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Abstract Protein–protein interactions (PPIs) offer great opportunities to expand the druggable proteome and therapeutically tackle various diseases, but remain challenging targets for drug discovery. Here, we provide a comprehensive pipeline that combines experimental and computational tools to identify and validate PPI targets and perform early-stage drug discovery. We have developed a machine learning approach that prioritizes interactions by analyzing quantitative data from binary PPI assays or AlphaFold-Multimer predictions. Using the quantitative assay LuTHy together with our machine learning algorithm, we identified high-confidence interactions among SARS-CoV-2 proteins for which we predicted three-dimensional structures using AlphaFold-Multimer. We employed VirtualFlow to target the contact interface of the NSP10-NSP16 SARS-CoV-2 methyltransferase complex by ultra-large virtual drug screening. Thereby, we identified a compound that binds to NSP10 and inhibits its interaction with NSP16, while also disrupting the methyltransferase activity of the complex, and SARS-CoV-2 replication. Overall, this pipeline will help to prioritize PPI targets to accelerate the discovery of early-stage drug candidates targeting protein complexes and pathways.

Viral infections are known to hijack the transcription and translation of the host cell. However, the extent to which viral proteins coordinate these perturbations remains unclear. Here we used a model system, the human T-cell leukemia virus type 1 (HTLV-1), and systematically analyzed the transcriptome and interactome of key effectors oncoviral proteins Tax and HBZ. We showed that Tax and HBZ target distinct but also common transcription factors. Unexpectedly, we also uncovered a large set of interactions with RNA-binding proteins, including the U2 auxiliary factor large subunit (U2AF2), a key cellular regulator of pre-mRNA splicing. We discovered that Tax and HBZ perturb the splicing landscape by altering cassette exons in opposing manners, with Tax inducing exon inclusion while HBZ induces exon exclusion. Among Tax- and HBZ-dependent splicing changes, we identify events that are also altered in Adult T cell leukemia/lymphoma (ATLL) samples from two independent patient cohorts, and in well-known cancer census genes. Our interactome mapping approach, applicable to other viral oncogenes, has identified spliceosome perturbation as a novel mechanism coordinated by Tax and HBZ to reprogram the transcriptome.

Comprehensive mapping of binary protein-protein interactions requires to combine several complementary assays. Here, the authors show that complete coverage could be reached with a minimal number of assays as long as they explore various experimental conditions.

Objectives: Macroscopic studies have suggested a link between distal border synovial invaginations of the navicular bone and the distal interphalangeal joint. However, many practitioners consider that these invaginations are directly and solely related to navicular disease. The objective was to investigate the communication pattern of these synovial invaginations with the synovial compartments of the distal interphalangeal joint and the navicular bursa, using minimally invasive imaging techniques., Methods: In a prospective observational study, 10 cadaveric limbs with radiographically evident distal border synovial invaginations were randomly assigned to computed tomography arthrography or bursography groups, using iopamidol., Results: In 5/5 limbs, contrast medium filled the invaginations following distal interphalangeal arthrography. In the other five limbs, no contrast medium filled the invaginations following bursography., Clinical Significance: Contrary to existing beliefs, these invaginations are more likely associated with distal interphalangeal joint synovitis and may not be directly linked to primary navicular bone pathology, but might reflect distal interphalangeal arthropathy. Therefore, the rationale for assessment of these invaginations in stallion selection or pre-purchase examinations as a predictive sign for navicular disease is questionable. Nonetheless, comorbidities are frequent in the equine distal limb. Enlarged synovial invaginations may also be seen in limbs with concomitant primary navicular disease. Further studies are needed to elucidate possible inter-related pathological processes.

Meniscal tears and subchondral bone marrow lesions have both been described in dogs with cranial cruciate ligament rupture, but their possible concurrence has not been evaluated. In a population of 14 dogs exhibiting signs of stifle pain with surgically confirmed cranial cruciate ligament rupture, a short presurgical 1.5T magnetic resonance (MR) imaging protocol including dorsal proton density, dorsal T1-weighted gradient recalled echo, and sagittal fat-saturated dual echo sequences was tested to further investigate these features and illustrate meniscal tears. Interobserver agreement for detection of medial meniscal tears (k=0.83) and bone marrow lesions (k=0.87) was excellent. Consensus MR reading allowed detection of nine out of 12 surgically confirmed medial meniscal tears and there was no false positive. All dogs had cruciate ligament enthesis-related bone marrow lesions in the tibia, femur or both bones. Additionally, among the 12 dogs with confirmed medial meniscal tears, subchondral bone marrow lesions were present in the caudomedial (9 dogs) and caudoaxial (11 dogs) regions of the tibial plateau, resulting in odds ratios (13.6, p=0.12, and 38.3, p=0.04, respectively) that had large confidence intervals due to the small group size of this study. The other two dogs had neither tibial bone marrow lesions in these locations nor medial meniscal tears. These encouraging preliminary results warrant further investigation using this clinically realistic preoperative MR protocol. As direct diagnosis of meniscal tears remained challenging in dogs even with high-field MR, identification of associated signs such as subchondral bone marrow lesions might indirectly allow suspicion of an otherwise unrecognized meniscal tear.