by ARM-1 delays the development of mutations and resistance acquisition, both in pure culture and during infection. Importantly, our data show that this compound prevents the evolution of AMR across highly divergent pathogens, including Pseudomonas aeruginosa, Staphylococcus aureus, Listeria monocytogenes, and Salmonella enterica serovar Typhimurium. The novel compound we present here has the potential to develop into a clinically useful "anti-evolution" drug. This work demonstrates that the molecular mechanisms of evolution are pharmaceutically targetable, and that this strategy could help prevent AMR development.

microbiology↗