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Cisplatin-based chemotherapy is the current standard care for lung cancer patients; however, drug resistance frequently develops during treatment, thereby limiting therapeutic efficacy.The molecular mechanisms underlying cisplatin resistance remain elusive. In this study, we conducted an analysis of microarray data from the Gene Expression Omnibus (GEO) database under the accession numbers GSE21656, which encompassed expression profiling of cisplatin-resistant H460 (DDP-H460)and the parental cells (H460). Subsequently, we calculated the differentially expressed genes (DEGs) between DDP-H460 and H460. Gene Ontology (GO) enrichment analysis and Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway analysis of DEGs demonstrated significant impact on the Rap1, PI3K/AKT and MAPK signaling pathways. Moreover, protein and protein interaction (PPI) network analysis identified PRKCA, DET1, and UBE2N as hub genes that potentially contribute predominantly to cisplatin resistance. Ultimately, PRKCA was selected for validation due to its significant prognostic effect, which predicts unfavorable overall survival and disease-free survival in patients with lung cancer. Network analysis conducted on The Cancer Genome Atlas (TCGA) database revealed a strong gene-level correlation between PRKCA and TP53, CDKN2A, BYR2, TTN, KRAS, and PIK3CA; whereas at the protein level, it exhibited a high correlation with EGFR, Lck, Bcl2, and Syk. The in vitro experiments revealed that PRKCA was upregulated in the cisplatin-resistant A549 cells (DDP-A549), while knockdown of PRKCA increased DDP-A549 apoptosis upon cisplatin treatment. Moreover, we observed that PRKCA knockdown attenuated DDP-A549 proliferation, migration and invasion ability. Western blot analysis demonstrated that PRKCA knockdown downregulated phosphorylation of PI3K expression while upregulated the genes involved in ferroptosis signaling. In summary, our results elucidate the role of PRKCA in acquiring resistance to cisplatin and underscore its potential as a therapeutic target for cisplatin-resistant lung cancer.

Abstract Background Polycystic ovary syndrome (PCOS) is a common reproductive endocrine disorder in women of reproductive age that still lacks effective treatment. Inflammation is one of the important features of PCOS. Asparagus (ASP) has anti-inflammatory, antioxidant, and anti-aging pharmacological effects, and its anti-tumor effects have been demonstrated in a variety of tumors. However, the role and mechanism of ASP in PCOS remain unclear. Methods The active components of ASP and the key therapeutic targets for PCOS were obtained by network pharmacology. Molecular docking was used to simulate the binding of PRKCA to the active components of ASP. The effects of ASP on inflammatory and oxidative stress pathways in PCOS, and the regulation of PRKCA were examined by KGN, a human derived granulosa cell line. PCOS mouse model validated the results of in vivo experiments. Results Network pharmacology identified 9 major active ingredients of ASP with 73 therapeutic targets for PCOS. Kyoto Encyclopedia of Genes and Genomes (KEGG) enrichment yielded 101 PCOS-related signaling pathways. The hub gene PRKCA was obtained after taking the gene intersection of the top 4 pathways. Molecular docking showed the binding of PRKCA to the 7 active components in ASP. In vitro and in vivo experiments showed that ASP alleviated the course of PCOS through antioxidant, anti-inflammatory effects. ASP can partially restore the low expression of PRKCA in the PCOS models. Conclusion The therapeutic effect of ASP on PCOS is mainly achieved by targeting PRKCA through the 7 active components of ASP. Mechanistically, ASP alleviated the course of PCOS through antioxidant, anti-inflammatory effects, and PRKCA was its potential target.

Abstract Background Intervertebral disk degeneration (IDD) is a degenerative disease that underlies various musculoskeletal and spinal disorders and is positively correlated with age. tRNA-derived small RNAs (tsRNA), as a new small noncoding RNAs, its function in IDD is unclear. Herein, our goal was to find the key tsRNA that affects IDD independently of age and explore the underlying mechanisms. Methods Small RNA sequencing was performed in nucleus pulposus (NP) tissues of traumatic lumbar fracture individuals, young IDD (IDDY) patients, and old IDD (IDDO) patients. The biological functions of tsRNA-04002 in NP cells (NPCs) were investigated by qRT-PCR, western blot, and flow cytometry analysis. The molecular mechanism of tsRNA-04002 was demonstrated by luciferase assays and rescue experiments. Furthermore, the therapeutic effects of tsRNA-04002 on IDD rat model were used and evaluated in vivo. Results Compared with fresh traumatic lumbar fracture patients, a total of 695 disordered tsRNAs is obtained (398 down-regulated tsRNAs and 297 up-regulated tsRNAs). These disordered tsRNAs were mainly involved in Wnt signaling pathway and MAPK signaling pathway. tsRNA-04002 was an age-independent key target in IDD, which was both lower expressed in IDDY and IDDO groups than control group. Overexpression of tsRNA-04002 restrained inflammatory cytokines IL-1β and TNF-α expression, increased the COL2A1, and inhibited the NPCs apoptosis. Furthermore, we determined that PRKCA was the target gene of tsRNA-04002 and was negatively regulated by tsRNA-04002. The rescue experiment results suggested that the high expression of PRKCA reversed the inhibitory effect of tsRNA-04002 mimics on NPCs inflammation and apoptosis, and promotive effect of COL2A1. Moreover, tsRNA-04002 treatment dramatically ameliorated the IDD process in the puncture-induced rat model, together with the blockade of PRKCA in vivo. Conclusion Collectively, our results substantiated that tsRNA-04002 could alleviate IDD by targeting PRKCA to inhibit apoptosis of NPCs. tsRNA-04002 may be a novel therapeutic target of IDD progression.

Background: Polycystic ovary syndrome (PCOS) is a common reproductive endocrine disorder in women of reproductive age that still lacks effective treatment. Inflammation is one of the important features of PCOS. Asparagus (ASP) has anti-inflammatory, antioxidant, and anti-aging pharmacological effects, and its anti-tumor effects have been demonstrated in a variety of tumors. However, the role and mechanism of ASP in PCOS remain unclear. Methods: The active components of ASP and the key therapeutic targets for PCOS were obtained by network pharmacology. Molecular docking was used to simulate the binding of PRKCA to the active components of ASP. The effects of ASP on inflammatory and oxidative stress pathways in PCOS, and the regulation of PRKCA were examined by KGN, a human derived granulosa cell line. PCOS mouse model validated the results of in vivo experiments. Results: Network pharmacology identified 9 major active ingredients of ASP with 73 therapeutic targets for PCOS. Kyoto Encyclopedia of Genes and Genomes (KEGG) enrichment yielded 101 PCOS-related signaling pathways. The hub gene PRKCA was obtained after taking the gene intersection of the top 4 pathways. Molecular docking showed the binding of PRKCA to the 7 active components in ASP. In vitro and in vivo experiments showed that ASP alleviated the course of PCOS through antioxidant, anti-inflammatory effects. ASP can partially restore the low expression of PRKCA in the PCOS models. Conclusion: The therapeutic effect of ASP on PCOS is mainly achieved by targeting PRKCA through the 7 active components of ASP. Mechanistically, ASP alleviated the course of PCOS through antioxidant, anti-inflammatory effects, and PRKCA was its potential target. [ABSTRACT FROM AUTHOR]

Background: Intervertebral disk degeneration (IDD) is a degenerative disease that underlies various musculoskeletal and spinal disorders and is positively correlated with age. tRNA-derived small RNAs (tsRNA), as a new small noncoding RNAs, its function in IDD is unclear. Herein, our goal was to find the key tsRNA that affects IDD independently of age and explore the underlying mechanisms. Methods: Small RNA sequencing was performed in nucleus pulposus (NP) tissues of traumatic lumbar fracture individuals, young IDD (IDDY) patients, and old IDD (IDDO) patients. The biological functions of tsRNA-04002 in NP cells (NPCs) were investigated by qRT-PCR, western blot, and flow cytometry analysis. The molecular mechanism of tsRNA-04002 was demonstrated by luciferase assays and rescue experiments. Furthermore, the therapeutic effects of tsRNA-04002 on IDD rat model were used and evaluated in vivo. Results: Compared with fresh traumatic lumbar fracture patients, a total of 695 disordered tsRNAs is obtained (398 down-regulated tsRNAs and 297 up-regulated tsRNAs). These disordered tsRNAs were mainly involved in Wnt signaling pathway and MAPK signaling pathway. tsRNA-04002 was an age-independent key target in IDD, which was both lower expressed in IDDY and IDDO groups than control group. Overexpression of tsRNA-04002 restrained inflammatory cytokines IL-1β and TNF-α expression, increased the COL2A1, and inhibited the NPCs apoptosis. Furthermore, we determined that PRKCA was the target gene of tsRNA-04002 and was negatively regulated by tsRNA-04002. The rescue experiment results suggested that the high expression of PRKCA reversed the inhibitory effect of tsRNA-04002 mimics on NPCs inflammation and apoptosis, and promotive effect of COL2A1. Moreover, tsRNA-04002 treatment dramatically ameliorated the IDD process in the puncture-induced rat model, together with the blockade of PRKCA in vivo. Conclusion: Collectively, our results substantiated that tsRNA-04002 could alleviate IDD by targeting PRKCA to inhibit apoptosis of NPCs. tsRNA-04002 may be a novel therapeutic target of IDD progression. [ABSTRACT FROM AUTHOR]

Ganoderma lucidum (GL) is known as a potent alleviator against chronic inflammatory disease like atherosclerosis (AS), but its mechanisms against AS have not been unveiled. This research aimed to identify the key compounds(s) and mechanism(s) of GL against AS through network pharmacology. The compounds from GL were identified by gas chromatography–mass spectrum (GC‐MS), and SwissADME screened their physicochemical properties. Then, the target(s) associated with the screened compound(s) or AS related targets were identified by public databases, and we selected the overlapping targets using a Venn diagram. The networks between overlapping targets and compounds were visualized, constructed, and analyzed by RStudio. Finally, we performed a molecular docking test (MDT) to explore key target(s), compound(s), on AutoDockVina. A total of 35 compounds in GL were detected via GC‐MS, and 34 compounds (accepted by Lipinski's rule) were selected as drug‐like compounds (DLCs). A total of 34 compounds were connected to the number of 785 targets, and DisGeNET and Online Mendelian Inheritance in Man (OMIM) identified 2,606 AS‐related targets. The final 98 overlapping targets were extracted between the compounds‐targets and AS‐related targets. On Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway enrichment, the number of 27 signaling pathways were sorted out, and a hub signaling pathway (MAPK signaling pathway), a core gene (PRKCA), and a key compound (Benzamide, 4‐acetyl‐N‐[2,6‐dimethylphenyl]) were selected among the 27 signaling pathways via MDT. Overall, we found that the identified 3 DLCs from GL have potent anti‐inflammatory efficacy, improving AS by inactivating the MAPK signaling pathway. Practical applications: Ganoderma lucidum (GL) has been used as a medicinal or edible mushroom for chronic inflammatory patients: diabetes mellitus and dyslipidemia, especially atherosclerosis (AS). Until now, the majority of mushroom research has been implemented regarding β‐glucan derivatives with very hydrophilic physicochemical properties. It implies that β‐glucan or its derivatives have poor bioavailability. Hence, we have involved GC‐MS in identifying lipophilic compounds from GL, which filtered them in silico to sort drug‐like compounds (DLCs). Then, we retrieved targets associated with the DLCs, and identified a key signaling pathway, key targets, and key compounds against AS. In this paper, we utilized bioinformatics and network pharmacology theory to understand the uncovered pharmacological mechanism of GL on AS. To sum things up, our analysis elucidates the relationships between signaling pathways, targets, and compounds in GL. Ultimately, this work provides biochemical evidence to identify the therapeutic effect of GL on AS, and a scientific basis for deciphering the key mechanism on DLCs of GL against AS. [ABSTRACT FROM AUTHOR]

Gene therapy aims to silence an oncogene through RNA interference, or replace an abnormal tumour suppressor via gene augmentation. In this study, we intended RNA interference for PRKCA oncogene and gene augmentation for PTEN tumour suppressor with a view to reduce tumour growth in a mouse model of breast cancer. Inorganic carbonate apatite nanoparticles (CA NPs) were utilized to deliver the synthetic siRNA and the purified gene-carrying plasmid DNA both in vitro and in vivo. Effects of PRKCA siRNA- and PTEN plasmid-loaded NPs on viability of MCF-7, MDA-MB-231 and 4T1 breast cancer cells were assessed by MTT assay. The cell viability data in MCF-7 cell line demonstrated that combined delivery of PRKCA specific siRNA and PTEN plasmid with CA NPs had an additive effect to significantly decrease cellular growth compared to individual treatments. In addition, we observed a similar pattern of cumulative influence for combined treatment in triple negative MDA-MB-231 breast cancer cell line. Upon treatment with PRKCA siRNA+PTEN plasmid-loaded NPs, a remarkable decrease in the phosphorylated form of AKT protein of PI3K/AKT pathway was observed in Western blot, indicative of diminished proliferative signal. Moreover, in vivo study in MCF-7 xenograft breast cancer mouse model demonstrated that the rate of growth and final tumour volume were reduced significantly in the mouse group that received intravenous treatment of PRKCA siRNA+NPs, and PTEN plasmid+NPs. Our findings demonstrated that PRKCA siRNA and PTEN plasmid loaded into CA NPs attenuated breast tumour growth, suggesting their therapeutic potential in the treatment of breast cancer. [ABSTRACT FROM AUTHOR]

Cisplatin-based chemotherapy is the current standard care for lung cancer patients; however, drug resistance frequently develops during treatment, thereby limiting therapeutic efficacy.The molecular mechanisms underlying cisplatin resistance remain elusive. In this study, we conducted an analysis of microarray data from the Gene Expression Omnibus (GEO) database under the accession numbers GSE21656, which encompassed expression profiling of cisplatin-resistant H460 (DDP-H460)and the parental cells (H460). Subsequently, we calculated the differentially expressed genes (DEGs) between DDP-H460 and H460. Gene Ontology (GO) enrichment analysis and Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway analysis of DEGs demonstrated significant impact on the Rap1, PI3K/AKT and MAPK signaling pathways. Moreover, protein and protein interaction (PPI) network analysis identified PRKCA, DET1, and UBE2N as hub genes that potentially contribute predominantly to cisplatin resistance. Ultimately, PRKCA was selected for validation due to its significant prognostic effect, which predicts unfavorable overall survival and disease-free survival in patients with lung cancer. Network analysis conducted on The Cancer Genome Atlas (TCGA) database revealed a strong gene-level correlation between PRKCA and TP53, CDKN2A, BYR2, TTN, KRAS, and PIK3CA; whereas at the protein level, it exhibited a high correlation with EGFR, Lck, Bcl2, and Syk. The in vitro experiments revealed that PRKCA was upregulated in the cisplatin-resistant A549 cells (DDP-A549), while knockdown of PRKCA increased DDP-A549 apoptosis upon cisplatin treatment. Moreover, we observed that PRKCA knockdown attenuated DDP-A549 proliferation, migration and invasion ability. Western blot analysis demonstrated that PRKCA knockdown downregulated phosphorylation of PI3K expression while upregulated the genes involved in ferroptosis signaling. In summary, our results elucidate the role of PRKCA in acquiring resistance to cisplatin and underscore its potential as a therapeutic target for cisplatin-resistant lung cancer., Competing Interests: The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (© 2024 The Authors.)

Diabetes increases in the worldwide by prevalence and numbers of complications related to it. The most common risk factor for foot ulcers in diabetic is peripheral neuropathy. In diabetic condition, Ca2+ concentration intracellular increases. NRF2 (Nuclear Factor Erythroid 2) acts as a bridging link in various inflammatory and apoptotic pathways impacting progress of diabetic neuropathy. The aim of this study is to predict the pathway of NRF2 activation that is mediated by the increased of intracellular Ca2+. Data interaction between Ca2+ and NRF2 were retrieved from STITCH (Search Tool for Interacting Chemicals) which were experimentally and prediction, then were analyzed computationally. Pathway analysis used Cytoscape software. The functional analysis was evaluated using STRING (Search Tool for the Retrieval of Interacting Genes/Proteins) database. The consequence of increased intracellular Ca2+ content is the increases of oxidative stress, ROS (Reactive Oxygen Species) and the translocation of the protein kinase Cα (PKRCA) to the plasma membrane as the initial step of their activations. This study reveals that activation NRF2 by Ca2+ intracellular which increases in diabetic condition through PRKCA, PRKCA phosphorylate NRF2 in its Neh2 domain at Ser-40.

Objective: To explore the relationship between SNPs in PRKCA‐HIF1A‐GLUT1 and diabetic kidney disease in Chinese Han people. Materials and methods: A total of 2552 participants from Shanghai Diabetes Institute Inpatient Database of Shanghai Jiao Tong University Affiliated Sixth People's Hospital were involved in the stage 1 cross‐sectional population. A total of 6015 subjects from the Hong Kong Diabetes Register were included for validation. Genotyping of participants was conducted by the MassARRAY Compact Analyzer (Agena Bioscience). The data were analysed by plink, SAS, Haploview. Results: We identified variants associated with diabetic kidney disease in stage 1. Rs1681851 (P =.0105, OR = 1.331) in GLUT1 as well as rs2301108 (P =.0085, OR = 1.289) and rs79865957 (P =.0204, OR = 1.263) in HIF1A were significantly associated with diabetic kidney disease. Regarding DKD‐related traits, rs1681851 was associated with plasma creatinine levels (P =.0169, β = 4.822) and eGFR (P =.0457, β = −6.956). Moreover, the results showed the interactions between PRKCA‐GLUT1 in the occurrence of DKD. We further sought validation of the 17 SNPs in a prospective cohort and found that rs900836 and rs844501 were associated with the percentage change in eGFR slope. We performed a meta‐analysis of case‐control analysis data from the Hong Kong samples together with the stage 1 data from Shanghai. Rs9894851 showed significant correlation with the serum creatinine level as well as eGFR and no SNP showed association with DKD after meta‐analysis. Conclusions: Our results suggest potential association between SNPs in PRKCA‐HIF1A‐GLUT1 and diabetic kidney disease in Chinese Han people. [ABSTRACT FROM AUTHOR]

Dry mature fruits of Hippophae rhamnoides L. (HRL), Elaeagnaceae , have traditional functions of invigorating spleen and improving spleen insufficiency. Traditional Chinese medicine (TCM) clinics have been proved that HRL is in favor of diabetes treatment. Modern pharmacological studies demonstrated that total flavones of Hippophae rhamnoides (TFH) are the main substance for HRL to develop anti-inflammation and anti-diabetes functions. However, chemical features, active ingredients and anti-diabetes pharmacological mechanism of HRL still remain unclear. Key targets and metabolites in anti-type-II diabetes mellitus (T2DM) of TFH have been explored based on AGE-RAGE signaling pathway in diabetic complications. The anti-T2DM mechanism of TFH has been elaborated from comprehensive perspectives, including target prediction, metabolites, potential metabolic pathways, and so on. In this study, a qualitative test of chemical composition of HRL was carried out based on ultra-high performance liquid chromatography-quadrupole time-of-flight mass spectrometry (UPLC-Q-TOF-MS). The anti-T2DM targets and pathways of HRL were predicted through network pharmacological approach. The T2DM rat model was induced by high-fat and high-glucose diet combined with streptozotocin (STZ). The T2DM model was evaluated through fasting blood glucose level, body weight, serum biochemical indicators, insulin levels and homeostatic model assessment of insulin resistance. The key metabolic pathways were screened through the correlation between metabolites and key targets. Finally, the quantitative analysis of key targets and metabolites was verified through experiments. After TFH intervention, the fasting blood-glucose level of T2DM rats induced by high-fat and high-glucose diet combined with streptozotocin (STZ) was downregulated significantly, while body weight, serum liquid level, insulin levels and homeostatic model assessment of insulin resistance (HOMA-IR) were improved. According to ELISA, Western blotting (WB) and reverse transcriptase polymerase chain reaction (RT-PCR), TFH significantly downregulates expression levels of diglyceride (DAG)-activated protein kinase C (PRKCA), mitogen activated protein kinase 10 (MAPK10), human nuclear factor κB subunit p65 (NF-κB p65) and tumor necrosis factor-α (TNF-α) in pancreas of STZ-induced rats. TFH downregulates expressions of PRKCA, MAPK10 and p65 TNF-α as well as level of the key metabolite DA in the DAG/PRKCA/MAPK10/TNF-α/p65 pathways, improves lipid metabolism disorder, inhibits inflammatory response and thereby relieves symptoms of T2DM. [Display omitted] • TFH administration is beneficial for high-fat and high-glucose diet combined with STZ induced T2DM. • By downregulating level of DAG, TFH can improve relieve symptoms of T2DM. • By downregulating expression levels of PRKCA, MAPK10, p65, TNF-α, relieve symptoms of T2DM. [ABSTRACT FROM AUTHOR]

Gene therapy aims to silence an oncogene through RNA interference, or replace an abnormal tumour suppressor via gene augmentation. In this study, we intended RNA interference for PRKCA oncogene and gene augmentation for PTEN tumour suppressor with a view to reduce tumour growth in a mouse model of breast cancer. Inorganic carbonate apatite nanoparticles (CA NPs) were utilized to deliver the synthetic siRNA and the purified gene-carrying plasmid DNA both in vitro and in vivo. Effects of PRKCA siRNA- and PTEN plasmid-loaded NPs on viability of MCF-7, MDA-MB-231 and 4T1 breast cancer cells were assessed by MTT assay. The cell viability data in MCF-7 cell line demonstrated that combined delivery of PRKCA specific siRNA and PTEN plasmid with CA NPs had an additive effect to significantly decrease cellular growth compared to individual treatments. In addition, we observed a similar pattern of cumulative influence for combined treatment in triple negative MDA-MB-231 breast cancer cell line. Upon treatment with PRKCA siRNA+PTEN plasmid-loaded NPs, a remarkable decrease in the phosphorylated form of AKT protein of PI3K/AKT pathway was observed in Western blot, indicative of diminished proliferative signal. Moreover, in vivo study in MCF-7 xenograft breast cancer mouse model demonstrated that the rate of growth and final tumour volume were reduced significantly in the mouse group that received intravenous treatment of PRKCA siRNA+NPs, and PTEN plasmid+NPs. Our findings demonstrated that PRKCA siRNA and PTEN plasmid loaded into CA NPs attenuated breast tumour growth, suggesting their therapeutic potential in the treatment of breast cancer.

MicroRNAs (miRNAs) are closely related with the posttranscriptional regulation of gene expression. Our past study showed that let‐7g‐5p decreased during pregnancy and lactation in mouse mammary gland, what suggested the prospective regulating role of let‐7g‐5p in mammary epithelial cells. In this study, to unravel the role of let‐7g‐5p, we found PRKCA (PKC‐alpha, PKC‐α) might serve as potential targets of let‐7g‐5p by bioinformatics. Then let‐7g‐5p was knocked down by an antisense oligonucleotide in mouse primary mammary epithelial cells. As predicted, PRKCA gene expression and mammary epithelial cells growth were increased by anti‐let‐7g‐5p regulation in vitro. By using reporter constructs, it showed that the let‐7g‐5p could direct binding with 3′‐the untranslated regions (3′‐UTR) of PRKCA mRNA. Furthermore, ectopic overexpression of let‐7g‐5p in vivo reduced PRKCA and β‐casein protein expression as well as inhibited mammary gland growth. These results suggested that let‐7g‐5p could inhibit the mammary epithelial cells differentiation and β‐casein protein synthesis and expression through suppression of PRKCA on the cycle of mammary cell differentiation and development and that let‐7g‐5p may be a novel important regulated target in mammary cells function. The let‐7g‐5p could inhibit the mammary epithelial cells differentiation and β‐casein protein synthesis and expression through suppression of PRKCA on the cycle of mammary cell differentiation and development, and that let‐7g‐5p may be a novel important regulated target in mammary cells function [ABSTRACT FROM AUTHOR]

Papillary glioneuronal tumor (PGNT) is a WHO-defined brain tumor entity that poses a major diagnostic challenge. Recently, SLC44A1–PRKCA fusions have been described in PGNT. We subjected 28 brain tumors from different institutions histologically diagnosed as PGNT to molecular and morphological analysis. Array-based methylation analysis revealed that 17/28 tumors exhibited methylation profiles typical for other tumor entities, mostly dysembryoplastic neuroepithelial tumor and hemispheric pilocytic astrocytoma. Conversely, 11/28 tumors exhibited a unique profile, thus constituting a distinct methylation class PGNT. By screening the extended Heidelberg cohort containing over 25,000 CNS tumors, we identified three additional tumors belonging to this methylation cluster but originally histologically diagnosed otherwise. RNA sequencing for the detection of SLC44A1–PRKCA fusions could be performed on 19 of the tumors, 10 of them belonging to the methylation class PGNT. In two additional cases, SLC44A1–PRKCA fusions were confirmed by FISH. We detected fusions involving PRKCA in all cases of this methylation class with material available for analyses: the canonical SLC44A1–PRKCA fusion was observed in 11/12 tumors, while the remaining case exhibited a NOTCH1-PRKCA fusion. Neither of the fusions was found in the tumors belonging to other methylation classes. Our results point towards a high misclassification rate of the morphological diagnosis PGNT and clearly demonstrate the necessity of molecular analyses. PRKCA fusions are highly diagnostic for PGNT, and detection by RNA sequencing enables the identification of rare fusion partners. Methylation analysis recognizes a unique methylation class PGNT irrespective of the nature of the PRKCA fusion. [ABSTRACT FROM AUTHOR]

Fusion genes involving homologs of protein kinase C (PKC) have been identified in a variety of tumors. We report the clinical and histologic presentation of 51 cutaneous melanocytic neoplasms with a PKC fusion gene (involving PRKCA in 35 cases, PRKCB in 15 cases, and PRKCG in a single case). Most tumors were in young adults (median age, 29.5 years; range, 1-73 years) but some presented in newborns. Histologically, 42 tumors were classified as benign, presenting predominantly as biphasic dermal proliferation (88%) with nests of small melanocytes surrounded by fibrosis with haphazardly arranged spindled and dendritic melanocytes, resembling those reported as "combined blue nevi." Most tumors (60%) were heavily pigmented and in 15%, hyperpigmented epithelioid melanocytes were present at the dermoepidermal junction. Two lesions were paucicellular and showed marked sclerosis. Three tumors, including 2 proliferating nodules, were considered intermediate grade. Six tumors had sheets of atypical melanocytes infiltrating the dermis and were classified as melanomas. Two of the melanomas displayed loss of BAP1 nuclear expression. The median follow-up time was 12 months, with 1 patient alive with metastatic disease and 1 dying of their melanoma. These results suggest that melanocytic tumors with PKC fusion genes have characteristic histopathologic features, which are more similar to blue nevi than to pigmented epithelioid melanocytomas. As is the case with GNA-mutated blue nevi, they can progress to melanomas via BAP1 inactivation and metastasize., (Copyright © 2023 United States & Canadian Academy of Pathology. Published by Elsevier Inc. All rights reserved.)

Diabetes increases in the worldwide by prevalence and numbers of complications related to it. The most common risk factor for foot ulcers in diabetic is peripheral neuropathy. In diabetic condition, Ca2+ concentration intracellular increases. NRF2 (Nuclear Factor Erythroid 2) acts as a bridging link in various inflammatory and apoptotic pathways impacting progress of diabetic neuropathy. The aim of this study is to predict the pathway of NRF2 activation that is mediated by the increased of intracellular Ca2+. Data interaction between Ca2+ and NRF2 were retrieved from STITCH (Search Tool for Interacting Chemicals) which were experimentally and prediction, then were analyzed computationally. Pathway analysis used Cytoscape software. The functional analysis was evaluated using STRING (Search Tool for the Retrieval of Interacting Genes/Proteins) database. The consequence of increased intracellular Ca2+ content is the increases of oxidative stress, ROS (Reactive Oxygen Species) and the translocation of the protein kinase Cα (PKRCA) to the plasma membrane as the initial step of their activations. This study reveals that activation NRF2 by Ca2+ intracellular which increases in diabetic condition through PRKCA, PRKCA phosphorylate NRF2 in its Neh2 domain at Ser-40. [ABSTRACT FROM AUTHOR]

Background The protein kinase C alpha ( PRKCA ) gene, coding for a Th17-cell-selective kinase, shows a complex splicing pattern, with at least 2 stable alternative transcripts characterized by an alternative upstream polyadenylation site. Polymorphisms in this gene were associated with several conditions, including multiple sclerosis, asthma, schizophrenia, and cancer. The presence of a microRNA (miRNA), i.e. miR-634, within intron 15 of the PRKCA gene, suggests the intriguing possibility that this miRNA might play a role in the susceptibility to these pathologies. Methods Here, we characterized miR-634 expression profile and searched for its putative targets using a combination of RT-PCR and gene reporter assays. Results The quantitative analysis of PRKCA and miR-634 transcripts in a panel of human tissues and cell lines revealed discordant expression profiles, suggesting the presence of an independent miR-634 promoter and/or a possible direct role of miR-634 in modulating PRKCA expression. Functional studies demonstrated the existence of a miRNA-specific promoter, which was shown to be Pol-III-dependent. Furthermore, transfection experiments showed that miR-634 is able to target its host gene by specifically down-regulating the shorter alternative-polyadenylated isoforms. Conclusions MiR-634 is a Pol III-dependent intronic miRNA, which could target its host gene through a “first-order” negative feedback. General significance MiR-634 is one of the few characterized examples of Pol-III-dependent intronic miRNAs. Its independent transcription from the host gene suggests caution in using expression profiles of host genes as proxies for the expression of the corresponding intronic miRNAs. [ABSTRACT FROM AUTHOR]

Although chromosome 17 shows high genetic instability, interstitial deletions within 17q24 region are very rare. The phenotype of affected patients is heterogeneous but generally includes microcephaly, dysmorphic features, heart defects, intellectual disability, Andersen-Tawil syndrome and Carney complex. We present a 10-month-old proband with rare 5.4 Mb microdeletion of 17q24.1-q24.3 region. Her main clinical manifestations include microcephaly, distinctive dysmorphic features, and hypotrophy. Among 30 deleted protein coding genes, we hypothesize that PRKCA, PSMD12, KPNA2, PRKAR1A, and KCNJ2 might be responsible for the proband's phenotype. Changes in PRKCA gene have been described in patients with abnormal cognitive function and haploinsufficiency of PSMD12 has been associated with neurodevelopmental disorders and dysmorphia. Deletions of KPNA2 gene, known to be involved in Nijmegen breakage syndrome, are characterized by microcephaly, short stature, and syndactyly. PRKAR1A gene is known to be involved in Carney complex. Haploinsufficiency of KCNJ2 gene in this region causes episodes of muscle weakness and arrhythmia. So far, these symptoms have not been observed in the proband. Nevertheless, as these and additional comorbidities might develop over time, continuous monitoring is required. [ABSTRACT FROM AUTHOR]

Acute lung injury (ALI) caused by sepsis is a common respiratory critical illness with high morbidity and mortality. Protein kinase C-alpha (PRKCA) plays a protective role in sepsis-induced ALI. However, the detailed molecular mechanism of PRKCA in ALI caused by sepsis is unclear. Animal and cell models of sepsis were established by cecal ligation and puncture (CLP)-surgery and lipopolysaccharide (LPS)/interferon-gamma (IFN-γ) treatment, respectively. Lentivirus transfection was used to overexpress PRKCA. H&E staining and lung injury in CLP-surgery mice were evaluated. Gene expression was evaluated using qPCR and Western blotting. The expression of TNF-α, IL-1β, and IL-6 was examined using qPCR and ELISA. The expression of LC3 and TOM20 was evaluated using immunofluorescence assays. Cell apoptosis was assessed using a flow cytometry assay. The bond between miR-15a-5p and PDK4 was confirmed by dual-luciferase reporter gene and RNA immunoprecipitation assays. In vivo and in vitro, PRKCA overexpression reduced lung injury to prompt mitophagy and inhibit the inflammatory response, ROS production, and cell apoptosis. miR-15a-5p was highly expressed in macrophages treated with LPS/IFN-γ and was negatively mediated by PRKCA. The overexpression of miR-15a-5p reduced the effects of PRKCA upregulation in macrophages. miR-15a-5p could restrain mitophagy in LPS/IFN-γ-treated macrophages by directly targeting PDK4. Furthermore, PDK4 knockdown reversed the inhibition of cell apoptosis and inflammatory factor release caused by miR-15a-5p silencing. The PRKCA/miR-15a-5p/PDK4 axis alleviated ALI caused by sepsis by promoting mitophagy and repressing anti-inflammatory response.

A full term female newborn presented with prominent forehead, bilateral microphthalmia, iris coloboma and cataract, wide intercanthal distance, large, low-set and protruding ears, skin tag at the left nasal nostril, imperforate anus with rectovestibular fistula, and postnatal growth delay with brachymicrocephaly. A marker chromosome was not detectable and the copy number of 22q11 was normal. However, array CGH revealed a 3.5 Mb microdeletion of chromosome region 3q26.32-3q26.33 (chr. 3: 178,598,162-182,114,483; hg19) which comprised the SOX2 gene. While SOX2 haploinsufficiency is known to cause microphthalmia and coloboma, it has not been described before in patients with anal atresia. © 2013 Wiley Periodicals, Inc. [ABSTRACT FROM AUTHOR]

Mixed neuronal-glial tumors are rare and challenging to subclassify. One recently recognized variant, papillary glioneuronal tumor ( PGNT), is characterized by prominent pseudopapillary structures and glioneuronal elements. We identified a novel translocation, t(9;17)(q31;q24), as the sole karyotypic anomaly in two PGNTs. A fluorescence in situ hybridization ( FISH)-based positional cloning strategy revealed SLC44A1, a member of the choline transporter-like protein family, and PRKCA, a protein kinase C family member of serine/threonine-specific protein kinases, as the 9q31 and 17q24 breakpoint candidate genes, respectively. Reverse transcription-polymerase chain reaction ( RT-PCR) analysis using a forward primer from SLC44A1 exon 5 and a reverse primer from PRKCA exon 10 confirmed the presence of a SLC44A1-PRKCA fusion product in both tumors. Sequencing of each chimeric transcript uncovered an identical fusion cDNA junction occurring between SLC44A1 exon 15 and PRKCA exon 9. A dual-color breakpoint-spanning probe set custom-designed for interphase cell recognition of the translocation event identified the fusion in a third PGNT. These results suggest that the t(9;17)(q31;q24) with the resultant novel fusion oncogene SLC44A1-PRKCA is the defining molecular feature of PGNT that may be responsible for its pathogenesis. The FISH and RT-PCR assays developed in this study can serve as valuable diagnostic adjuncts for this rare disease entity. [ABSTRACT FROM AUTHOR]

We earlier reported a genome-wide significant linkage to schizophrenia at chromosome 17 that was identified in a single pedigree (C702) consisting of six affected, male siblings with DSM-IV schizophrenia and prominent mood symptoms. In this study, we adopted several approaches in an attempt to map the putative disease locus. First, mapping the source of linkage to chromosome 17 in pedigree C702. We refined the linkage region in family C702 to a 21-marker segment spanning 11.7 Mb at 17q23-q24 by genotyping a total of 50 microsatellites across chromosome 17 in the pedigree. Analysis of data from 1028 single nucleotide polymorphisms (SNPs) across the refined linkage region identified a single region of homozygosity present in pedigree C702 but not in 2938 UK controls. This spanned ∼432 kb of the gene encoding protein kinase C, alpha (PRKCA), the encoded protein of which has been implicated in the pathogenesis of psychiatric disorders. Analysis of pedigree C702 by oligonucleotide-array comparative genome hybridization excluded the possibility that this region of homozygosity was because of a deletion. Mutation screening of PRKCA identified a rare, four-marker haplotype (C-HAP) in the 3′ untranslated region of the gene, which was present in the homozygous state in all six affected members of pedigree C702. No other homozygotes were observed in genotype data for a total of 6597 unrelated Europeans (case N=1755, control N=3580 and parents of probands N=1262). Second, association analysis of C702 alleles at PRKCA. The low-frequency haplotype (C-HAP) showed a trend for association in a study of unrelated schizophrenia cases and controls from the UK (661 cases, 2824 controls, P=0.078 and odd ratio (OR)=1.9) and significant evidence for association when the sample was expanded to include cases with bipolar (N=710) and schizoaffective disorder (N=50) (psychosis sample: 1421 cases, 2824 controls, P=0.037 and OR=1.9). Given that all the affected members of C702 are male, we also undertook sex-specific analyses. This revealed that the association was strongest in males for both schizophrenia (446 male cases, 1421 male controls, P=0.008 and OR=3.9) and in the broader psychosis group (730 male cases, 1421 male controls, P=0.008 and OR=3.6). Analysis of C-HAP in follow-up samples from Ireland and Bulgaria revealed no evidence for association in either the whole sample or in males alone, and meta-analysis of all male psychosis samples yielded no significant evidence of association (969 male cases, 1939 male controls, 311 male probands P=0.304 and OR=1.4). Third, association mapping of the pedigree C702 linkage region. Independent of pedigree C702, genotype data from the Affymetrix 500k GeneChip set were available for 476 patients with schizophrenia and 2938 controls from the United Kingdom. SNPs in PRKCA showed evidence for association with schizophrenia that achieved gene-wide significance (P=0.027). Moreover, the same SNP was the most significantly associated marker out of the 1028 SNPs genotyped across the linkage region (rs873417, allelic P=0.0004). Follow-up genotyping in samples from Ireland, Bulgaria and Germany did not show consistent replication, but meta-analysis of all samples (4116 cases and 6491 controls) remained nominally significant (meta-analysis P=0.026, OR=1.1). We conclude that, although we have obtained convergent lines of evidence implicating both rare and common schizophrenia risk variants at PRKCA, none of these is individually compelling. However, the evidence across all approaches suggests that further study of this locus is warranted. [ABSTRACT FROM AUTHOR]

Abstract: The vitamin D receptor (VDR) mediates the actions of 1,25-dihydroxyvitamin D3 (1,25(OH)2D3) in target cells and tissues by orchestrating the expression of gene networks responsible for vitamin D-induced phenotypes. The molecular mechanisms of these regulatory systems have been studied for decades under the principle that transcriptional regulation occurs near the transcriptional start site of the gene. However, this now appears to be an outdated view of transcriptional control. In this study, we examined the genome-wide chromatin immunoprecipitation on microarray (ChIP-chip) across pre-osteoblastic cells for VDR, retinoid X receptor (RXR), RNA polymerase II, and histone H4 acetylation (H4ac). We uncovered potential regulatory mechanisms for genes important to osteoblast biology as well as skeletal formation under the control of 1,25(OH)2D3. We found that VDR, along with RXR and H4ac, binds to distal regions 43% of the time; and within gene introns and exons 44%, leaving only 13% of activation at traditional promoter regions. Here, we briefly summarize our findings for all the VDR/RXR cis-acting transcriptional elements (VDR/RXR cistrome) in pre-osteoblastic cells, MC3T3-E1, provide a few examples of this dynamic control by VDR and 1,25(OH)2D3, and demonstrate that distal transcriptional control contributes to the majority of vitamin D3-mediated transcription. [ABSTRACT FROM AUTHOR]

Cryptosporidium infection is a leading cause of diarrhea-associated morbidity and mortality in young children globally. Single nucleotide polymorphisms (SNPs) in the human protein kinase C-α ( PRKCA ) gene region have been associated with susceptibility to cryptosporidiosis. Here, we examined the role of protein kinase C-α (PKCα) activity in human HCT-8 intestinal epithelial cells during infection with Cryptosporidium parvum sporozoites. To delineate the role of PKCα in infection, we developed a fluorescence-based imaging assay to differentiate adherent from intracellular parasites. We tested pharmacological agonists and antagonists of PKCα and measured the effect on C. parvum sporozoite adherence to and invasion of HCT-8 cells. We demonstrate that both PKCα agonists and antagonists significantly alter parasite adherence and invasion in vitro . We found that HCT-8 cell PKCα is activated by C. parvum infection. Our findings suggest intestinal epithelial cell PKCα as a potential host-directed therapeutic target for cryptosporidiosis and implicate PKCα activity as a mediator of parasite adherence and invasion.

Simple Summary: Over the last decades, the incidence of tongue cancer has risen among young patients who often show an aggressive course of disease. At the same time, exposure to the major risk factors, alcohol and tobacco, has decreased, indicating that a novel risk factor may be involved. In this study, we found that high expression of protein kinase C alpha (PRKCA) is frequent among young patients without alcohol and smoking history and associated with a poor prognosis. Our results suggest that PRKCA levels may serve as a molecular marker of an emerging high-risk subgroup of young tongue cancer patients. Elucidation of the underlying mechanisms may clarify whether PRKCA expression itself promotes disease progression and which genetic or environmental factors trigger its upregulation. Oral tongue squamous cell carcinomas (OTSCCs) have an increasing incidence in young patients, and many have an aggressive course of disease. The objective of this study was to identify candidate prognostic protein markers associated with early-onset OTSCC. We performed an exploratory screening for differential protein expression in younger (≤45 years) versus older (>45 years) OTSCC patients in The Cancer Genome Atlas (TCGA) cohort (n = 97). Expression of candidate markers was then validated in an independent Austrian OTSCC patient group (n = 34) by immunohistochemistry. Kaplan–Meier survival estimates were computed, and genomic and mRNA enrichment in silico analyses were performed. Overexpression of protein kinase C alpha (PRKCA) was significantly more frequent among young patients of both the TCGA (p = 0.0001) and the Austrian cohort (p = 0.02), associated with a negative anamnesis for alcohol consumption (p = 0.009) and tobacco smoking (p = 0.02) and poorer overall survival (univariate p = 0.02, multivariate p< 0.01). Within the young subgroup, both overall and disease-free survival were significantly decreased in patients with PRKCA overexpression (both p < 0.001). TCGA mRNA enrichment analysis revealed 332 mRNAs with significant differential expression in PRKCA-upregulated versus PRKCA-downregulated OTSCC (all FDR ≤ 0.01). Our findings suggest that PRKCA overexpression may be a hallmark of a novel molecular subtype of early-onset alcohol- and tobacco-negative high-risk OTSCC. Further analysis of the molecular PRKCA interactome may decipher the underlying mechanisms of carcinogenesis and clinicopathological behavior of PRKCA-overexpressing OTSCC. [ABSTRACT FROM AUTHOR]

To date, efforts to improve non-small-cell lung cancer (NSCLC) outcomes with increased radiation dose have not been successful. Identification of novel druggable targets that are capable to modulate NSCLC radiosensitivity may provide a way forward. Mediator complex is implicated in gene expression control, but it remains unclear how Mediator dysfunction is involved in cancer radiotherapy. Methods: The biologic functions of miR-4497, MED13L and PRKCA in NSCLC radiosensitivity were examined through biochemical assays including gene expression profilling, cell proliferation assay, colony formation assay, wound healing assay, transwell assay, dual luciferase reporter assay, xenograft models, immunoprecipitation, and chromatin immunoprecipitation sequencing. Clinical implications of miR-4497, MED13L and PRKCA in radiosensitivity were evaluated in NSCLC patients treated with concurrent chemoradiotherapy or radiotherapy alone. Results: We found that radiation can trigger disassemble of Mediator complex via silencing of MED13L by miR-4497 in NSCLC. Although not interrupting structure integrity of the core Mediator or the CDK8 kinase module, suppression of MED13L attenuated their physical interactions and reduced recruitment of acetyltransferase P300 to chromatin via Mediator. Silencing of MED13L therefore diminishes global H3K27ac signals written by P300, activities of enhancer and/or promoters and expression of multiple oncogenes, especially PRKCA . Inhibition of PRKCA expression potentiates the killing effect of radiotherapy in vitro and in vivo . Remarkably, high PRKCA expression in NSCLC tissues is correlated with poor prognosis of patients received radiotherapy. Conclusions: Our study linking PRKCA to radiosensitivity through a novel mechanism may enable the rational targeting of PRKCA to unlock therapeutic potentials of NSCLC., Competing Interests: Competing Interests: The authors have declared that no competing interest exists., (© The author(s).)

Non-small cell lung cancer (NSCLC) is a highly malignant tumor. Many circular RNAs (circRNAs) are reportedly in regulating the progression of NSCLC. To identify potential therapeutic targets for NSCLC, we conducted a bioinformatics analysis of circRNAs differentially expressed between NSCLC tissues and adjacent normal tissues. Hsa&#95;circ&#95;0007580 was upregulated in NSCLC tumor tissues, and the expression of its host gene (protein kinase Ca) correlated negatively with overall survival. Short-hairpin RNAs were used to knock down hsa&#95;circ&#95;0007580 in NSCLC cells, and gene and protein levels were measured with qRT-PCR and Western blotting, respectively. NSCLC cell proliferation, migration and apoptosis were evaluated with CCK-8 assays, Ki-67 staining, Transwell assays and flow cytometry, respectively. Knocking down hsa&#95;circ&#95;0007580 inhibited proliferation and invasion by NSCLC cells and induced their apoptosis. Dual luciferase reporter assays indicated that miR-545-3p can bind to hsa&#95;circ&#95;0007580 (suggesting that hsa&#95;circ&#95;0007580 sponges miR-545-3p) and to protein kinase Ca (suggesting that miR-545-3p directly inhibits this gene). In a xenograft tumor model, downregulating hsa&#95;circ&#95;0007580 inhibited NSCLC tumorigenesis by inactivating p38/mitogen-activated protein kinase signaling. Thus, silencing hsa&#95;circ&#95;0007580 notably inhibited NSCLC progression in vitro and in vivo , suggesting this circRNA could be a novel treatment target for NSCLC.

Traumatic injury is a leading cause of mortality and morbidity. MicroRNAs (miRNAs) regulate the cellular responses when traumatic injury occurs. Previously, we reported that miR-3945, miR-125a-5p, miR-363-3p, and miR-150-5p were significantly altered in neutrophils of patients who suffered traumatic injury. In the present study, by comparing neutrophils of patients suffering from major trauma with neutrophils of patients with a inflammatory disease, we found that the variation trend of miR-150-5p was relatively different in the process of these two diseases. Gene Ontology and pathway analysis of miR-150-5p revealed that it may activate the mitogen-activated protein kinase and Toll-like receptor signaling pathways and cell adhesion molecules when the traumatic injury occurs. In addition, protein kinase C alpha (PRKCA) was also identified as a direct target of miR-150-5p by establishing a miRNA-mRNA network, and this target was validated via dual-luciferase reporter and western blot analysis. Our results suggested that the expression of miR-150-5p was down-regulated in neutrophils after a major traumatic injury. miR-150-5p and its identified target PRKCA play important roles in the development of traumatic process., (© The Author 2015. Published by ABBS Editorial Office in association with Oxford University Press on behalf of the Institute of Biochemistry and Cell Biology, Shanghai Institutes for Biological Sciences, Chinese Academy of Sciences.)