Selective Labeling and Identification of the Tumor Cell Proteome of Pancreatic Cancer In Vivo

Pancreatic ductal adenocarcinoma (PDAC) is among the deadliest cancers. Dissecting the tumor cell proteome from that of the non-tumor cells in the PDAC tumor bulk is critical for tumorigenesis studies, biomarker discovery, and development of therapeutics. However, investigating the tumor cell proteome has proven evasive due to the tumor’s extremely complex cellular composition. To circumvent this technical barrier, we have combined bioorthogonal noncanonical amino acid tagging (BONCAT) and data-independent acquisition mass spectrometry (DIA-MS) in an orthotopic PDAC model to specifically identify the tumor cell proteome in vivo. Utilizing the tumor cell-specific expression of a mutant tRNA synthetase transgene, this approach provides tumor cells with the exclusive ability to incorporate an azide-bearing methionine analogue into newly synthesized proteins. The azide-tagged tumor cell proteome is subsequently enriched and purified via a bioorthogonal reaction and then identified and quantified using DIA-MS. Applying this workflow to the orthotopic PDAC model, we have identified thousands of proteins expressed by the tumor cells. Furthermore, by comparing the tumor cell and tumor bulk proteomes, we showed that the approach can distinctly differentiate proteins produced by tumor cells from those of non-tumor cells within the tumor microenvironment. Our study, for the first time, reveals the tumor cell proteome of PDAC under physiological conditions, providing broad applications for tumorigenesis, therapeutics, and biomarker studies in various human cancers

Functionality of redox-active crysteines is required for restriction of retroviral replication by SAMHD1

Open access article. Creative Commons 4.0 International License (CC BY 4.0) appliesSAMHD1 is a dNTP triphosphohydrolase (dNTPase)that impairs retroviral replication in a subset of non-cycling immune cells. Here we show that SAMHD1is a redox-sensitive enzyme and identify threeredox-active cysteines within the protein: C341,C350, and C522. The three cysteines reside nearone another and the allosteric nucleotide bindingsite. Mutations C341S and C522S abolish the abilityof SAMHD1 to restrict HIV replication, whereas theC350S mutant remains restriction competent. TheC522S mutation makes the protein resistant to inhibi-tion by hydrogen peroxide but has no effect onthe tetramerization-dependent dNTPase activity ofSAMHD1in vitroor on the ability of SAMHD1 todeplete cellular dNTPs. Our results reveal that enzy-matic activation of SAMHD1 via nucleotide-depen-dent tetramerization is not sufficient for the estab-lishment of the antiviral state and that retroviralrestriction depends on the ability of the protein to un-dergo redox transformations.Ye

Evolving trends in the management of acute appendicitis during COVID-19 waves. The ACIE appy II study (vol 46, pg 2021, 2022)

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Global attitudes in the management of acute appendicitis during COVID-19 pandemic: ACIE Appy Study

Background: Surgical strategies are being adapted to face the COVID-19 pandemic. Recommendations on the management of acute appendicitis have been based on expert opinion, but very little evidence is available. This study addressed that dearth with a snapshot of worldwide approaches to appendicitis. Methods: The Association of Italian Surgeons in Europe designed an online survey to assess the current attitude of surgeons globally regarding the management of patients with acute appendicitis during the pandemic. Questions were divided into baseline information, hospital organization and screening, personal protective equipment, management and surgical approach, and patient presentation before versus during the pandemic. Results: Of 744 answers, 709 (from 66 countries) were complete and were included in the analysis. Most hospitals were treating both patients with and those without COVID. There was variation in screening indications and modality used, with chest X-ray plus molecular testing (PCR) being the commonest (19\ub78 per cent). Conservative management of complicated and uncomplicated appendicitis was used by 6\ub76 and 2\ub74 per cent respectively before, but 23\ub77 and 5\ub73 per cent, during the pandemic (both P < 0\ub7001). One-third changed their approach from laparoscopic to open surgery owing to the popular (but evidence-lacking) advice from expert groups during the initial phase of the pandemic. No agreement on how to filter surgical smoke plume during laparoscopy was identified. There was an overall reduction in the number of patients admitted with appendicitis and one-third felt that patients who did present had more severe appendicitis than they usually observe. Conclusion: Conservative management of mild appendicitis has been possible during the pandemic. The fact that some surgeons switched to open appendicectomy may reflect the poor guidelines that emanated in the early phase of SARS-CoV-2