Directed evolution of a family of AAV capsid variants enabling potent muscle-directed gene delivery across species

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An NK-like CAR T cell transition in CAR T cell dysfunction

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Genome-wide programmable transcriptional memory by CRISPR-based epigenome editing

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Structural Insight into Chromatin Recognition by Multiple Domains of the Tumor Suppressor RBBP1

Retinoblastoma-binding protein 1 (RBBP1) is involved in gene regulation, epigenetic regulation, and disease processes. RBBP1 contains five domains with DNA-binding or histone-binding activities, but how RBBP1 specifically recognizes chromatin is still unknown. An AT-rich interaction domain (ARID) in RBBP1 was proposed to be the key region for DNA-binding and gene suppression. Here, we first determined the solution structure of a tandem PWWP-ARID domain mutant of RBBP1 after deletion of a long flexible acidic loop L12 in the ARID domain. NMR titration results indicated that the ARID domain interacts with DNA with no GC- or AT-rich preference. Surprisingly, we found that the loop L12 binds to the DNA-binding region of the ARID domain as a DNA mimic and inhibits DNA binding. The loop L12 can also bind weakly to the Tudor and chromobarrel domains of RBBP1, but binds more strongly to the DNA-binding region of the histone H2A-H2B heterodimer. Furthermore, both the loop L12 and DNA can enhance the binding of the chromobarrel domain to H3K4me3 and H4K20me3. Based on these results, we propose a model of chromatin recognition by RBBP1, which highlights the unexpected multiple key roles of the disordered acidic loop L12 in the specific binding of RBBP1 to chromatin.     

Free-living amoebae promote Candida auris survival and proliferation in water

Candida auris is an emerging species responsible for life-threatening infections. Its ability to be resistant to most systemic antifungal classes and its capacity to persist in a hospital environment have led to health concerns. Currently, data about environmental reservoirs are limited but remain essential in control of C. auris spread. The aim of our study was to explore the interactions between C. auris and two free-living amoeba (FLA) species, Vermamoeba vermiformis and Acanthamoeba castellanii, potentially found in the same water environment. Candida auris was incubated with FLA trophozoites or their culture supernatants. The number of FLA and yeasts was determined at different times and transmission electron microscopy (TEM) was performed. Supernatants of FLAs promoted yeast survival and proliferation. Internalization of viable C. auris within both FLA species was also evidenced by TEM. A water environmental reservoir of C. auris can therefore be considered through FLAs and contamination of the hospital water networks would consequently be possible.     

Mysterious bio-duck sound attributed to the Antarctic minke whale (Balaenoptera bonaerensis)

For decades, the bio-duck sound has been recorded in the Southern Ocean, but the animal producing it has remained a mystery. Heard mainly during austral winter in the Southern Ocean, this ubiquitous sound has been recorded in Antarctic waters and contemporaneously off the Australian west coast. Here, we present conclusive evidence that the bio-duck sound is produced by Antarctic minke whales (Balaenoptera bonaerensis). We analysed data from multi-sensor acoustic recording tags that included intense bio-duck sounds as well as singular downsweeps that have previously been attributed to this species. This finding allows the interpretation of a wealth of long-term acoustic recordings for this previously acoustically concealed species, which will improve our understanding of the distribution, abundance and behaviour of Antarctic minke whales. This is critical information for a species that inhabits a difficult to access sea-ice environment that is changing rapidly in some regions and has been the subject of contentious lethal sampling efforts and ongoing international legal action.     

Cystine-knot peptides: emerging tools for cancer imaging and therapy

Cystine-knot miniproteins, also known as knottins, constitute a large family of structurally related peptides with diverse amino acid sequences and biological functions. Knottins have emerged as attractive candidates for drug development as they potentially fill a niche between small molecules and protein biologics, offering drug-like properties and the ability to bind to clinical targets with high affinity and selectivity. Due to their extremely high stability and unique structural features, knottins also demonstrate promise in addressing challenging drug development goals, including the potential for oral delivery and the ability to access intracellular drug targets. Several naturally-occurring knottins have recently received approval for treating chronic pain and irritable bowel syndrome, while others are under development for tumor imaging applications. To expand beyond nature’s repertoire, rational and combinatorial protein engineering methods are generating tumor-targeting knottins for use as cancer diagnostics and therapeutics.     

Development of a human three-dimensional organotypic skin-melanoma spheroid model for in vitro drug testing

Despite remarkable efforts, metastatic melanoma (MM) still presents with significant mortality. Recently, mono-chemotherapies are increasingly replenished by more cancer-specific combination therapies involving death ligands and drugs interfering with cell signaling. Still, MM remains a fatal disease because tumors rapidly develop resistance to novel therapies thereby regaining tumorigenic capacity. Although genetically engineered mouse models for MM have been developed, at present no model is available that reliably mimics the human disease and is suitable for studying mechanisms of therapeutic obstacles including cell death resistance. To improve the increasing requests on new therapeutic alternatives, reliable human screening models are demanded that translate the findings from basic cellular research into clinical applications. By developing an organotypic full skin equivalent, harboring melanoma tumor spheroids of defined sizes we have invented a cell-based model that recapitulates both the 3D organization and multicellular complexity of an organ/tumor in vivo but at the same time accommodates systematic experimental intervention. By extending our previous findings on melanoma cell sensitization toward TRAIL (tumor necrosis factor-related apoptosis-inducing ligand) by co-application of sublethal doses of ultraviolet-B radiation (UVB) or cisplatin, we show significant differences in the therapeutical outcome to exist between regular two-dimensional (2D) and complex in vivo-like 3D models. Of note, while both treatment combinations killed the same cancer cell lines in 2D culture, skin equivalent-embedded melanoma spheroids are potently killed by TRAIL+cisplatin treatment but remain almost unaffected by the TRAIL+UVB combination. Consequently, we have established an organotypic human skin-melanoma model that will facilitate efforts to improve therapeutic outcomes for malignant melanoma by providing a platform for the investigation of cytotoxic treatments and tailored therapies in a more physiological setting.     

Skp-cullin-F box E3 ligase component FBXL2 ubiquitinates Aurora B to inhibit tumorigenesis

Aurora B kinase is an integral regulator of cytokinesis, as it stabilizes the intercellular canal within the midbody to ensure proper chromosomal segregation during cell division. Here we identified that the ubiquitin E3 ligase complex SCF^FBXL2 mediates Aurora B ubiquitination and degradation within the midbody, which is sufficient to induce mitotic arrest and apoptosis. Three molecular acceptor sites (K¹⁰², K¹⁰³ and K²⁰⁷) within Aurora B protein were identified as important sites for its ubiquitination. A triple Lys mutant of Aurora B (K¹⁰²/¹⁰³/^207R) exhibited optimal resistance to SCF^FBXL2-directed polyubiquitination, and overexpression of this variant resulted in a significant delay in anaphase onset, resulting in apoptosis. A unique small molecule F-box/LRR-repeat protein 2 (FBXL2) activator, BC-1258, stabilized and increased levels of FBXL2 protein that promoted Aurora B degradation, resulting in tetraploidy, mitotic arrest and apoptosis of tumorigenic cells, and profoundly inhibiting tumor formation in athymic nude mice. These findings uncover a new proteolytic mechanism targeting a key regulator of cell replication that may serve as a basis for chemotherapeutic intervention in neoplasia.