Multiple roles of arsenic compounds in phase separation and membraneless organelles formation determine their therapeutic efficacy in tumors.

Arsenic compounds are widely used for the therapeutic intervention of multiple diseases. Ancient pharmacologists discovered the medicinal utility of these highly toxic substances, and modern pharmacologists have further recognized the specific active ingredients in human diseases. In particular, Arsenic trioxide (ATO), as a main component, has therapeutic effects on various tumors (including leukemia, hepatocellular carcinoma, lung cancer, etc.). However, its toxicity limits its efficacy, and controlling the toxicity has been an important issue. Interestingly, recent evidence has pointed out the pivotal roles of arsenic compounds in phase separation and membraneless organelles formation, which may determine their toxicity and therapeutic efficacy. Here, we summarize the arsenic compounds-regulating phase separation and membraneless organelles formation. We further hypothesize their potential involvement in the therapy and toxicity of arsenic compounds, highlighting potential mechanisms underlying the clinical application of arsenic compounds.

Exosomal miR-486 derived from bone marrow mesenchymal stem cells promotes angiogenesis following cerebral ischemic injury by regulating the PTEN/Akt pathway.

Bone marrow mesenchymal stem cell-derived exosomes (BMSC-Exos) have been shown to promote angiogenesis after ischemic stroke, in which microRNAs (miRs) are believed to play an important role in exosome-mediated therapeutic effects, though the mechanism is still not clear. In this study, a series of molecular biological and cellular assays, both in vitro and in vivo, were performed to elucidate the role of exosomal miR-486 in angiogenesis following cerebral ischemic and its molecular mechanisms. Our results revealed that BMSC-Exos significantly improved neurological function and increased microvessel density in ischemic stroke rats. In vitro assays showed that BMSC-Exos promoted the proliferation, migration, and tube formation ability of oxygen-glucose deprivation/reoxygenation (OGD/R) injured rat brain microvascular endothelial cells (RBMECs). Importantly, BMSC-Exos increased the expression of miR-486 and phosphorylated protein kinase B (p-Akt) and down-regulated the protein level of phosphatase and tensin homolog (PTEN) in vivo and in vitro. Mechanistic studies demonstrated that transfection with miR-486 mimic enhanced RBMECs angiogenesis and increased p-Akt expression, while inhibited PTEN expression. On the other hand, the miR-486 inhibitor induced an opposite effect, which could be blocked by PTEN siRNA. It was thus concluded that exosomal miR-486 from BMSCs may enhance the functional recovery by promoting angiogenesis following cerebral ischemic injury, which might be related to its regulation of the PTEN/Akt pathway.

Vitamin C inactivates c-Jun N-terminal kinase to stabilize heart and neural crest derivatives expressed 1 (Hand1) in regulating placentation and maintenance of pregnancy.

Vitamin C (VC) serves as a pivotal nutrient for anti-oxidation process, metabolic responses, and stem cell differentiation. However, its precise contribution to placenta development and gestation remains obscure. Here, we demonstrated that physiological levels of VC act to stabilize Hand1, a key bHLH transcription factor vital for the development trajectory of trophoblast giant cell (TGC) lineages, thereby promoting the differentiation of trophoblast stem cells into TGC. Specifically, VC administration inactivated c-Jun N-terminal kinase (JNK) signaling, which directly phosphorylates Hand1 at Ser48, triggering the proteasomal degradation of Hand1. Conversely, a loss-of-function mutation at Ser48 on Hand1 not only significantly diminished both intrinsic and VC-induced stabilization of Hand1 but also underscored the indispensability of this residue. Noteworthy, the insufficiency of VC led to severe defects in the differentiation of diverse TGC subtypes and the formation of labyrinth's vascular network in rodent placentas, resulting in failure of maintenance of pregnancy. Importantly, VC deficiency, lentiviral knockdown of JNK or overexpression of Hand1 mutants in trophectoderm substantially affected the differentiation of primary and secondary TGC in E8.5 mouse placentas. Thus, these findings uncover the significance of JNK inactivation and consequential stabilization of Hand1 as a hitherto uncharacterized mechanism controlling VC-mediated placentation and perhaps maintenance of pregnancy.

Smoking-induced CCNA2 expression promotes lung adenocarcinoma tumorigenesis by boosting AT2/AT2-like cell differentiation.

Lung adenocarcinoma (LUAD), a type of non-small cell lung cancer (NSCLC), originates from not only bronchial epithelial cells but also alveolar type 2 (AT2) cells, which could differentiate into AT2-like cells. AT2-like cells function as cancer stem cells (CSCs) of LUAD tumorigenesis to give rise to adenocarcinoma. However, the mechanism underlying AT2 cell differentiation into AT2-like cells in LUAD remains unknown. We analyze genes differentially expressed and genes with significantly different survival curves in LUAD, and the combination of these two analyses yields 147 differential genes, in which 14 differentially expressed genes were enriched in cell cycle pathway. We next analyze the protein levels of these genes in LUAD and find that Cyclin-A2 (CCNA2) is closely associated with LUAD tumorigenesis. Unexpectedly, high CCNA2 expression in LUAD is restrictedly associated with smoking and independent of other driver mutations. Single-cell sequencing analyses reveal that CCNA2 is predominantly involved in AT2-like cell differentiation, while inhibition of CCNA2 significantly reverses smoking-induced AT2-like cell differentiation. Mechanistically, CCNA2 binding to CDK2 phosphorylates the AXIN1 complex, which in turn induces ubiquitination-dependent degradation of ß-catenin and inhibits the WNT signaling pathway, thereby failing AT2 cell maintenance. These results uncover smoking-induced CCNA2 overexpression and subsequent WNT/ß-catenin signaling inactivation as a hitherto uncharacterized mechanism controlling AT2 cell differentiation and LUAD tumorigenesis.

Smurf1 polyubiquitinates on K285/K282 of the kinases Mst1/2 to attenuate their tumor-suppressor functions.

Sterile 20-like kinases Mst1 and Mst2 (Mst1/2) and large tumor suppressor 1/2 are core kinases to mediate Hippo signaling in maintaining tissue homeostasis. We have previously demonstrated that Smad ubiquitin (Ub) regulatory factor 1 (Smurf1), a HECT-type E3 ligase, ubiquitinates and in turn destabilizes large tumor suppressor 1/2 to induce the transcriptional output of Hippo signaling. Here, we unexpectedly find that Smurf1 interacts with and polyubiquitinates Mst1/2 by virtue of K27- and K29-linked Ub chains, resulting in the proteasomal degradation of Mst1/2 and attenuation of their tumor-suppressor functions. Among the potential Ub acceptor sites on Mst1/2, K285/K282 are conserved and essential for Smurf1-induced polyubiquitination and degradation of Mst1/2 as well as transcriptional output of Hippo signaling. As a result, K285R/K282R mutation of Mst1/2 not only negates the transcriptional output of Hippo signaling but enhances the tumor-suppressor functions of Mst1/2. Together, we demonstrate that Smurf1-mediated polyubiquitination on K285/K282 of Mst1/2 destabilizes Mst1/2 to attenuate their tumor-suppressor functions. Thus, the present study identifies Smurf1-mediated ubiquitination of Mst1/2 as a hitherto uncharacterized mechanism fine-tuning the Hippo signaling pathway and may provide additional targets for therapeutic intervention of diseases associated with this important pathway.

SUMOylation of Rho-associated protein kinase 2 induces goblet cell metaplasia in allergic airways.

Allergic asthma is characterized by goblet cell metaplasia and subsequent mucus hypersecretion that contribute to the morbidity and mortality of this disease. Here, we explore the potential role and underlying mechanism of protein SUMOylation-mediated goblet cell metaplasia. The components of SUMOylaion machinery are specifically expressed in healthy human bronchial epithelia and robustly upregulated in bronchial epithelia of patients or mouse models with allergic asthma. Intratracheal suppression of SUMOylation by 2-D08 robustly attenuates not only allergen-induced airway inflammation, goblet cell metaplasia, and hyperreactivity, but IL-13-induced goblet cell metaplasia. Phosphoproteomics and biochemical analyses reveal SUMOylation on K1007 activates ROCK2, a master regulator of goblet cell metaplasia, by facilitating its binding to and activation by RhoA, and an E3 ligase PIAS1 is responsible for SUMOylation on K1007. As a result, knockdown of PIAS1 in bronchial epithelia inactivates ROCK2 to attenuate IL-13-induced goblet cell metaplasia, and bronchial epithelial knock-in of ROCK2(K1007R) consistently inactivates ROCK2 to alleviate not only allergen-induced airway inflammation, goblet cell metaplasia, and hyperreactivity, but IL-13-induced goblet cell metaplasia. Together, SUMOylation-mediated ROCK2 activation is an integral component of Rho/ROCK signaling in regulating the pathological conditions of asthma and thus SUMOylation is an additional target for the therapeutic intervention of this disease.

Multiple post-translational modifications ensure EGFR functionality: Potential therapeutic targets to overcome its drug-resistance mutations.

Epidermal growth factor receptor (EGFR) mutation is the most common driver mutation in non-small cell lung cancer (NSCLC). The first-line therapy for advanced NSCLC patients with EGFR-sensitive mutation is the EGFR tyrosine kinase inhibitor (EGFR-TKI). However, most NSCLC patients with EGFR mutation will develop resistant mutations in EGFR-TKI therapy. With further studies, resistance mechanisms represented by EGFR-T790M mutations have revealed the impact of EGFR mutations in situ on EGFR-TKIs sensitivity. The third-generation EGFR-TKIs inhibit both EGFR-sensitive mutations and T790M mutations. The emergence of novel mutations such as EGFR-C797S and EGFR-L718Q may decrease efficacy. Searching for new targets to overcome EGFR-TKI resistance becomes a key challenge. Therefore, an in-depth understanding of the regulatory mechanisms of EGFR is essential to find novel targets to overcome drug-resistant mutations in EGFR-TKIs. EGFR, as a receptor-type tyrosine kinase, undergoes homo/heterodimerization and autophosphorylation upon binding to ligands, which activates multiple downstream signaling pathways. Interestingly, there is growing evidence that the kinase activity of EGFR is affected not only by phosphorylation but also by various post-translational modifications (PTMs, such as S-palmitoylation, S-nitrosylation, Methylation, etc.). In this review, we systematically review the effects of different protein PTMs on EGFR kinase activity and its functionality and suggest that influencing EGFR kinase activity by modulating multiple EGFR sites are potential targets to overcome EGFR-TKIs resistance mutations.

SUMOylation activates large tumour suppressor 1 to maintain the tissue homeostasis during Hippo signalling.

Large tumour suppressor (LATS) 1/2, the core kinases of Hippo signalling, are critical for maintaining tissue homeostasis. Here, we investigate the role of SUMOylation in the regulation of LATS activation. High cell density induces the expression of components of the SUMOylation machinery and enhances the SUMOylation and activation of Lats1 but not Lats2, whereas genetic deletion of the SUMOylation E2 ligase, Ubc9, abolishes this Lats1 activation. Moreover, SUMOylation occurs at the K830 (mouse K829) residue to activate LATS1 and depends on the PIAS1/2 E3 ligase. Whereas the K830 deSUMOylation mutation of LATS1 found in the human metastatic prostate cancers eliminates the kinase activity by attenuating the formation of the phospho-MOB1/phospho-LATS1 complex. As a result, the LATS1(K830R) transgene phenocopies Yap transgene to cause the oversized livers in mice, whereas Lats1(K829R) knock-in phenocopies the deletion of Lats1 in causing the reproductive and endocrine defects and ovary tumours in mice. Thus, SUMOylation-mediated LATS1 activation is an integral component of Hippo signalling in the regulation of tissues homeostasis.