Renal cell carcinoma (RCC) occurs in a number of cancer predisposition syndromes, but the genetic architecture of susceptibility to RCC is not well defined. We investigated the frequency of pathogenic and likely pathogenic (P/LP) germline variants in cancer susceptibility genes (CSGs) within a large series of unselected RCC participants. Whole-genome sequencing data on 1336 RCC participants and 5834 controls recruited to the UK 100 000 Genomes Project, a nationwide multicentre study, was analyzed to identify rare P/LP short variants (single nucleotide variants and insertions/deletions ranging from 1 to 50 base pairs) and structural variants in 121 CSGs. Among 1336 RCC participants [mean: 61.3 years (±12 SD), range: 13–88 years; 64% male], 85 participants [6.4%; 95% CI (5.1, 7.8)] had one or more P/LP germline variant in a wider range of CSGs than previously recognized. A further 64 intragenic variants in CSGs previously associated with RCC were classified as a variant of uncertain significance (VUS) (24 ‘hot VUSs’) and were considered to be of potential clinical relevance as further evaluation might results in their reclassification. Most patients with P variants in well-established CSGs known to predispose to renal cell carcinoma (RCC-CSGs) were aged <50 years. Burden test analysis for filtered variants in CSGs demonstrated a significant excess of CHEK2 variants in European RCC participants compared with the healthy European controls (P = 0.0019). Approximately, 6% of the patients with RCC unselected for family history have a germline variant requiring additional follow-up analysis. To improve diagnostic yield, we suggest expanding the panel of RCC-CSGs tested to include CHEK2 and all SDHx subunits and raising the eligibility criteria for age-based testing.

The transcription factor Gli-similar 3 (Glis3) plays a critical role in the generation of pancreatic ß cells and the regulation insulin gene transcription and has been implicated in the development of several pathologies, including type 1 and 2 diabetes and polycystic kidney disease. However, little is known about the proteins and posttranslational modifications that regulate or mediate Glis3 transcriptional activity. In this study, we identify by mass-spectrometry and yeast 2-hybrid analyses several proteins that interact with the N-terminal region of Glis3. These include the WW-domain-containing HECT E3 ubiquitin ligases, Itch, Smurf2, and Nedd4. The interaction between Glis3 and the HECT E3 ubiquitin ligases was verified by co-immunoprecipitation assays and mutation analysis. All three proteins interact through their WW-domains with a PPxY motif located in the Glis3 N-terminus. However, only Itch significantly contributed to Glis3 polyubiquitination and reduced Glis3 stability by enhancing its proteasomal degradation. Itch-mediated degradation of Glis3 required the PPxY motif-dependent interaction between Glis3 and the WW-domains of Itch as well as the presence of the Glis3 zinc finger domains. Transcription analyses demonstrated that Itch dramatically inhibited Glis3-mediated transactivation and endogenous Ins2 expression by increasing Glis3 protein turnover. Taken together, our study identifies Itch as a critical negative regulator of Glis3-mediated transcriptional activity. This regulation provides a novel mechanism to modulate Glis3-driven gene expression and suggests that it may play a role in a number of physiological processes controlled by Glis3, such as insulin transcription, as well as in Glis3-associated diseases.

The earliest step in DNA replication is origin licensing, which is the DNA loading of minichromosome maintenance (MCM) helicase complexes. The Cdc10-dependent transcript 1 (Cdt1) protein is essential for MCM loading during the G1 phase of the cell cycle, but the mechanism of Cdt1 function is still incompletely understood. We examined a collection of rare Cdt1 variants that cause a form of primordial dwarfism (the Meier-Gorlin syndrome) plus one hypomorphic Drosophila allele to shed light on Cdt1 function. Three hypomorphic variants load MCM less efficiently than wild-type (WT) Cdt1, and their lower activity correlates with impaired MCM binding. A structural homology model of the human Cdt1-MCM complex positions the altered Cdt1 residues at two distinct interfaces rather than the previously described single MCM interaction domain. Surprisingly, one dwarfism allele (Cdt1-A66T) is more active than WT Cdt1. This hypermorphic variant binds both cyclin A and SCFSkp2 poorly relative to WT Cdt1. Detailed quantitative live-cell imaging analysis demonstrated no change in the stability of this variant, however. Instead, we propose that cyclin A/CDK inhibits the Cdt1 licensing function independent of the creation of the SCFSkp2 phosphodegron. Together, these findings identify key Cdt1 interactions required for both efficient origin licensing and tight Cdt1 regulation to ensure normal cell proliferation and genome stability.

Uveal melanoma (UM) is an uncommon but highly aggressive ocular malignancy. Poor overall survival is associated with deleterious BAP1 alterations, which frequently occur with monosomy 3 (LOH3) and a characteristic gene expression profile. Tumor DNA from a cohort of 100 UM patients from Moorfields Biobank (UK) that had undergone enucleation were sequenced for known UM driver genes (BAP1, SF3B1, EIF1AX, GNAQ, and GNA11). Immunohistochemical staining of BAP1 and interphase FISH for chromosomes 3 and 8 was performed, and cellular localization of BAP1 was correlated with BAP1 mutations. Wildtype (WT) BAP1 staining was characterized by nBAP1 expression with 10% cBAP1.

Background: Head and neck paragangliomas (HNPGLs) are typically slow-growing, hormonally inactive tumors of parasympathetic paraganglia. Inactivation of prolyl-hydroxylase domain-containing 2 protein causing indirect gain-of-function of hypoxia-inducible factor-2α (HIF-2α), encoded by EPAS1, was recently shown to cause carotid body hyperplasia. We previously described a syndrome with multiple sympathetic paragangliomas caused by direct gain-of-function variants in EPAS1 (Pacak-Zhuang syndrome, PZS) and developed a corresponding mouse model. Methods: We evaluated a cohort of patients with PZS (n = 9) for HNPGL by positron emission tomography, magnetic resonance imaging, and computed tomography and measured carotid body size compared to literature reference values. Resected tumors were evaluated by histologic sectioning and staining. We evaluated the corresponding mouse model at multiple developmental stages (P8 and adult) for lesions of the head and neck by high resolution ex vivo imaging and performed immunohistochemical staining on histologic sections of the identified lesions. Results: Three patients had imaging consistent with HNPGL, one of which warranted resection and was confirmed on histology. Three additional patients had carotid body enlargement (Z-score > 2.0), and 3 had carotid artery malformations. We found that 9 of 10 adult variant mice had carotid body tumors and 6 of 8 had a paraganglioma on the cranio-caval vein, the murine homologue of the superior vena cava; these were also found in 4 of 5 variant mice at post-natal day 8. These tumors and the one resected from a patient were positive for tyrosine hydroxylase, synaptophysin, and chromogranin A. Brown fat in a resected patient tumor carried the EPAS1 pathogenic variant. Conclusions: These findings (1) suggest HNPGL as a feature of PZS and (2) show that these pathogenic variants are sufficient to cause the development of these tumors, which we believe represents a continuous spectrum of disease starting from hyperplasia.

Background: The coronavirus disease-2019 stressed health science education across many science disciplines. At one public medical school, faculty educators responded with the design and delivery of an online COVID-19 pandemic course for clinical phase students. However, faculty resources were limited, and no official course was offered to preclinical students, despite reported interest. Subsequently, clinical students created an 8-week online, student-run curriculum. We aimed to assess how effectively the course improved medical student knowledge on COVID-19, created community, and improved other professional skills. Methods: Participants and instructors completed four surveys—two pre-course surveys and two post-course surveys. Average ratings for each question were calculated and analyzed using independent sample t tests. Results: Of the 188 students in the preclinical cohort, 143 enrolled in the course with 63.64% attending all, 91% attending ≥7 out of 8, and 96% attending ≥6 out of 8 sessions. Students and instructors reported an overall mean increase in knowledge on all course topics, peer teaching, learning in an online format, and overall preparedness with regards to a public health crisis. Overall, 79.2% of student participants and instructors reported being satisfied with the experience. Conclusion: Peer teaching and learning offers a valuable educational experience for participants and instructors alike. Student-driven curricula may provide an effective teaching model for medical education, contributing to efforts that improve student confidence and leadership skills, provide role models for junior students, and prepare students for their future role as educations.