Extra (embryonic) dialogues: Keys to improved stem cell-based embryo models.

Despite advances in stem cell cultures, modeling early human development with stem cells in a dish remains challenging. Research by Hislop et al., Okubo et al.

βTrCP1 promotes SLC35F2 protein ubiquitination and inhibits cancer progression in HeLa cells.

The solute carrier family 35 F2 (SLC35F2) belongs to membrane-bound carrier proteins that are associated with multiple cancers. The main factor that determines cancer progression is the expression level of SLC35F2. Thus, identifying the E3 ligase that controls SLC35F2 protein abundance in cancer cells is critical.

Quantification of Genetic Heterogeneity Using Long-Read Targeted Individual DNA Molecule Sequencing.

Understanding genetic heterogeneity is of paramount importance in unraveling the intricate functioning of biological systems, as it contributes to the diversity of phenotypes of gene-environment interactions. We have developed a method termed targeted Individual DNA Molecule Sequencing (IDMseq) to accurately quantify genetic heterogeneity within cell populations, even those with rare variants present at low frequencies. IDMseq ensures that each original DNA molecule is distinctively represented by one unique molecule identifier (UMI) group, preventing false UMI groups and enabling precise quantification of allele frequency within the original population.

E3 ubiquitin ligase APC/C regulates SLC35F2 protein turnover and inhibits cancer progression in HeLa cells.

Background: The solute carrier family 35 F2 (SLC35F2), belongs to membrane-bound carrier proteins that control various physiological functions and are activated in several cancers. However, the molecular mechanism regulating SLC35F2 protein turnover and its implication in cancer progression remains unexplored. Therefore, screening for E3 ligases that promote SLC35F2 protein degradation is essential during cancer progression.

USP19 Negatively Regulates p53 and Promotes Cervical Cancer Progression.

p53 is a tumor suppressor gene activated in response to cellular stressors that inhibits cell cycle progression and induces pro-apoptotic signaling. The protein level of p53 is well balanced by the action of several E3 ligases and deubiquitinating enzymes (DUBs). Several DUBs have been reported to negatively regulate and promote p53 degradation in tumors.

CRISPR/Cas9-based genome-wide screening for deubiquitinase subfamily identifies USP1 regulating MAST1-driven cisplatin-resistance in cancer cells.

Cisplatin is one of the frontline anticancer agents. However, development of cisplatin-resistance limits the therapeutic efficacy of cisplatin-based treatment. The expression of microtubule-associated serine/threonine kinase 1 (MAST1) is a primary factor driving cisplatin-resistance in cancers by rewiring the MEK pathway.

Dual role of deubiquitinating enzyme USP19 regulates mitotic progression and tumorigenesis by stabilizing survivin.

Survivin is a component of the chromosomal passenger complex, which includes Aurora B, INCENP, and Borealin, and is required for chromosome segregation and cytokinesis. We performed a genome-wide screen of deubiquitinating enzymes for survivin. For the first time, we report that USP19 has a dual role in the modulation of mitosis and tumorigenesis by regulating survivin expression.

The role of the CRISPR-Cas system in cancer drug development: Mechanisms of action and therapy.

Background: The recent emergence of gene editing using Clustered regularly interspaced short palindromic repeats (CRISPR)-CRISPR associated system (Cas) tools and advances in genomics and proteomics has revolutionized drug discovery and personalized medicine.

Purpose And Scope: The CRISPR-Cas system has enabled gene and cell-based therapies, screening for novel drug targets, a new generation of disease models, elucidation of drug resistance mechanisms, and drug efficacy testing. Here, we summarized recent investigations and strategies involved in cancer-related drug discovery using the CRISPR-Cas system.

USP32 confers cancer cell resistance to YM155 via promoting ER-associated degradation of solute carrier protein SLC35F2.

The most commonly preferred chemotherapeutic agents to treat cancers are small-molecule drugs. However, the differential sensitivity of various cancer cells to small molecules and untargeted delivery narrow the range of potential therapeutic applications. The mechanisms responsible for drug resistance in a variety of cancer cells are also largely unknown.

Smart Vitamin Micelles as Cancer Nanomedicines for Enhanced Intracellular Delivery of Doxorubicin.

Chemotherapy is one of the most effective treatments for cancer. However, intracellular delivery of many anticancer drugs is hindered by their hydrophobicity and low molecular weight. Here, we describe highly biocompatible and biodegradable amphiphilic vitamin conjugates comprising hydrophobic vitamin E and hydrophilic vitamin B labeled with dual pH and glutathione-responsive degradable linkages.

Ubiquitin Specific Protease 29 Functions as an Oncogene Promoting Tumorigenesis in Colorectal Carcinoma.

Colorectal carcinoma is the third foremost cause of cancer-related deaths and accounts for 5.8% of all deaths globally. The molecular mechanisms of colon cancer progression and metastasis control are not well studied.

Ubiquitin-Specific Protease 29 Regulates Cdc25A-Mediated Tumorigenesis.

Cell division cycle 25A (Cdc25A) is a dual-specificity phosphatase that is overexpressed in several cancer cells and promotes tumorigenesis. In normal cells, Cdc25A expression is regulated tightly, but the changes in expression patterns in cancer cells that lead to tumorigenesis are unknown. In this study, we showed that ubiquitin-specific protease 29 (USP29) stabilized Cdc25A protein expression in cancer cell lines by protecting it from ubiquitin-mediated proteasomal degradation.

E3 Ubiquitin Ligase APC/C Regulation of Phenylalanine Hydroxylase Stability and Function.

Phenylketonuria (PKU) is an autosomal recessive metabolic disorder caused by the dysfunction of the enzyme phenylalanine hydroxylase (PAH). Alterations in the level of PAH leads to the toxic accumulation of phenylalanine in the blood and brain. Protein degradation mediated by ubiquitination is a principal cellular process for maintaining protein homeostasis.

YM155 sensitizes HeLa cells to TRAIL-mediated apoptosis via cFLIP and survivin downregulation.

Tumor necrosis factor-related apoptosis inducing ligand (TRAIL)-mediated apoptosis is a safe method for the treatment of various types of cancer. However, TRAIL therapy is less effective in certain types of cancer, including cervical cancer. To address this problem, a combinatorial approach was employed to sensitize cervical cancer at low dosages.

HAUSP stabilizes Cdc25A and protects cervical cancer cells from DNA damage response.

Resistance to DNA-damaging agents is one of the main reasons for the low survival of cervical cancer patients. Previous reports have suggested that the Cdc25A oncoprotein significantly affects the level of susceptibility to DNA-damaging agents, but the molecular mechanism remains unclear. In this study, we used Western blot and flow cytometry analyses to demonstrate that the deubiquitinating enzyme HAUSP stabilizes Cdc25A protein level.

Genome-scale screening of deubiquitinase subfamily identifies USP3 as a stabilizer of Cdc25A regulating cell cycle in cancer.

Conventional screening methods for deubiquitinating enzymes (DUBs) have important limitations. A loss-of-function study based on the knockout of DUB genes in mammalian cells can provide an excellent model for exploring DUB function. Here, we used CRISPR-Cas9 to perform genome-scale knockout of the entire set of genes encoding ubiquitin-specific proteases (USPs), a DUB subfamily, and then systematically screened for DUBs that stabilize the Cdc25A oncoprotein.

CRISPR-mediated upregulation of DR5 and downregulation of cFLIP synergistically sensitize HeLa cells to TRAIL-mediated apoptosis.

Tumor necrosis factor-related apoptosis-inducing ligand (TRAIL) has received attention as an anticancer therapy because it mediates apoptosis of several cancer cell types but not normal human cell types. In this study, we implemented genome editing techniques to upregulate DR5 and downregulate cFLIP in HeLa cells to stimulate TRAIL-induced apoptosis. We designed and validated sgRNAs to enrich the endogenous level of DR5 by dead Cas9 (dCas9).

Deubiquitinating enzymes as cancer biomarkers: new therapeutic opportunities?

Cancer remains a life-threatening disease and accounts for the major mortality rates worldwide. The practice of using biomarkers for early detection, staging, and customized therapy may increase cancer patients' survival. Deubiquitinating enzymes (DUBs) are a family of proteases that remove ubiquitin tags from proteins of interest undergoing proteasomal degradation.

The stability and oncogenic function of LIN28A are regulated by USP28.

RNA-binding protein LIN28A is often highly expressed in human malignant tumors and is involved in tumor metastasis and poor prognosis. Knowledge about post-translational regulatory mechanisms governing LIN28A protein stability and function is scarce. Here, we investigated the role of ubiquitination and deubiquitination on LIN28A protein stability and report that LIN28A protein undergoes ubiquitination.

Different Methods of Delivering CRISPR/Cas9 Into Cells.

Clustered regularly interspaced short palindromic repeats (CRISPR)/CRISPR-associated system (Cas) is comprised of repetitive bases followed by short fragments of DNA from a previously invading organism that provide immunity to the most prokaryotic organisms. An RNA-dependent spacer is required for CRISPR/Cas9 to recognize the target DNA. Delivery of the CRISPR/Cas9-guide RNA (gRNA) complex to any cell results in modification of the target sequence.

Efficient genome editing by FACS enrichment of paired D10A Cas9 nickases coupled with fluorescent proteins.

Targeted genome editing by clustered regularly interspaced short palindromic repeats (CRISPR-Cas9) raised concerns over off-target effects. The use of double-nicking strategy using paired Cas9 nickase has been developed to minimize off-target effects. However, it was reported that the efficiency of paired nickases were comparable or lower than that of either corresponding nuclease alone.

Genome editing: a robust technology for human stem cells.

Human pluripotent stem cells comprise induced pluripotent and embryonic stem cells, which have tremendous potential for biological and therapeutic applications. The development of efficient technologies for the targeted genome alteration of stem cells in disease models is a prerequisite for utilizing stem cells to their full potential. Genome editing of stem cells is possible with the help of synthetic nucleases that facilitate site-specific modification of a gene of interest.

Concise Review: Fate Determination of Stem Cells by Deubiquitinating Enzymes.

Post-translational modification by ubiquitin molecules is a key regulatory process for stem cell fate determination. Ubiquitination and deubiquitination are the major cellular processes used to balance the protein turnover of several transcription factors that regulate stem cell differentiation. Deubiquitinating enzymes (DUBs), which facilitate the processing of ubiquitin, significantly influence stem cell fate choices.