An SPNS1-dependent lysosomal lipid transport pathway that enables cell survival under choline limitation.

Lysosomes degrade macromolecules and recycle their nutrient content to support cell function and survival. However, the machineries involved in lysosomal recycling of many nutrients remain to be discovered, with a notable example being choline, an essential metabolite liberated via lipid degradation. Here, we engineered metabolic dependency on lysosome-derived choline in pancreatic cancer cells to perform an endolysosome-focused CRISPR-Cas9 screen for genes mediating lysosomal choline recycling. We identified the orphan lysosomal transmembrane protein SPNS1 as critical for cell survival under choline limitation. SPNS1 loss leads to intralysosomal accumulation of lysophosphatidylcholine (LPC) and lysophosphatidylethanolamine (LPE). Mechanistically, we reveal that SPNS1 is a proton gradient-dependent transporter of LPC species from the lysosome for their re-esterification into phosphatidylcholine in the cytosol. Last, we establish that LPC efflux by SPNS1 is required for cell survival under choline limitation. Collectively, our work defines a lysosomal phospholipid salvage pathway that is essential under nutrient limitation and, more broadly, provides a robust platform to deorphan lysosomal gene function.

Medical student's perception of the COVID-19 pandemic effect on their education and well-being: a cross-sectional survey in the United States.

BACKGROUND: The effects of drastic curricular changes necessitated by the COVID-19 pandemic on medical students' education and wellbeing have remained largely unstudied. Out study aimed to characterize how medical students were affected by the pandemic, specifically how limitations introduced by the pandemic may have affected the quality, delivery, and experience of medical education. METHODS: Three hundred students from 5 U.S. allopathic medical schools were surveyed to determine students' perceptions about their quality of medical education, professional development, and mental health during the COVID-19 pandemic (October 2020-December 2020). RESULTS: A large majority of students report that while lecture-based learning has not been significantly affected by the pandemic, small-group and clinical learning have greatly declined in quality. Students also reported higher levels of depression, anxiety, and uncertainty with regards to their futures as physicians. CONCLUSIONS: The COVID-19 pandemic has greatly affected the medical student education and wellbeing. Although medical schools have implemented measures to continue to train medical students as effectively as they can, further strategies must be devised to ensure the well-being of students in the present and for future national emergencies.

The renal clear cell carcinoma immune landscape.

A comprehensive evaluation of the clear cell renal cell carcinoma (ccRCC) immune landscape was found using 584 RNA-sequencing datasets from The Cancer Genome Atlas (TCGA), we identified 17 key dysregulated immune-associated genes in ccRCC based on association with clinical variables and important immune pathways. Of the numerous findings from our analyses, we found that several of the 17 key dysregulated genes are heavily involved in interleukin and NF-kB signaling and that somatic copy number alteration (SCNA) hotspots may be causally associated with gene dysregulation. More importantly, we also found that key immune-associated genes and pathways are strongly upregulated in ccRCC. Our study may lend novel insights into the clinical implications of immune dysregulation in ccRCC and suggests potential immunotherapeutic targets for further evaluation.

Computational methods reveal novel functionalities of PIWI-interacting RNAs in human papillomavirus-induced head and neck squamous cell carcinoma.

Human papillomavirus (HPV) infection is the fastest growing cause of head and neck squamous cell carcinoma (HNSCC) today, but its role in malignant transformation remains unclear. This study aimed to conduct a comprehensive investigation of PIWI-interacting RNA (piRNA) alterations and functionalities in HPV-induced HNSCC. Using 77 RNA-sequencing datasets from TCGA, we examined differential expression of piRNAs between HPV16(+) HNSCC and HPV(-) Normal samples, identifying a panel of 30 HPV-dysregulated piRNAs. We then computationally investigated the potential mechanistic significances of these transcripts in HPV-induced HNSCC, identifying our panel of piRNAs to associate with the protein PIWIL4 as well as the RTL family of retrotransposon-like genes, possibly through direct binding interactions. We also recognized several HPV-dysregulated transcripts for their correlations with well-documented mutations and copy number variations in HNSCC as well as HNSCC clinical variables, demonstrating the potential ability of our piRNAs to play important roles in large-scale modulation of HNSCC in addition to their direct, smaller-scale interactions in this malignancy. The differential expression of key piRNAs, including NONHSAT077364, NONHSAT102574, and NONHSAT128479, was verified in vitro by evaluating endogenous expression in HPV(+) cancer vs. HPV(-) normal cell lines. Overall, our novel study provides a rigorous investigation of piRNA dysregulation in HPV-related HNSCC, and lends critical insight into the idea that these small regulatory transcripts may play crucial and previously unidentified roles in tumor pathogenesis and progression.

A comprehensive study of smoking-specific microRNA alterations in head and neck squamous cell carcinoma.

OBJECTIVE: While tobacco smoking is a well-known risk factor for head and neck squamous cell carcinoma (HNSCC), the molecular mechanisms underlying tobacco-induced HNSCC remain unclear. This study sought to comprehensively identify microRNA (miRNA) alterations and evaluate their clinical relevance in smoking-induced HNSCC pathogenesis and progression. MATERIALS AND METHODS: Using small RNA-sequencing data and clinical data from 145 HNSCC patients, we performed a series of differential expression and correlation analyses to identify a panel of tobacco-dysregulated miRNAs associated with key clinical characteristics in HNSCC. We then examined the expression patterns of these miRNAs in normal epithelial cell lines following exposure to cigarette smoke extract. RESULTS: Our analyses revealed distinct panels of miRNAs to be dysregulated with smoking status and associated with additional clinical features, including tumor stage, metastasis, anatomic site, and patient survival. The differential expression of key miRNAs, including miR-101, miR-181b, miR-486, and miR-1301, was verified in cigarette-treated epithelial cell lines, suggesting their potential roles in the early development of smoking-related HNSCCs. CONCLUSION: Specific alterations in miRNA expression may be traced to tobacco use and are associated with important HNSCC clinical characteristics. Future studies of these miRNAs may be valuable for furthering the understanding and targeted treatment of smoking-associated HNSCC.

Smoking status regulates a novel panel of PIWI-interacting RNAs in head and neck squamous cell carcinoma.

OBJECTIVE: Smoking remains a primary etiological factor in head and neck squamous cell carcinoma (HNSCC). Given that non-coding RNAs (ncRNAs), including PIWI-interacting RNAs (piRNAs), have emerged as mediators of initiation and progression in head and neck malignancies, we undertook a global study of piRNA expression patterns in smoking-associated HNSCC. MATERIALS AND METHODS: Using RNA-sequencing data from 256 current smoker and lifelong nonsmoker samples in The Cancer Genome Atlas (TCGA), we analyzed the differential expression patterns of 27,127 piRNAs across patient cohorts stratified by tobacco use, with HPV16 status and tumor status taken into account. We correlated their expression to clinical characteristics and to smoking-induced alterations of PIWI proteins, the functional counterparts of piRNAs. Finally, we correlated our identified piRNAs and PIWI proteins to known chromosomal aberrations in HNSCC to understand their wider-ranging genomic effects. RESULTS AND CONCLUSION: Our analyses implicated a 13-member piRNA panel in smoking-related HNSCC, among which NONHSAT123636 and NONHSAT113708 associated with tumor stage, NONHSAT067200 with patient survival, and NONHSAT081250 with smoking-altered PIWIL1 protein expression. 6 piRNAs as well as PIWIL1 correlated with genomic alterations common to HNSCC, including TP53 mutation, TP53-3p co-occurrence, and 3q26, 8q24, and 11q13 amplification. Collectively, our findings provide novel insights into the etiology-specific piRNA landscape of smoking-induced HNSCC.

Electronic cigarettes induce DNA strand breaks and cell death independently of nicotine in cell lines.

OBJECTIVES: Evaluate the cytotoxicity and genotoxicity of short- and long-term e-cigarette vapor exposure on a panel of normal epithelial and head and neck squamous cell carcinoma (HNSCC) cell lines. MATERIALS AND METHODS: HaCaT, UMSCC10B, and HN30 were treated with nicotine-containing and nicotine-free vapor extract from two popular e-cigarette brands for periods ranging from 48 h to 8 weeks. Cytotoxicity was assessed using Annexin V flow cytometric analysis, trypan blue exclusion, and clonogenic assays. Genotoxicity in the form of DNA strand breaks was quantified using the neutral comet assay and γ-H2AX immunostaining. RESULTS: E-cigarette-exposed cells showed significantly reduced cell viability and clonogenic survival, along with increased rates of apoptosis and necrosis, regardless of e-cigarette vapor nicotine content. They also exhibited significantly increased comet tail length and accumulation of γ-H2AX foci, demonstrating increased DNA strand breaks. CONCLUSION: E-cigarette vapor, both with and without nicotine, is cytotoxic to epithelial cell lines and is a DNA strand break-inducing agent. Further assessment of the potential carcinogenic effects of e-cigarette vapor is urgently needed.