Chemoproteomic discovery of a human RNA ligase.

RNA ligases are present across all forms of life. While enzymatic RNA ligation between 5'-PO4 and 3'-OH termini is prevalent in viruses, fungi, and plants, such RNA ligases are yet to be identified in vertebrates. Here, using a nucleotide-based chemical probe targeting human AMPylated proteome, we have enriched and identified the hitherto uncharacterised human protein chromosome 12 open reading frame 29 (C12orf29) as a human enzyme promoting RNA ligation between 5'-PO4 and 3'-OH termini. C12orf29 catalyses ATP-dependent RNA ligation via a three-step mechanism, involving tandem auto- and RNA AMPylation. Knock-out of C12ORF29 gene impedes the cellular resilience to oxidative stress featuring concurrent RNA degradation, which suggests a role of C12orf29 in maintaining RNA integrity. These data provide the groundwork for establishing a human RNA repair pathway.

Chemical proteomics reveals interactors of the alarmone diadenosine triphosphate in the cancer cell line H1299.

Intracellular dinucleoside polyphosphates (Npn Ns) have been known for decades but the functional role remains enigmatic. Diadenosine triphosphate (Ap3 A) is one of the most prominent examples, and its intercellular concentration was shown to increase upon cellular stress. By employment of previously reported Ap3 A-based photoaffinity-labeling probes (PALPs) in chemical proteomics, we investigated the Ap3 A interactome in the human lung carcinoma cell line H1299. The cell line is deficient of the fragile histidine triade (Fhit) protein, a hydrolase of Ap3 A and tumor suppressor. Overall, the number of identified potential interaction partners was significantly lower than in the previously investigated HEK293T cell line. Gene ontology term analysis revealed that the identified proteins participate in similar pathways as for HEK293T, but the percentage of proteins involved in RNA-related processes is higher for H1299. The obtained results highlight similarities and differences of the Ap3 A interaction network in different cell lines and give further indications regarding the importance of the presence of Fhit.

Chemical proteomic profiling reveals protein interactors of the alarmones diadenosine triphosphate and tetraphosphate.

The nucleotides diadenosine triphosphate (Ap3A) and diadenosine tetraphosphate (Ap4A) are formed in prokaryotic and eukaryotic cells. Since their concentrations increase significantly upon cellular stress, they are considered to be alarmones triggering stress adaptive processes. However, their cellular roles remain elusive. To elucidate the proteome-wide interactome of Ap3A and Ap4A and thereby gain insights into their cellular roles, we herein report the development of photoaffinity-labeling probes and their employment in chemical proteomics. We demonstrate that the identified ApnA interactors are involved in many fundamental cellular processes including carboxylic acid and nucleotide metabolism, gene expression, various regulatory processes and cellular response mechanisms and only around half of them are known nucleotide interactors. Our results highlight common functions of these ApnAs across the domains of life, but also identify those that are different for Ap3A or Ap4A. This study provides a rich source for further functional studies of these nucleotides and depicts useful tools for characterization of their regulatory mechanisms in cells.

Comparison of massive and emergency transfusion prediction scoring systems after trauma with a new Bleeding Risk Index score applied in-flight.

BACKGROUND: Assessment of blood consumption (ABC), shock index (SI), and Revised Trauma Score (RTS) are used to estimate the need for blood transfusion and triage. We compared Bleeding Risk Index (BRI) score calculated with trauma patient noninvasive vital signs and hypothesized that prehospital BRI has better performance compared with ABC, RTS, and SI for predicting the need for emergent and massive transfusion (MT). METHODS: We analyzed 2-year in-flight data from adult trauma patients transported directly to a Level I trauma center via helicopter. The BRI scores 0 to 1 were derived from continuous features of photoplethymographic and electrocardiographic waveforms, oximetry values, blood pressure trends. The ABC, RTS, and SI were calculated using admission data. The area under the receiver operating characteristic curve (AUROC) with 95% confidence interval (CI) was calculated for predictions of critical administration threshold (CAT, ≥3 units of blood in the first hour) or MT (≥10 units of blood in the first 24 hours). DeLong's method was used to compare AUROCs for different scoring systems. p < 0.05 was considered statistically significant. RESULTS: Among 1,396 patients, age was 46.5 ± 20.1 years (SD), 67.1% were male. The MT rate was 3.2% and CAT was 7.6%, most (92.8%) were blunt injury. Mortality was 6.6%. Scene arrival to hospital time was 35.3 ± (10.5) minutes. The BRI prediction of MT with AUROC 0.92 (95% CI, 0.89-0.95) was significantly better than ABC, SI, or RTS (AUROCs = 0.80, 0.83, 0.78, respectively; 95% CIs 0.73-0.87, 0.76-0.90, 0.71-0.85, respectively). The BRI prediction of CAT had an AUROC of 0.91 (95% CI, 0.86-0.94), which was significantly better than ABC (AUROC, 077; 95% CI, 0.73-0.82) or RTS (AUROC, 0.79; 95% CI, 0.74-0.83) and better than SI (AUROC, 0.85; 95% CI, 0.80-0.89). The BRI score threshold for optimal prediction of CAT was 0.25 and for MT was 0.28. CONCLUSION: The autonomous continuous noninvasive patient vital signs-based BRI score performs better than ABC, RTS, and SI predictions of MT and CAT. Bleeding Risk Index does not require additional data entry or expert interpretation. LEVEL OF EVIDENCE: Prognostic test, level III.

Tablets made from paper.

The possibility to compress ordinary paper into tablets was systematically investigated in this study. Results proved that tablets can be made from paper, independent of the type of paper used. The tablets appear shiny and with a smooth surface. The pharmaceutical quality was acceptable, i.e. all tablets fulfilled the requirements for tablets according to the European Pharmacopeia. Drug-loaded tablets were produced by compression of drug-loaded paper. Drug loading did not alter the pharmaceutical quality. However, the uncoated tablets possessed an extremely fast disintegration, i.e. intense swelling upon contact with water, which might hamper the swallowing after oral administration. To avoid swelling tablets were successfully coated with a polymer film, leading to a prevention of swelling but immediate disintegration in simulated gastric fluid. In fact, tablets made from paper are a novel and promising strategy for improved oral drug delivery. They can be easily produced without any further excipients and possess pharmaceutical quality according to the European Pharmacopeia.