KRAS is mutated in over 90% of pancreatic ductal adenocarcinomas (PDAC), where hotspot alterations in codons 12, 13, and 61 drive tumor initiation and progression. Although distinct biochemical properties have been described for individual KRAS mutants, whether they generate unique allele-specific signaling programs in PDAC cells remains unresolved. Here, we systematically interrogated the molecular consequences of seven common KRAS mutant variants in reconstituted isogenic, KRAS-deficient PDAC cell lines by integrated transcriptomic, proteomic, and phosphoproteomic profiling. We found that baseline cellular state, rather than allele identity, was the predominant driver of molecular variation. Comparisons with established KRAS reference signatures revealed significant but moderate overlap at the mRNA level and less so at the proteome level. Pathway analyses highlighted interferon response and mitochondrial translation as recurrently altered across alleles, while phosphoproteomic data confirmed robust ERK1/2 activity and suppression of DYRK kinase substrates by mutant KRAS expression. Importantly, no robust allele-specific molecular programs were identified. Together, our study establishes a comprehensive multi-omics resource for KRAS signaling in PDAC and demonstrates that cellular context exerts a stronger influence than allele identity in shaping molecular profiles, with implications for interpreting putative allele-specific signaling dependencies and therapeutic vulnerabilities.

How aneuploid cells tolerate chromosome arm gains or losses remains an open question. Using an isogenic human lung cell model with either chromosome 3p loss or 3q gain, combined with quantitative mass spectrometry and isotopic labeling, we reveal distinct proteostasis mechanisms for gain- and loss-type aneuploidy. Surprisingly, while compensation for 3q gain is primarily driven by increased degradation of excess protein complex subunits, 3p loss is neither counteracted by global protein degradation nor selectively reduced degradation. Rather, there is a relative upregulation in protein synthesis of those 3p-encoded proteins that participate in stable protein complexes to maintain functional complex stoichiometry. Additionally, 3p-encoded proteins that are in a complex show increased thermal stability in loss-type aneuploidy, potentially via their interactions with other proteins from euploid chromosomes. Together, our findings uncover distinct proteomic buffering strategies that enable cells to tolerate either excessive or deficient single-arm aneuploidy.

To support PTM proteomic analysis and annotation in different species, we developed PTMoreR, a user-friendly tool that considers the surrounding amino acid sequences of PTM sites during BLAST, enabling a motif-centric analysis across species. By controlling sequence window similarity, PTMoreR can map phosphoproteomic results between any two species, perform site-level functional enrichment analysis, and generate kinase-substrate networks. We demonstrate that the majority of real P-sites in mice can be inferred from experimentally derived human P-sites with PTMoreR mapping. Furthermore, the compositions of 129 mammalian phosphoproteomes can also be predicted using PTMoreR. The method also identifies cross-species phosphorylation events that occur on proteins with an increased tendency to respond to the environmental factors. Moreover, the classic kinase motifs can be extracted across mammalian species, offering an evolutionary angle for refining current motifs. PTMoreR supports PTM proteomics in non-human species and facilitates quantitative phosphoproteomic analysis.

The serine/threonine kinase AKT is a central node in cell signaling. While aberrant AKT activation underlies the development of a variety of human diseases, how different patterns of AKT-dependent phosphorylation dictate downstream signaling and phenotypic outcomes remains largely enigmatic. Herein, we perform a systems-level analysis that integrates methodological advances in optogenetics, mass spectrometry-based phosphoproteomics, and bioinformatics to elucidate how different intensity, duration, and pattern of Akt1 stimulation lead to distinct temporal phosphorylation profiles in vascular endothelial cells. Through the analysis of ~35,000 phosphorylation sites across multiple conditions precisely controlled by light stimulation, we identify a series of signaling circuits activated downstream of Akt1 and interrogate how Akt1 signaling integrates with growth factor signaling in endothelial cells. Furthermore, our results categorize kinase substrates that are preferably activated by oscillating, transient, and sustained Akt1 signals. We validate a list of phosphorylation sites that covaried with Akt1 phosphorylation across experimental conditions as potential Akt1 substrates. Our resulting dataset provides a rich resource for future studies on AKT signaling and dynamics.

Phosphorylation is one of the most important post-translational modifications (PTMs) of proteins, governing critical protein functions. Most human proteins have been shown to undergo phosphorylation, and phosphoproteomic studies have been widely applied due to recent advancements in high-resolution mass spectrometry technology. Although the experimental workflow for phosphoproteomics has been well-established, it would be useful to optimize and summarize a detailed, feasible protocol that combines phosphoproteomics and data-independent acquisition (DIA), along with follow-up data analysis procedures due to the recent instrumental and bioinformatic advances in measuring and understanding tens of thousands of site-specific phosphorylation events in a single experiment. Here, we describe an optimized Phos-DIA protocol, from sample preparation to bioinformatic analysis, along with practical considerations and experimental configurations for each step. The protocol is designed to be robust and applicable for both small-scale phosphoproteomic analysis and large-scale quantification of hundreds of samples for studies in systems biology and systems medicine.

Adding the plasmon optical material into DSSC photoanode has been demonstrated to be an promising method for enhancing the photoelectric conversion ability of photoanode sensitized with presently commercial dye. Zirconium nitride (ZrN) based on relatively cheap Earth-abundant elements is a highly potential alternative to noble plasmon gold. Herein plasmonic ZrN@TiO2 core-shell nanostructures are fabricated and incorporated into the photoanode of DSSC. Related results show that ZrN@TiO2 nanostructure indeed can efficiently improve the photovoltaic performance of corresponding DSSC. The DSSC doped with ZrN@TiO2 yields the champion power conversion efficiency (PCE) of 8.6%. Spectroscopic investigation and theoretical study demonstrate that plasmon effects originated from ZrN can effectively boost the light absorption of dye molecules and accelerate the photogenerated carrier separation, thus elevating the overall harvesting-conversion efficiency of photoanode for light and contributing to the improvement of DSSC photovoltaic performance. This work may offer a kind of new material support to further develop DSSC and provide experimental foundation for the application of ZrN in other solar energy utilization field.

Benefiting from the rapid development on chloride incorporated perovskite materials, nowadays, novel device fabricating techniques have sparked a new vision of ultraviolet photodetectors (UVPDs), which is moving toward higher precision, lower energy consumption, and greater miniaturization. Nevertheless, the natural poor solubility of chloride precursors for perovskites has limited their processability for achieving high‐performance self‐driven photovoltaic‐type UVPDs. Herein, a ball‐milling pretreated single‐source vapor deposition strategy is presented, which can facilely fabricate a pure‐phase CsPbCl 3 perovskite film with supermicrometer crystal size and no grain boundaries in the vertical direction. Based on this excellent quality UV harvesting perovskite layer, an optimized self‐driven visible‐blind UVPD exhibits outstanding detection properties, including a superior peak responsivity of 118 mA W −1 , a maximum specific detectivity of 6.62 × 10 12 Jones, and a wide linear dynamic range of 136 dB, coupled with >1000 h operational durability in air ambient. Most notably, a recorded ultrafast rise/fall response time of 120/820 ns is realized as the fastest responsive perovskite UVPDs to date. In addition, this CsPbCl 3 UVPD offers adequate visible‐transparency (>50% average visible transmittance) for a glazing integration by using a transparent back electrode, providing a promising pathway to realize a transparent sensor system for future smart window applications.

Herein, dispersed Ni species over the surface of plasmonic TiN nanocrystals (TiN@Ni) are manufactured by using wetness impregnation method. This developmental material holds abundant surface sites and local surface plasmon resonance property. To further satisfy the requirement as the electrocatalyst for dye-sensitized solar cell (DSSC), bifunctional TiN@Ni nanocrystals are incorporated with monolayer MXene to construct the continuous conductive matrix. The yielded TiN@Ni-MXene film serves as counter electrode, power conversion efficiency (PCE) of corresponding DSSC under conventional irradiation condition is 8.08%, which surpasses as-reference Pt-based DSSC(7.59%). When further adding the NIR irradiation from counter electrode side of device, DSSC achieves an impressive PCE of 8.45%. The superior performance of TiN@Ni-MXene electrode should be attrib-uted to the created active sites on the surface of TiN support, and the plasmonic effect from TiN@Ni nano-particles via utilizing NIR light. Ni species provide more adsorption sites for triiodide ions, meanwhile the elevated temperature from plasmon-induced photothermal effect can effectively boost the triiodide reducing reaction rates at the interface of electrode and electrolyte. Thus electrocatalytic performance of TiN@Ni-MXene counter electrode is remarkablely enhanced. The strategy here will be beneficial for the design of highly active and stable electrocatalyst for DSSC, as well as realizing the efficient utilization for wide-spectrum solar energy.

Human cancer cell lines are widely used in pharmacological and systems biological studies. The rapid documentation of the steady-state gene expression landscape of the cells used in a particular experiment may help to improve the reproducibility of scientific research. Here we applied a data-independent acquisition mass spectrometry (DIA-MS) method, coupled with a peptide spectral-library-free data analysis workflow, to measure both the proteome and phosphoproteome of a melanoma cell line panel with different metastatic properties. For each cell line, the single-shot DIA-MS detected 8100 proteins and almost 40 000 phosphopeptides in the respective measurements of two hours. Benchmarking the DIA-MS data towards the RNA-seq data and tandem mass tag (TMT)-MS results from the same set of cell lines demonstrated comparable qualitative coverage and quantitative reproducibility. Our data confirmed the high but complex mRNA-protein and protein-phospsite correlations. The results successfully established DIA-MS as a strong and competitive proteotyping approach for cell lines. The data further showed that all subunits of the glycosylphosphatidylinositol (GPI)-anchor transamidase complex were overexpressed in metastatic melanoma cells and identified altered phosphoprotein modules such as the BAF complex and mRNA splicing between metastatic and primary cells. This study provides a high-quality resource for calibrating DIA-MS performance, benchmarking DIA bioinformatic algorithms, and exploring the metastatic proteotypes in melanoma cells.

The data-independent acquisition (DIA) performed in the latest high-resolution, high-speed mass spectrometers offers a powerful analytical tool for biological investigations. The DIA mass spectrometry (DIA-MS) combined with the isotopic labeling approach holds a particular promise for increasing the multiplexity of DIA-MS analysis, which could assist the relative protein quantification and the proteome-wide turnover profiling. However, the wide MS1 isolation windows employed in conventional DIA methods lead to a limited efficiency in identifying and quantifying isotope-labeled peptide pairs through peptide fragment ions. Here, we optimized a high-selectivity DIA-MS named BoxCarmax that supports the analysis of complex samples, such as those generated from Stable isotope labeling by amino acids in cell culture (SILAC) and pulse SILAC (pSILAC) experiments. BoxCarmax enables multiplexed acquisition at both MS1 and MS2 levels, through the integration of BoxCar and MSX features, as well as a gas-phase separation strategy. We found BoxCarmax significantly improved the quantitative accuracy in SILAC and pSILAC samples by mitigating the ratio suppression of isotope-peptide pairs. We further applied BoxCarmax to measure protein degradation regulation during serum starvation stress in cultured cells, revealing valuable biological insights. Our study offered an alternative and accurate approach for the MS analysis of protein turnover and complex samples.

To date, the effects of specific modification types and sites on protein lifetime have not been systematically illustrated. Here, we describe a proteomic method, DeltaSILAC, to quantitatively assess the impact of site-specific phosphorylation on the turnover of thousands of proteins in live cells. Based on the accurate and reproducible mass spectrometry-based method, a pulse labeling approach using stable isotope-labeled amino acids in cells (pSILAC), phosphoproteomics, and a unique peptide-level matching strategy, our DeltaSILAC profiling revealed a global, unexpected delaying effect of many phosphosites on protein turnover. We further found that phosphorylated sites accelerating protein turnover are functionally selected for cell fitness, enriched in Cyclin-dependent kinase substrates, and evolutionarily conserved, whereas the glutamic acids surrounding phosphosites significantly delay protein turnover. Our method represents a generalizable approach and provides a rich resource for prioritizing the effects of phosphorylation sites on protein lifetime in the context of cell signaling and disease biology.

The development of self-chargeable lithium-ion batteries is of great significance for expanding the usable range of the lithium-ion battery and it has received intensive attention from numerous researchers.

Post-translational modifications such as phosphorylation can have profound effects on the physicochemical and biological properties of proteins. However, high-throughput and systematic approaches have not yet been developed to assess the effects of specific modification types and sites on protein lifetime, which represents a key parameter for understanding signaling rewiring and drug development. Here we describe a proteomic method, DeltaSILAC, to quantify the impact of site-specific phosphorylation on the endurance of thousands of proteins in live cells. Being configured on the reproducible data-independent acquisition mass spectrometry (DIA-MS), the pulse labeling approach using stable isotope-labeled amino acids in cells (SILAC), together with a novel peptide-level matching strategy, this multiplexed assay revealed the global delaying effect of phosphorylation on protein turnover in growing cancer cells. Further, we identified local sequence and structural features in proximity to the phosphorylated sites that could be associated with protein endurance alterations. We found that phosphorylated sites accelerating protein turnover are functionally selected for cell fitness and evolutionarily conserved. DeltaSILAC provides a generalizable approach for prioritizing the effects of phosphorylation sites on protein lifetime in the context of cell signaling and disease biology, which is highly complementary to existing methods. Finally, DeltaSILAC is widely applicable to diverse post-translational modification types and different cell systems.

The ionic effects of three halide salts (NaI, NaCl and NaBr) on thermosensitive poly(N-isopropylacrylamide) and poly(N,N-diethylacrylamide) have been investigated by NMR, DFT-D calculations and DLS.

Mass spectrometry (MS) quantification has become an essential tool for both modern biology and translational medicine research. Data-independent acquisition (DIA) has emerged recently and combines the merits of data-dependent acquisition (DDA) methods and selected reaction monitoring (SRM) approaches. Unfortunately, making a quantitative comparison between samples in a single DIA injection is extremely restricted due to its poor compatibility with the traditional DDA labeling approach. As a consequence, a multiplex quantitative reagent suitable for the DIA strategy would be strongly desirable to improve these uninformative situations. As counterparts to traditional DDA quantitative reagents, such as tandem mass tags (TMT) and isobaric tags for relative and absolute quantification (iTRAQ), here, we have designed a novel mass-defect-based carbonyl activated tag (mdCAT) allowing the DIA method to quantify eight samples in parallel for the first time. The integration of the mdCATs with the DIA strategy can not only allow the popular and powerful merits of the DIA strategy, such as broad proteomic coverage, good reproducibility, and accuracy, but it can also eliminate spectral complexity due to their minuscule mass differences. In doing so, it offers new opportunities for the investigation of clinical samples, driving the field of DIA toward unprecedented multiplexing and improving its ability to search for diagnostic biomarkers.

The scale-up application of dye-sensitized solar cells (DSSCs) will desire the efficient and low-cost electrocatalytic counter electrodes. Carbon materials derived from the natural materials have been extensively used in the new energy fields owing to numerously distinct advantages. In this work, a new-style carbon matrix incorporated with Ni species is manufactured through pyrolyzing the humic acid-Ni complex compounds under an inert atmosphere. Systematic electrochemical measurements reveal that the Ni-incorporated carbon holds an elevated triiodide reducing properties in contrast to the single carbonized humic acid. The improvement of electrocatalytic ability may be ascribed to Ni incorporated in carbon matrix that offer more electroactive sites. As a result, when the prepared Ni-incorporated material serves as electrocatalyst in iodide-mediated DSSC, the corresponding device yields a PCE of 7.01%, which increases by 14% in contrast to the single carbonized humic acid (6.14%) and is close to that (7.1%) of as-reference Pt electrode. This study indicates that the proposed carbonaceous material may be in favour of large-scale production of low cost and high efficiency DSSCs.

In this research, the bimetallic NiCoP nanoparticles and NiCoP/carbon nanotubes (CNTs) composites were successfully synthesized, and their performances as counter electrodes (CEs) for dye sensitized solar cells were investigated. The optimal composite CE (NiCoP-CNTs-3) exhibits outstanding photoelectric conversion efficiency (PCE) of 7.24%, which exceeds the single NiCoP (4.71%) or CNTs electrode (6.05%) and is on par with the standard Pt CE (7.12%) under same conditions. The electrochemical properties of NiCoP-CNTs electrodes were also characterized by utilizing the electrochemical impedance spectroscopy (EIS), Tafel polarization curves and cyclic-voltammetry (CV). The results indicate that NiCoP-CNTs-3 electrode holds Pt-like good electrocatalytic activity towards reduction of I3−. In addtion, the composite CE shows a reliably electrochemical stability. The present materials in this study may pave a way for the practical applications of Pt-free DSSCs.

Protein sialylation is ubiquitous and essential in a wide range of biological processes. Herein, a mass defect-based chemical-directed proteomics method (MdCDPM) was presented for targeted analysis of intact sialylglycopeptides (SGPs). The process starts by specific oxidation of dihydroxy in sialic acid to aldehyde, which was then chemically labeled by two arginine isotopologues (Arg-15N4 and Arg-D4, differs by 36 mDa). The equally mixed precursor partners, spacing tens of mDa apart, enable the direct recognition of SGPs in MS1 level and benefit the subsequent targeted MS2 characterization. The mass envelope of two labeled forms falling into a narrow m/z window strengthens recognition uniqueness greatly, and the proposed 1:1 intensity ratio of doublets will not be readily distorted. More important, such subtle mass differences permit multiple sialic acids labeling without additional complexity of precursor patterns. Also, the partner m/z shifts detail the number of sialic acids contained in the precursor species. By applying MdCDPM, femtomole quantities of SGPs could be detected from total cell lysates, even at a signal-to-noise ratio of as low as 3:1. In addition, assays were performed to estimate the false positive rate and demonstrated high confidence of MdCDPM. Furthermore, it was designed and successfully exploited to analyze SGPs in human serum, which highlighted the feasibility of this strategy for biological applications.

Excellent electrocatalytic properties and low cost are main requirements for the suitable counter electrodes of dye sensitized solar cells (DSSCs). In this work, serials of electrocatalytic films were facilely fabricated by blending the PEDOT polymers and transition metal phosphides nanoparticles (Ni2P or Co2P). All yielded films as counter electrodes were assembled into intact cells and their electrocatalytic performances were carefully investigated. Compared to the PEDOT-only electrode or phosphide-only electrode, the achieved composite electrodes display the superior electrocatalytic performances. The continuous PEDOT polymers can significantly alleviate phosphides nanoparticles aggregations and thereby generate more active area for redox couple. DSSCs with the optimal PEDOT-Ni2P-3 electrode and PEDOT-Co2P-3 electrode show the power conversion efficiency (PCE) of 7.14% and PCE of 6.85%, respectively. And PCE value of the optimal PEDOT-Ni2P-3 is comparable to that (7.09%) of Pt electrode on the same test condition. Moreover, electrochemical measurements reveal the type of composite electrode holding decent dissolution-resistant ability. Considering the outstanding and available electrocatalytic activities, the potential application of PEDOT-phosphides electrodes as the substitute of Pt electrode is worth to be expected.

4-Hydroxy-2-nonenal (HNE)-modified proteins are closely associated with cellular functions and diseases, so qualitative and quantitative analysis of HNE-modified proteins is very necessary in order to further understand their structures and molecular functions. In this study, we described a six-plex isobaric labeling affinity purification (SiLAP) method based on the interaction of aminoxyTMT six-plex and anti-TMT antibody resin to identify and quantify the HNE modifications simultaneously. The labeling efficiency, ionization efficiency of the aminoxyTMT-tagged peptides, and reliability of the quantification method were investigated in detail. The mass tags were labeled on the modification sites, which could also significantly increase the ionization efficiency, contributing to site-specific identification and quantification of HNE peptides. The SiLAP strategy possessed high sensitivity, accuracy, and good reproducibility to qualitatively and quantitatively analyze HNE-modified proteins/peptides, which could be used to analyze both endogenously and exogenously modified proteins. Using the SiLAP strategy, 2257 HNE-modified peptides mapping 1121 proteins were collectively quantified, which was the largest data set of HNE-modified proteins with detailed modification sites, and 101 proteins were found to be differentially modified by HNE in six liver cell lines. At the same time, 33 endogenously HNE-modified peptides mapping 33 proteins were identified with modification sites.