Targeting N-Myc in neuroblastoma with selective Aurora kinase A degraders.

The N-Myc transcription factor, encoded by MYCN, is a mechanistically validated, yet challenging, target for neuroblastoma (NB) therapy development. In normal neuronal progenitors, N-Myc undergoes rapid degradation, while, in MYCN-amplified NB cells, Aurora kinase A (Aurora-A) binds to and stabilizes N-Myc, resulting in elevated protein levels. Here, we demonstrate that targeted protein degradation of Aurora-A decreases N-Myc levels.

Structural Insights into the Assembly and Regulation of 2'- RNA Methylation by SARS-CoV-2 nsp16/nsp10.

2'- -ribose methylation of the first transcribed base (adenine or A in SARS-CoV-2) of viral RNA mimics the host RNAs and subverts the innate immune response. How nsp16, with its obligate partner nsp10, assembles on the 5'-end of SARS-CoV-2 mRNA to methylate the A has not been fully understood. We present a ∼ 2.

Mammalian ZAP and KHNYN independently restrict CpG-enriched avian viruses.

Unlabelled: Zoonotic viruses are an omnipresent threat to global health. Influenza A virus (IAV) transmits between birds, livestock, and humans. Proviral host factors involved in the cross-species interface are well known.

Regulatory interactions between APOBEC3B N- and C-terminal domains.

APOBEC3B (A3B) is implicated in DNA mutations that facilitate tumor evolution. Although structures of its individual N- and C-terminal domains (NTD and CTD) have been resolved through X-ray crystallography, the full-length A3B (fl-A3B) structure remains elusive, limiting understanding of its dynamics and mechanisms. In particular, the APOBEC3B C-terminal domain (A3Bctd) active site is frequently closed in models and structures.

The Impact of Sugar Conformation on the Single-Stranded DNA Selectivity of APOBEC3A and APOBEC3B Enzymes.

The APOBEC3 family of polynucleotide cytidine deaminases has diverse roles as viral restriction factors and oncogenic mutators. These enzymes convert cytidine to uridine in single-stranded (ss)DNA, inducing genomic mutations that promote drug resistance and tumor heterogeneity. Of the seven human APOBEC3 members, APOBEC3A (A3A) and APOBEC3B (A3B) are most implicated in driving pro-tumorigenic mutations.

Differentiation signals induce APOBEC3A expression via GRHL3 in squamous epithelia and squamous cell carcinoma.

Two APOBEC DNA cytosine deaminase enzymes, APOBEC3A and APOBEC3B, generate somatic mutations in cancer, thereby driving tumour development and drug resistance. Here, we used single-cell RNA sequencing to study APOBEC3A and APOBEC3B expression in healthy and malignant mucosal epithelia, validating key observations with immunohistochemistry, spatial transcriptomics and functional experiments. Whereas APOBEC3B is expressed in keratinocytes entering mitosis, we show that APOBEC3A expression is confined largely to terminally differentiating cells and requires grainyhead-like transcription factor 3 (GRHL3).

RADD: A real-time FRET-based biochemical assay for DNA deaminase studies.

In recent years, the connection between APOBEC3 cytosine deaminases and cancer mutagenesis has become ever more apparent. This growing awareness and lack of inhibitory drugs has created a distinct need for biochemical tools that can be used to identify and characterize potential inhibitors of this family of enzymes. In response to this challenge, we have developed a Real-time APOBEC3-mediated DNA Deamination (RADD) assay.

A compact stem-loop DNA aptamer targets a uracil-binding pocket in the SARS-CoV-2 nucleocapsid RNA-binding domain.

SARS-CoV-2 nucleocapsid (N) protein is a structural component of the virus with essential roles in the replication and packaging of the viral RNA genome. The N protein is also an important target of COVID-19 antigen tests and a promising vaccine candidate along with the spike protein. Here, we report a compact stem-loop DNA aptamer that binds tightly to the N-terminal RNA-binding domain of SARS-CoV-2 N protein.

A comprehensive study of SARS-CoV-2 main protease (Mpro) inhibitor-resistant mutants selected in a VSV-based system.

Nirmatrelvir was the first protease inhibitor specifically developed against the SARS-CoV-2 main protease (3CLpro/Mpro) and licensed for clinical use. As SARS-CoV-2 continues to spread, variants resistant to nirmatrelvir and other currently available treatments are likely to arise. This study aimed to identify and characterize mutations that confer resistance to nirmatrelvir.

SARS-CoV-2 M inhibitor identification using a cellular gain-of-signal assay for high-throughput screening.

Severe Acute Respiratory Syndrome Coronavirus-2 (SARS-CoV-2, SARS2) is responsible for the COVID-19 pandemic and infections that continue to affect the lives of millions of people worldwide, especially those who are older and/or immunocompromised. The SARS2 main protease enzyme, M (also called 3C-like protease, 3CL), is a bona fide drug target as evidenced by potent inhibition with nirmatrelvir and ensitrelvir, the active components of the drugs Paxlovid and Xocova, respectively. However, the existence of nirmatrelvir and ensitrelvir-resistant isolates underscores the need to develop next-generation drugs with different resistance profiles and/or distinct mechanisms of action.

A high-throughput cell-based screening method for Zika virus protease inhibitor discovery.

Zika virus (ZIKV) continues to pose a significant global public health threat, with recurring regional outbreaks and potential for pandemic spread. Despite often being asymptomatic, ZIKV infections can have severe consequences, including neurological disorders and congenital abnormalities. Unfortunately, there are currently no approved vaccines or antiviral drugs for the prevention or treatment of ZIKV.

A real-time biochemical assay for quantitative analyses of APOBEC-catalyzed DNA deamination.

Over the past decade, the connection between APOBEC3 cytosine deaminases and cancer mutagenesis has become increasingly apparent. This growing awareness has created a need for biochemical tools that can be used to identify and characterize potential inhibitors of this enzyme family. In response to this challenge, we have developed a Real-time APOBEC3-mediated DNA Deamination assay.

A real-time biochemical assay for quantitative analyses of APOBEC-catalyzed DNA deamination.

Over the past decade, the connection between APOBEC3 cytosine deaminases and cancer mutagenesis has become increasingly apparent. This growing awareness has created a need for biochemical tools that can be used to identify and characterize potential inhibitors of this enzyme family. In response to this challenge, we have developed a Real-time APOBEC3-mediated DNA Deamination (RADD) assay.

APOBEC3 mutagenesis drives therapy resistance in breast cancer.

Acquired genetic alterations commonly drive resistance to endocrine and targeted therapies in metastatic breast cancer , however the underlying processes engendering these diverse alterations are largely uncharacterized. To identify the mutational processes operant in breast cancer and their impact on clinical outcomes, we utilized a well-annotated cohort of 3,880 patient samples with paired tumor-normal sequencing data. The mutational signatures associated with apolipoprotein B mRNA-editing enzyme catalytic polypeptide-like 3 (APOBEC3) enzymes were highly prevalent and enriched in post-treatment compared to treatment-naïve hormone receptor-positive (HR+) cancers.

Development of Allosteric Small Molecule APOBEC3B Inhibitors from Screening.

APOBEC3B cytosine deaminase contributes to the mutational burdens of tumors, resulting in tumor progression and therapy resistance. Small molecule APOBEC3B inhibitors have potential to slow or mitigate these detrimental outcomes. Through molecular dynamics (MD) simulations and computational solvent mapping analysis, we identified a novel putative allosteric pocket on the C-terminal domain of APOBEC3B (A3Bctd), and virtually screened the ChemBridge Diversity Set (N~110,000) against both the active and potential allosteric sites.

APOBEC shapes tumor evolution and age at onset of lung cancer in smokers.

APOBEC enzymes are part of the innate immunity and are responsible for restricting viruses and retroelements by deaminating cytosine residues. Most solid tumors harbor different levels of somatic mutations attributed to the off-target activities of APOBEC3A (A3A) and/or APOBEC3B (A3B). However, how APOBEC3A/B enzymes shape the tumor evolution in the presence of exogenous mutagenic processes is largely unknown.

Mesoscale DNA features impact APOBEC3A and APOBEC3B deaminase activity and shape tumor mutational landscapes.

Antiviral DNA cytosine deaminases APOBEC3A and APOBEC3B are major sources of mutations in cancer by catalyzing cytosine-to-uracil deamination. APOBEC3A preferentially targets single-stranded DNAs, with a noted affinity for DNA regions that adopt stem-loop secondary structures. However, the detailed substrate preferences of APOBEC3A and APOBEC3B have not been fully established, and the specific influence of the DNA sequence on APOBEC3A and APOBEC3B deaminase activity remains to be investigated.

Differentiation signals induce expression via GRHL3 in squamous epithelia and squamous cell carcinoma.

Two APOBEC (apolipoprotein-B mRNA editing enzyme catalytic polypeptide-like) DNA cytosine deaminase enzymes (APOBEC3A and APOBEC3B) generate somatic mutations in cancer, driving tumour development and drug resistance. Here we used single cell RNA sequencing to study and expression in healthy and malignant mucosal epithelia, validating key observations with immunohistochemistry, spatial transcriptomics and functional experiments. Whereas is expressed in keratinocytes entering mitosis, we show that expression is confined largely to terminally differentiating cells and requires Grainyhead-like transcription factor 3 (GRHL3).

Redefining high risk multiple myeloma with an APOBEC/Inflammation-based classifier.

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Deaminase deluge yields new opportunities for biotechnology and genome engineering.

In this issue of Molecular Cell, Vaisvila et al. report a tour de force functional characterization of a large and highly diverse set of polynucleotide cytosine deaminase (PCD) enzymes, which is already propelling new biotechnology applications.

Mutational impact of APOBEC3A and APOBEC3B in a human cell line and comparisons to breast cancer.

A prominent source of mutation in cancer is single-stranded DNA cytosine deamination by cellular APOBEC3 enzymes, which results in signature C-to-T and C-to-G mutations in TCA and TCT motifs. Although multiple enzymes have been implicated, reports conflict and it is unclear which protein(s) are responsible. Here we report the development of a selectable system to quantify genome mutation and demonstrate its utility by comparing the mutagenic activities of three leading candidates-APOBEC3A, APOBEC3B, and APOBEC3H.

Acute expression of human APOBEC3B in mice results in RNA editing and lethality.

Background: RNA editing has been described as promoting genetic heterogeneity, leading to the development of multiple disorders, including cancer. The cytosine deaminase APOBEC3B is implicated in tumor evolution through DNA mutation, but whether it also functions as an RNA editing enzyme has not been studied.

Results: Here, we engineer a novel doxycycline-inducible mouse model of human APOBEC3B-overexpression to understand the impact of this enzyme in tissue homeostasis and address a potential role in C-to-U RNA editing.