Cell cycle progression of under-replicated cells.

Cell cycle checkpoints are the regulatory mechanisms that secure the strict order of cellular events for cell division that ensure genome integrity. It has been proposed that mitosis initiation depends on the completion of DNA replication, which must be tightly controlled to guarantee genome duplication. Contrary to these conventional hypotheses, we showed here that cells were able to enter mitosis without completion of DNA replication.

DePARylation is critical for S phase progression and cell survival.

Poly(ADP-ribose)ylation or PARylation by PAR polymerase 1 (PARP1) and dePARylation by poly(ADP-ribose) glycohydrolase (PARG) are equally important for the dynamic regulation of DNA damage response. PARG, the most active dePARylation enzyme, is recruited to sites of DNA damage via pADPr-dependent and PCNA-dependent mechanisms. Targeting dePARylation is considered an alternative strategy to overcome PARP inhibitor resistance.

Proteomic analysis of ferroptosis pathways reveals a role of CEPT1 in suppressing ferroptosis.

Ferroptosis has been recognized as a unique cell death modality driven by excessive lipid peroxidation and unbalanced cellular metabolism. In this study, we established a protein interaction landscape for ferroptosis pathways through proteomic analyses, and identified choline/ethanolamine phosphotransferase 1 (CEPT1) as a lysophosphatidylcholine acyltransferase 3 (LPCAT3)-interacting protein that regulates LPCAT3 protein stability. In contrast to its known role in promoting phospholipid synthesis, we showed that CEPT1 suppresses ferroptosis potentially by interacting with phospholipases and breaking down certain pro-ferroptotic polyunsaturated fatty acid (PUFA)-containing phospholipids.

DePARylation is critical for S phase progression and cell survival.

Poly(ADP-ribose)ylation or PARylation by PAR polymerase 1 (PARP1) and dePARylation by poly(ADP-ribose) glycohydrolase (PARG) are equally important for the dynamic regulation of DNA damage response. PARG, the most active dePARylation enzyme, is recruited to sites of DNA damage via pADPr-dependent and PCNA-dependent mechanisms. Targeting dePARylation is considered an alternative strategy to overcome PARP inhibitor resistance.

FACS-based genome-wide CRISPR screens define key regulators of DNA damage signaling pathways.

DNA damage-activated signaling pathways are critical for coordinating multiple cellular processes, which must be tightly regulated to maintain genome stability. To provide a comprehensive and unbiased perspective of DNA damage response (DDR) signaling pathways, we performed 30 fluorescence-activated cell sorting (FACS)-based genome-wide CRISPR screens in human cell lines with antibodies recognizing distinct endogenous DNA damage signaling proteins to identify critical regulators involved in DDR. We discovered that proteasome-mediated processing is an early and prerequisite event for cells to trigger camptothecin- and etoposide-induced DDR signaling.

Genome-Wide CRISPR Screens Reveal ZATT as a Synthetic Lethal Target of TOP2-Poison Etoposide That Can Act in a TDP2-Independent Pathway.

Etoposide (ETO) is an anticancer drug that targets topoisomerase II (TOP2). It stabilizes a normally transient TOP2-DNA covalent complex (TOP2cc), thus leading to DNA double-strand breaks (DSBs). Tyrosyl-DNA phosphodiesterases two (TDP2) is directly involved in the repair of TOP2cc by removing phosphotyrosyl peptides from 5'-termini of DSBs.

Actin cytoskeleton vulnerability to disulfide stress mediates disulfidptosis.

SLC7A11-mediated cystine uptake suppresses ferroptosis yet promotes cell death under glucose starvation; the nature of the latter cell death remains unknown. Here we show that aberrant accumulation of intracellular disulfides in SLC7A11 cells under glucose starvation induces a previously uncharacterized form of cell death distinct from apoptosis and ferroptosis. We term this cell death disulfidptosis.

USP7 substrates identified by proteomics analysis reveal the specificity of USP7.

Deubiquitylating enzymes (DUBs) remove ubiquitin chains from proteins and regulate protein stability and function. USP7 is one of the most extensively studied DUBs, since USP7 has several well-known substrates important for cancer progression, such as MDM2, N-MYC, and PTEN. Thus, USP7 is a promising drug target.

Discovery of toxoflavin, a potent IRE1α inhibitor acting through structure-dependent oxidative inhibition.

Inositol-requiring enzyme 1α (IRE1α) is the most conserved endoplasmic reticulum (ER) stress sensor with two catalytic domains, kinase and RNase, in its cytosolic portion. IRE1α inhibitors have been used to improve existing clinical treatments against various cancers. In this study we identified toxoflavin (TXF) as a new-type potent small molecule IRE1α inhibitor.

Integrated screens uncover a cell surface tumor suppressor gene involved in Hippo pathway.

Cell surface proteins play essential roles in various biological processes and are highly related to cancer development. They also serve as important markers for cell identity and targets for pharmacological intervention. Despite their great potentials in biomedical research, comprehensive functional analysis of cell surface proteins remains scarce.

Genome-wide CRISPR screens using isogenic cells reveal vulnerabilities conferred by loss of tumor suppressors.

Exploiting cancer vulnerabilities is critical for the discovery of anticancer drugs. However, tumor suppressors cannot be directly targeted because of their loss of function. To uncover specific vulnerabilities for cells with deficiency in any given tumor suppressor(s), we performed genome-scale CRISPR loss-of-function screens using a panel of isogenic knockout cells we generated for 12 common tumor suppressors.

Beyond proteins: Ubiquitylation of lipopolysaccharide to fight bacteria.

Mechanisms and functions of protein ubiquitylation and LPS ubiquitylation. LPS ubiquitylation serves as a scaffold to recruit E3 ligases for the ubiquitylation of the membrane of bacteria. Polyubiquitin coat on the bacterial cell surface is one type of "eat-me" signal recognized by the host cells.

Low-density-lipoprotein-receptor-related protein 1 mediates Notch pathway activation.

The Notch signaling pathway controls cell growth, differentiation, and fate decisions, and its dysregulation has been linked to various human genetic disorders and cancers. To comprehensively understand the global organization of the Notch pathway and identify potential drug targets for Notch-related diseases, we established a protein interaction landscape for the human Notch pathway. By combining and analyzing genetic and phenotypic data with bioinformatics analysis, we greatly expanded this pathway and identified many key regulators, including low-density-lipoprotein-receptor-related protein 1 (LRP1).

Genetic vulnerabilities upon inhibition of DNA damage response.

Because of essential roles of DNA damage response (DDR) in the maintenance of genomic integrity, cellular homeostasis, and tumor suppression, targeting DDR has become a promising therapeutic strategy for cancer treatment. However, the benefits of cancer therapy targeting DDR are limited mainly due to the lack of predictive biomarkers. To address this challenge, we performed CRISPR screens to search for genetic vulnerabilities that affect cells' response to DDR inhibition.

Interactomes of SARS-CoV-2 and human coronaviruses reveal host factors potentially affecting pathogenesis.

Host-virus protein-protein interactions play key roles in the life cycle of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2). We conducted a comprehensive interactome study between the virus and host cells using tandem affinity purification and proximity-labeling strategies and identified 437 human proteins as the high-confidence interacting proteins. Further characterization of these interactions and comparison to other large-scale study of cellular responses to SARS-CoV-2 infection elucidated how distinct SARS-CoV-2 viral proteins participate in its life cycle.

Non-canonical function of DGCR8 in DNA double-strand break repair signaling and tumor radioresistance.

In response to DNA double-strand breaks (DSBs), repair proteins are recruited to the damaged sites. Ubiquitin signaling plays a critical role in coordinating protein recruitment during the DNA damage response. Here, we find that the microRNA biogenesis factor DGCR8 promotes tumor resistance to X-ray radiation independently of its Drosha-binding ability.

mTORC1 couples cyst(e)ine availability with GPX4 protein synthesis and ferroptosis regulation.

Glutathione peroxidase 4 (GPX4) utilizes glutathione (GSH) to detoxify lipid peroxidation and plays an essential role in inhibiting ferroptosis. As a selenoprotein, GPX4 protein synthesis is highly inefficient and energetically costly. How cells coordinate GPX4 synthesis with nutrient availability remains unclear.

Selective Labeling and Identification of the Tumor Cell Proteome of Pancreatic Cancer .

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.

Bufalin exerts antitumor effects in neuroblastoma via the induction of reactive oxygen species‑mediated apoptosis by targeting the electron transport chain.

The prognosis of high‑risk neuroblastoma remains poor. Clinical first‑line drugs for treating neuroblastoma have been developed over the previous half‑century; however, progress in the identification of new drugs with high efficiency is required. Bufalin, one of the major components of extracts obtained from the venom of the Chinese toad Bufo gargarizans, which is used to treat heart failure in Asian Pacific countries, has been reported to be a potential drug against multiple types of tumor; however, the detailed mechanisms underlying its antitumor activities remain unclear, largely due to lack of knowledge regarding its targets.

Extracellular signal-regulated kinases associate with and phosphorylate DHPS to promote cell proliferation.

The ERK1/2 pathway is one of the most commonly dysregulated pathways in human cancers and controls many vital cellular processes. Although many ERK1/2 kinase substrates have been identified, the diversity of ERK1/2 mediated processes suggests the existence of additional targets. Here, we identified Deoxyhypusine synthase (DHPS), an essential hypusination enzyme regulating protein translation, as a major and direct-binding protein of ERK1/2.

Proteome-wide Analysis Reveals Substrates of E3 Ligase RNF146 Targeted for Degradation.

Specific E3 ligases target tumor suppressors for degradation. Inhibition of such E3 ligases may be an important approach to cancer treatment. RNF146 is a RING domain and PARylation-dependent E3 ligase that functions as an activator of the β-catenin/Wnt and YAP/Hippo pathways by targeting the degradation of several tumor suppressors.