Inhibition of non-muscular myosin light chain kinase accelerates the clearance of inflammatory cells by promoting the lysosome-mediated cell death.

Infections like COVID-19 are the primary cause of death around the world because they can cause acute lung injury (ALI), acute respiratory distress syndrome (ARDS), and sepsis. Inflammatory cells serve as crucial protective barriers in these diseases. However, excessive accumulation of inflammatory cells is also one of the major causes of organ damage. The non-muscular myosin light chain kinase (nmMLCK) plays crucial of cytoskeletal components involved in endothelial cell-matrix and cell-cell adhesion, integrity, and permeability. Our previous investigations found that ML-7, a specific inhibitor of MLCK, promoted neutrophil apoptosis through various signaling pathways. In this study, we found that knockout of MLCK significantly promote apoptosis of neutrophils and macrophages in the BALF of the LPS-induced ALI, meanwhile it had no effect on the apoptosis of neutrophils in the circulatory system. RNA-sequencing revealed that the effect of MLCK knockout in inducing apoptosis of inflammatory cells was mediated through lysosomes. Administering ML-7 into the lungs significantly promoted neutrophil apoptosis, accelerating their clearance. In the LPS- or CLP-induced sepsis models, ML-7 administration significantly improves the apoptosis of inflammatory cells, especially neutrophils, at the infection site but had no impact on neutrophils in the circulatory system. ML-7 also significantly improved the survival rate of mice with LPS- or CLP-induced sepsis. Taken together, we found that MLCK plays a crucial role in the survival of inflammatory cells at the infection site. Inhibiting MLCK significantly induces apoptosis of inflammatory cells at the infection site, promoting inflammation resolution, with no impact of the circulatory system.

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.

Sonic hedgehog signaling: Alternative splicing and pathogenic role in medulloblastoma.

Alternative splicing (AS) produces the different mRNA splicing bodies, which are then translated into multiple protein isoforms and participate in various biological functions. With a deeper understanding of alternative splicing through the study of transcriptomes using high-throughput sequencing-based methods, the correlation between aberrant AS and diseases triggered a great concern, especially abnormal AS and cancer. Medulloblastoma (MB) is an intracranial tumor in children. Sonic hedgehog MB (SHH-MB) accounted for approximately 30% of MB, which is associated with the activation of SHH signaling. Growing evidence shows that aberrant AS is closely related to the tumorigenesis of MB. Here, we briefly introduced the AS and its mechanism. Next, we described canonical/noncanonical hedgehog signaling and its correlation with MB. The main description focused on AS of various regulators in canonical hedgehog signaling in MB. In addition, we also described AS of various regulators in noncanonical hedgehog signaling. Meanwhile, activated hedgehog signaling also induces AS in MB. Then, we pointed out that aberrant AS of hedgehog signaling is associated with different MB subgroups. Finally, we summarized the therapeutic applications of targeted AS in cancer treatment. In summary, further understanding of AS in SHH-MB could develop therapeutic targets for splicing factors which may be a novel therapeutic strategy.

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.

Control of mitochondria-associated endoplasmic reticulum membranes by protein S-palmitoylation: Novel therapeutic targets for neurodegenerative diseases.

Mitochondria-associated endoplasmic reticulum membranes (MAMs) are dynamic coupling structures between mitochondria and the endoplasmic reticulum (ER). As a new subcellular structure, MAMs combine the two critical organelle functions. Mitochondria and the ER could regulate each other via MAMs. MAMs are involved in calcium (Ca2+) homeostasis, autophagy, ER stress, lipid metabolism, etc. Researchers have found that MAMs are closely related to metabolic syndrome and neurodegenerative diseases (NDs). The formation of MAMs and their functions depend on specific proteins. Numerous protein enrichments, such as the IP3R-Grp75-VDAC complex, constitute MAMs. The changes in these proteins govern the interaction between mitochondria and the ER; they also affect the biological functions of MAMs. S-palmitoylation is a reversible protein post-translational modification (PTM) that mainly occurs on protein cysteine residues. More and more studies have shown that the S-palmitoylation of proteins is closely related to their membrane localization. Here, we first briefly describe the composition and function of MAMs, reviewing the component and biological roles of MAMs mediated by S-palmitoylation, elaborating on S-palmitoylated proteins in Ca2+ flux, lipid rafts, and so on. We try to provide new insight into the molecular basis of MAMs-related diseases, mainly NDs. Finally, we propose potential drug compounds targeting S-palmitoylation.

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.

HSP90ß chaperoning SMURF1-mediated LATS proteasomal degradation in the regulation of bone formation.

Heat shock protein 90 (HSP90) molecular chaperone is responsible for the stabilization and biological activity of a diverse set of client proteins. We have previously demonstrated that inhibition of HSP90 by 17-Demethoxy-17-allyaminogeldanmycin (17-AAG) not only reverses the glucocorticoid-induced bone loss but also enhances the basal level of bone mass in mice. Here, we investigate the potential mechanism underlying HSP90-associated osteoblast differentiation and bone formation. Knockdown of HSP90ß but not HSP90α or inhibition of HSP90 by 17-AAG or NVP-BEP800 negates the protein levels of large tumor suppressor (LATS), the core kinases of Hippo signaling, resulting in the inactivation of LATS and activation of Yes-associated protein (YAP) and transcriptional coactivator with PDZ-binding motif (TAZ), in the enhancement of osteoblastic differentiation. In contrast, genetic ablation of Lats1 in mesenchymal stem cells is sufficient to abolish the HSP90 inhibition-induced osteoblastic differentiation and bone formation. Mechanistically, HSP90ß but not HSP90α chaperones and prevents the SMAD specific E3 ubiquitin protein ligase 1 (SMURF1)-mediated and ubiquitination-dependent LATS protein proteasomal degradation, whereas 17-AAG abolishes these effects of HSP90ß. Thus, these results uncover the HSP90ß chaperoning SMURF1-mediated LATS protein proteasomal degradation and the subsequent YAP/TAZ activation as a hitherto uncharacterized mechanism controlling osteoblastic differentiation and bone formation.

Suppression of VEGFD expression by S-nitrosylation promotes the development of lung adenocarcinoma.

BACKGROUND: Vascular endothelial growth factor D (VEGFD), a member of the VEGF family, is implicated in angiogenesis and lymphangiogenesis, and is deemed to be expressed at a low level in cancers. S-nitrosylation, a NO (nitric oxide)-mediated post-translational modification has a critical role in angiogenesis. Here, we attempt to dissect the role and underlying mechanism of S-nitrosylation-mediated VEGFD suppression in lung adenocarcinoma (LUAD). METHODS: Messenger RNA and protein expression of VEGFD in LUAD were analyzed by TCGA and CPTAC database, respectively, and Assistant for Clinical Bioinformatics was performed for complex analysis. Mouse models with urethane (Ure)-induced LUAD or LUAD xenograft were established to investigate the role of S-nitrosylation in VEGFD expression and of VEGFD mutants in the oncogenesis of LUAD. Molecular, cellular, and biochemical approaches were applied to explore the underlying mechanism of S-nitrosylation-mediated VEGFD suppression. Tube formation and wound healing assays were used to examine the role of VEGFD on the angiogenesis and migration of LUAD cells, and the molecular modeling was applied to predict the protein stability of VEGFD mutant. RESULTS: VEGFD mRNA and protein levels were decreased to a different extent in multiple primary malignancies, especially in LUAD. Low VEGFD protein expression was closely related to the oncogenesis of LUAD and resultant from excessive NO-induced VEGFD S-nitrosylation at Cys277. Moreover, inhibition of S-nitrosoglutathione reductase consistently decreased the VEGFD denitrosylation at Cys277 and consequently promoted angiogenesis of LUAD. Finally, the VEGFDC277S mutant decreased the secretion of mature VEGFD by attenuating the PC7-dependent proteolysis and VEGFDC277S mutant thus reversed the effect of VEGFD on angiogenesis of LUAD. CONCLUSION: Low-expression of VEGFD positively correlates with LUAD development. Aberrant S-nitrosylation of VEGFD negates itself to induce the tumorigenesis of LUAD, whereas normal S-nitrosylation of VEGFD is indispensable for its secretion and repression of angiogenesis of LUAD.

Hedgehog Signaling: Linking Embryonic Lung Development and Asthmatic Airway Remodeling.

The development of the embryonic lung demands complex endodermal-mesodermal interactions, which are regulated by a variety of signaling proteins. Hedgehog (Hh) signaling is vital for lung development. It plays a key regulatory role during several morphogenic mechanisms, such as cell growth, differentiation, migration, and persistence of cells. On the other hand, abnormal expression or loss of regulation of Hh signaling leads to airway asthmatic remodeling, which is characterized by cellular matrix modification in the respiratory system, goblet cell hyperplasia, deposition of collagen, epithelial cell apoptosis, proliferation, and activation of fibroblasts. Hh also targets some of the pathogens and seems to have a significant function in tissue repairment and immune-related disorders. Similarly, aberrant Hh signaling expression is critically associated with the etiology of a variety of other airway lung diseases, mainly, bronchial or tissue fibrosis, lung cancer, and pulmonary arterial hypertension, suggesting that controlled regulation of Hh signaling is crucial to retain healthy lung functioning. Moreover, shreds of evidence imply that the Hh signaling pathway links to lung organogenesis and asthmatic airway remodeling. Here, we compiled all up-to-date investigations linked with the role of Hh signaling in the development of lungs as well as the attribution of Hh signaling in impairment of lung expansion, airway remodeling, and immune response. In addition, we included all current investigational and therapeutic approaches to treat airway asthmatic remodeling and immune system pathway diseases.

Revamping of Chronic Respiratory Diseases in Low- and Middle-Income Countries.

Low- and middle-income countries (LMICs) endure an asymmetrically high burden of worldwide disease and death caused by chronic respiratory diseases (CRDs), i.e., asthma, emphysema, bronchiectasis, and post-tuberculosis lung disease (PTLD). CRDs are firmly related with indigence, infectious diseases, and other non-communicable diseases (NCDs) and add to complex multi-disease with great impact on the lives and livelihood of those affected. The pertinence of CRDs to health and demographic wellbeing is relied upon to increment in the long time ahead, as expectations of life rise and the contending dangers of right on time youth mortality and irresistible infections level. The WHO has distinguished the counteraction and control of NCDs as an earnest improvement issue and crucial for the sustainable development goals (SDSs) by 2030. In this review, we center on CRDs in LMICs. We examine the early life roots of CRDs, challenges in their avoidance, identification and administration in LMICs, and the pathways to resolve for accomplish valid widespread wellbeing inclusion.

Acute Respiratory Distress Syndrome and COVID-19: A Literature Review.

Acute respiratory distress syndrome (ARDS) is an overwhelming inflammatory disorder of the lung due to direct and indirect insults to the lungs. ARDS is characterized by increased vascular permeability, protein-rich edema, diffuse alveolar infiltrate, and loss of aerated lung tissue, leading to decreased lung compliance, tachypnea, and severe hypoxemia. COVID-19 is generally associated with ARDS, and it has gained prime importance since it started. The mortality rate is alarmingly high in COVID-19-related ARDS patients regardless of advances in mechanical ventilation. Several pharmacological agents, including corticosteroids, nitric oxide, neuromuscular blocker, anti-TNF, statins, and exogenous surfactant, have been studied and some are under investigation, like ketoconazole, lisofylline, N-acetylcysteine, prostaglandins, prostacyclin, and fish oil. The purpose of this review is to appraise the understanding of the pathophysiology of ARDS, biomarkers, and clinical trials of pharmacological therapies of ARDS and COVID-19-related ARDS.

Epidemic Spread and Its Management Through Governance and Leadership Response Influencing the Arising Challenges Around COVID-19 in Pakistan-A Lesson Learnt for Low Income Countries With Limited Resource.

The coronavirus disease (COVID-19) was first reported in China (Wuhan) at the end of 2019. It has rapidly spread over 216 countries, including the USA, UK, Europe, Russia, and many Asian countries. It has affected more than 4.5 million people, and around 0.3 million deaths have been reported globally. Many preventive measures have been adopted worldwide to mitigate its spread. The government of Pakistan has also taken many preventive measures to combat the COVID-19 outbreak, such as rapid response by governance, continuous monitoring of the pandemic spread in the affected areas, and integration of resources from multiple sectors, including health, education, defense, and media. According to global statistics, the number of COVID-19 cases in the country remained remarkably lower than the expected number for the first 169 days, as compared to other countries. A total of 286,674 confirmed cases, including 16,475 active, 6,139 deaths, and 264,060 (92%) recoveries were reported. The study finds that strict adherence to national policies, effective governance, and unity at the national level resulted in better outcomes. Hence, the preventive measures, rapid responses, and strategies adopted for combating the challenges could be adopted as a learning tool for other countries having similar work environments and financial constraints. This paper can help and guide governance/public actions in response to the possible rebound of coronavirus this fall/winter.