Wolf-Hirschhorn Syndrome Candidate Research Articles

Wolf-Hirschhorn syndrome candidate 1 (Whsc1) methyltransferase signals via a Pitx2-miR-23/24 axis to effect tooth development

Wolf-Hirschhorn syndrome (WHS) is a developmental disorder attributed to a partial deletion on the short arm of chromosome 4. WHS patients suffer from oral manifestations including cleft lip and palate, hypodontia, and taurodontism. WHS candidate 1 (WHSC1) gene is a H3K36-specific methyltransferase that is deleted in every reported case of WHS. Mutation in this gene also results in tooth anomalies in patients. However, the correlation between genetic abnormalities and the tooth anomalies has remained controversial. In our study, we aimed to clarify the role of WHSC1 in tooth development. We profiled the Whsc1 expression pattern during mouse incisor and molar development by immunofluorescence staining and found Whsc1 expression is reduced as tooth development proceeds. Using real-time quantitative reverse transcription PCR, Western blot, chromatin immunoprecipitation, and luciferase assays, we determined that Whsc1 and Pitx2, the initial transcription factor involved in tooth development, positively and reciprocally regulate each other through their gene promoters. miRNAs are known to regulate gene expression posttranscriptionally during development. We previously reported miR-23a/b and miR-24-1/2 were highly expressed in the mature tooth germ. Interestingly, we demonstrate here that these two miRs directly target Whsc1 and repress its expression. Additionally, this miR cluster is also negatively regulated by Pitx2. We show the expression of these two miRs and Whsc1 are inversely correlated during mouse mandibular development. Taken together, our results provide new insights into the potential role of Whsc1 in regulating tooth development and a possible molecular mechanism underlying the dental defects in WHS.

WHSC1L1-mediated epigenetic downregulation of VMP1 participates in herpes simplex virus 1 infection-induced mitophagy impairment and neuroinflammation

Microglia are the first-line defenders against invading pathogens in the brain whose activation mediates virus clearance and leads to neurotoxicity as well. This work studies the role of Wolf-Hirschhorn syndrome candidate 1-like 1 (WHSC1L1)/vacuole membrane protein 1 (VMP1) interaction in the activation of microglia and neuroinflammation following herpes simplex virus 1 (HSV-1) infection. Aberrantly expressed genes after HSV-1 infection were screened by analyzing the GSE35943 dataset. C57BL/6J mice and mouse microglia BV2 were infected with HSV-1 for in vivo and in vitro assays. VMP1 was downregulated but WHSC1L1 was upregulated in HSV-1-infected mouse brain tissues as well as in BV2 cells. The VMP1 overexpression enhanced mitophagy activity and suppressed oxidative stress and inflammatory activation of BV2 cells, but these effects were blocked by the autophagy antagonist 3-methyladenine. WHSC1H1 suppressed VMP1 transcription through H3K36me2-recruited DNMT3A. Downregulation of WHSC1H1 similarly enhanced mitophagy in BV2 cells, and it alleviated microglia activation, nerve cell inflammation, and brain tissue damage in HSV-1-infected mice. However, the alleviating roles of WHSC1H1 silencing were negated by further VMP1 silencing. Taken together. this study demonstrates that WHSC1L1 upregulation following HSV-1 infection leads to mitophagy impairment and neuroinflammation through epigenetic suppression of VMP1.

WHSC1 is involved in DNA damage, cellular senescence and immune response in hepatocellular carcinoma progression

Wolf-Hirschhorn syndrome candidate 1 (WHSC1) is a transcriptional regulatory protein that encodes a histone methyltransferase to control H3K36me2 modification. WHSC1 was upregulated and associated with poor prognosis in HCC. The elevated WHSC1 likely due to the alterations of DNA methylation or RNA modification. WHSC1 perhaps form a chromatin cross talk with H3K27me3 and DNA methylation to regulate transcription factors expression in HCC. Functional analysis indicated that WHSC1 was involved in DNA damage repair, cell cycle, cellular senescence and immune regulations. Furthermore, WHSC1 was associated with the infiltrating levels of B cell, CD4+, Tregs and macrophage cells. Therefore, our findings suggested that WHSC1 might function as a promotor regulator to affect the development and progression of HCC. Thus, WHSC1 could be a potential biomarker in predicting the prognosis and therapeutic target for patients with HCC.

NSD3, a member of nuclear receptor-binding SET domain family, is a potential prognostic biomarker for pancreatic cancer.

Members of the nuclear receptor-binding SET domain (NSD) family of histone H3 lysine 36 methyltransferases comprise NSD1, NSD2 (MMSET/WHSC1), and NSD3 (Wolf-Hirschhorn syndrome candidate 1-like 1, WHSC1L1). While the expression of NSD genes is essential to normal biological processes and cancer, knowledge of their expression levels to prognosticate in cancer remains unclear. We analyzed the expression patterns for NSD family genes across multiple cancer types and examined their association with clinical features and patient survival profiles. Next, we explored the association between NSD3 expression and described features of the tumor microenvironment (TME) in PAAD, a severe type of pancreatic cancer. In particular, we correlated promoter methylation levels for NSD3 with patient outcomes in PAAD. Finally, we explored the putative oncogenic roles for NSD3 using a series of experiments with pancreatic cancer cells. We report that the expression of NSD family members is correlated with clinical prognosis across multiple types of cancers. Also, we demonstrate that NSD3 variants are most prevalent among NSD genes across cancers we analyzed. Notably, when compared with NSD1 and NSD2, we find that NSD3 is prominently expressed, and its expression is significantly linked with clinical outcome in pancreatic cancer. Furthermore, NSD3 is frequently amplified, exhibits low promoter methylation, and is correlated with immune cell infiltration and enhanced proliferation of pancreatic cancer. Finally, we demonstrate that knockdown of NSD3 alters H3K36me2 methylation, downstream gene expression and EGFR/ERK signaling in pancreatic cancer cells. We find that expression levels, the presence of genetic variants of NSD family genes, as well as their promoter methylation are correlated with clinical outcomes in cancer, including pancreatic cancer. Our in vitro experiments suggest that NSD3 may be relevant to gene expression regulation and growth factor signaling in pancreatic cancer.

Tumor-augmenting Effect of Histone Methyltransferase WHSC1 on Colorectal Cancer Via Epigenetic Upregulation of TACC3 and PI3K/Akt Activation

Wolf-Hirschhorn syndrome candidate 1 (WHSC1) is a histone methyltransferase which is frequently expressed in an aberrant manner in tumors, but its specific roles and targets in colorectal cancer (CRC) remain unclear. Expression of WHSC1 in CRC tissues and cells was examined by RT-qPCR, immunohistochemistry and immunoblot assays. WHSC1 was knocked down in CRC cells to examine its effect on cell proliferation, invasiveness, migration, epithelial-mesenchymal transition (EMT) activity and apoptosis. The binding between WHSC1 and transforming acidic coiled-coil containing protein 3 (TACC3) was predicted using bioinformatics systems and subsequently validated. In addition, altered expression of TACC3 was introduced into CRC cells to perform functional assays. Stably transfected cells were injected into nude mice to induce xenograft tumors. WHSC1 was highly expressed in CRC tumor tissues and cells with a positive correlation with TACC3 concentration. Specific downregulation of WHSC1 in two CRC cell lines blocked cell proliferation, invasiveness, EMT and migration activity and promoted cell apoptosis. WHSC1 triggers H3K36me2 modification at the level of the TACC3 promoter to induce TACC3 activation. TACC3 suppression similarly blocked the malignant activity of CRC cells, whereas TACC3 restoration in cells counteracted the cancer-suppressive functions of WHSC1 silencing. TACC3 levels were correlated with increased phosphorylation of the PI3K/Akt signaling pathway in cells. Likewise, WHSC1 suppression blocked tumor growth in nude mice. The results of this study revealed TACC3 as a target of WHSC1 in CRC that is positively correlated with PI3K/Akt pathway activation and tumor development.

CircWHSC1 expedites cervical cancer progression via miR-532-3p/LTBP2 axis.

Dysregulated circRNAs have potential roles in the progression of various cancer types, including cervical cancer (CaCx). The carcinogenic roles of circRNA Wolf-Hirschhorn syndrome candidate gene-1 (circWHSC1) are described in the development of diverse cancers. The objective of this study was to investigate the expression and the underlying role of circWHSC1 in CaCx. The expression of circWHSC1 was detected by real-time PCR. After the suppression of circWHSC1 expression, the changes in the proliferation, migration, invasion, and apoptosis capacities were detected by CCK-8 assay, colony formation assay, Transwell assays, flow cytometry, and the determination of apoptosis-related proteins. The interplay among circWHSC1, miR-532-3p, and latent transforming growth factor-β binding protein 2 (LTBP2) was confirmed by luciferase reporter and biotinylated RNA pull-down assays. A nude mice xenograft tumor model was established to evaluate the anti-tumorigenic role of circWHSC1 silencing in vivo. CircWHSC1 was overexpressed in CaCx tissues and cell lines and its high expression was inversely associated with the survival rate of patients with CaCx. CircWHSC1 silencing was capable of suppressing the proliferation, metastasis, and invasion of tumor cells and inducing apoptosis. Investigation to its molecular mechanism revealed that circWHSC1 functioned as a competitive endogenous RNA (ceRNA), mediating LTBP2 expression by targeting miR-532-3p. The in vivo experiments further confirmed the inhibition of tumor growth and metastasis by circWHSC1 knockdown. The circWHSC1-mediated miR-532-3p/LTBP2 signaling axis might be a novel therapeutic target for CaCx.

Glucocorticoid receptor gene mutations confer glucocorticoid resistance in B-cell precursor acute lymphoblastic leukemia

Glucocorticoid (GC) is a key drug in the treatment of B-cell precursor acute lymphoblastic leukemia (BCP-ALL), and the initial GC response is an important prognostic factor. GC receptors play an essential role in GC sensitivity, and somatic mutations of the GC receptor gene, NR3C1, are reportedly identified in some BCP-ALL cases, particularly at relapse. Moreover, associations of somatic mutations of the CREB-binding protein (CREBBP) and Wolf-Hirschhorn syndrome candidate 1 (WHSC1) genes with the GC-resistance of ALL have been suggested. However, the significance of these mutations in the GC sensitivity of BCP-ALL remains to be clarified in the intrinsic genes. In the present study, we sequenced NR3C1, WHSC1, and CREBBP genes in 99 BCP-ALL and 22 T-ALL cell lines (32 and 67 cell lines were known to be established at diagnosis and at relapse, respectively), and detected their mutations in 19 (2 cell lines at diagnosis and 15 cell lines at relapse), 26 (6 and 15), and 38 (11 and 15) cell lines, respectively. Of note, 14 BCP-ALL cell lines with the NR3C1 mutations were significantly more resistant to GC than those without mutations. In contrast, WHSC1 and CREBBP mutations were not associated with GC resistance. However, among the NR3C1 unmutated BCP-ALL cell lines, WHSC1 mutations tended to be associated with GC resistance and lower NR3C1 gene expression. Finally, we successfully established GC-resistant sublines of the GC-sensitive BCP-ALL cell line (697) by disrupting ligand binding and DNA binding domains of the NR3C1 gene using the CRISPR/Cas9 system. These observations demonstrated that somatic mutations of the NR3C1 gene, and possibly the WHSC1 gene, confer GC resistance in BCP-ALL.

Wolf-Hirschhorn syndrome candidate 1 facilitates alveolar macrophage pyroptosis in sepsis-induced acute lung injury through NEK7-mediated NLRP3 inflammasome activation.

Sepsis is a complex clinical syndrome with high incidence and mortality. Acute lung injury (ALI) is a common complication of sepsis. At present, there is no effective therapeutic strategy to treat ALI. The SET domain–containing histone methyltransferase Wolf–Hirschhorn syndrome candidate 1 (WHSC1) regulates cancer progression, while its role in sepsis-induced ALI remains unclear. Thus, this study aimed to study the effect of WHSC1 on sepsis-induced ALI and to explore the potential mechanism of action. In the study, LPS treatment induced lung injury. WHSC1 was highly expressed in LPS-induced ALI. Knockdown of WHSC1 attenuated LPS-induced ALI and pyroptosis in vivo. Besides, knockdown of WHSC1 attenuated LPS-induced alveolar macrophage pyroptosis in vitro. Furthermore, NIMA-related kinase-7 (NEK7) expression could be regulated by WHSC1, and NEK7 bound to NLRP3 in alveolar macrophages. Moreover, WHSC1 regulated alveolar macrophage pyroptosis through modulating NEK7-mediated NLRP3 inflammasome activation. In conclusion, WHSC1 was highly expressed in LPS-induced ALI. WHSC1 facilitated alveolar macrophage pyroptosis in sepsis-induced ALI through NEK7-mediated NLRP3 inflammasome activation. WHSC1 may be a valuable target for the therapy of sepsis-induced ALI.

High WHSC1L1 Expression Reduces Survival Rates in Operated Breast Cancer Patients with Decreased CD8+ T Cells: Machine Learning Approach.

Nuclear receptor-binding SET domain protein (NSD), a histone methyltransferase, is known to play an important role in cancer pathogenesis. The WHSC1L1 (Wolf-Hirschhorn syndrome candidate 1-like 1) gene, encoding NSD3, is highly expressed in breast cancer, but its role in the development of breast cancer is still unknown. The purpose of this study was to analyze the survival rates and immune responses of breast cancer patients with high WHSC1L1 expression and to validate the results using gradient boosting machine (GBM) in breast cancer. We investigated the clinicopathologic parameters, proportions of immune cells, pathway networks and in vitro drug responses according to WHSC1L1 expression in 456, 1500 and 776 breast cancer patients from the Hanyang University Guri Hospital, METABRIC and TCGA, respectively. High WHSC1L1 expression was associated with poor prognosis, decreased CD8+ T cells and high CD274 expression (encoding PD-L1). In the pathway networks, WHSC1L1 was indirectly linked to the regulation of the lymphocyte apoptotic process. The GBM model with WHSC1L1 showed improved prognostic performance compared with the model without WHSC1L1. We found that VX-11e, CZC24832, LY2109761, oxaliplatin and erlotinib were effective in inhibiting breast cancer cell lines with high WHSC1L1 expression. High WHSC1L1 expression could play potential roles in the progression of breast cancer and targeting WHSC1L1 could be a potential strategy for the treatment of breast cancer.

Characterization of Wolf‐Hirschhorn Syndrome Candidate 1 ( Whsc1 ) in Pitx2 ‐ miR‐23/24 ‐mediated Tooth Development

Objective Wolf-Hirschhorn syndrome (WHS) is a developmental disorder attributed to a partial deletion on the short arm of chromosome 4. WHS patients suffer from oral manifestations including cleft lip and palate (CLP), hypodontia and taurodontism. However, the causative factors and underlying mechanisms of these oral anomalies are relatively unknown. Wolf-Hirschhorn syndrome candidate 1 (WHSC1) is a H3K36-specific methyltransferase that is frequently deleted in WHS. This gene has been associated with craniofacial defects including CLP and defects in occipital ossification. In our study, we aim to understand the role of WHSC1 in tooth development. Methods To characterize the role of Whsc1 in tooth development, we profiled the Whsc1 expression pattern during mouse tooth development by immunofluorescence staining (IF). To investigate the regulatory effects between Whsc1 and Pitx2, we overexpressed either Whsc1 or Pitx2 in oral epithelial cell line LS8 and detected their expression. Then, ChIP-PCR and luciferase assays were performed to confirm the binding on the promoter regions. To determine the negative regulation of Whsc1 by miR-23-3p and miR-24-3p, miRs were inhibited by our Plasmid-based microRNA inhibition system in PMIS-miR-23-27-24 embryos and LS8 cells followed by detecting the mRNA and protein level of Whsc1. Luciferase assays were also conducted to confirm the direct binding between miRs and Whsc1 3’UTR. Results Whsc1 expresses from early stage of tooth development and restricts to stem cell niches in homeostatic tooth germ. Whsc1 and Pitx2 reciprocally activate each other's expression through chromosomal and transcriptional regulation. miR-23-3p and miR-24-3p, two miRs that regulated by Pitx2, directly inhibit Whsc1. Conclusion Whsc1 expresses in the developing tooth germ and participates the Whsc1-Pitx2-miR23/24 regulatory network in oral epithelium cells. Our observations provide new insights into the potential role of Whsc1 in regulating tooth development and a possible causer of the dental defects in WHS.

CircWHSC1 serves as an oncogene to promote hepatocellular carcinoma progression.

Circular RNAs (circRNAs) function as vital regulators in multifarious cancers, including hepatocellular carcinoma (HCC). However, the roles of circRNA Wolf-Hirschhorn syndrome candidate gene-1 (circWHSC1) in HCC are barely known. Quantitative real-time polymerase chain reaction (qRT-PCR) was conducted for the levels of circWHSC1, miR-142-3p, miR-421, miR-665 and homeobox A1 (HOXA1) mRNA. Cell Counting Kit-8 (CCK-8) assay, colony formation assay and 5'-ethynyl-2'-deoxyuridine (EdU) assay were used to evaluate cell proliferation ability. Transwell assay was adopted for cell migration and invasion. Western blot assay was employed for protein levels. RNA pull-down assay, dual-luciferase reporter assay and RNA immunoprecipitation (RIP) assay were executed to verify the interaction between miR-142-3p and circWHSC1 or HOXA1. Murine xenograft model assay was conducted for the role of circWHSC1 in vivo. The morphology of exosomes was observed by transmission electron microscopy (TEM). CircWHSC1 was elevated in HCC tissues and cells, and high level of circWHSC1 was associated with worse overall survival of HCC patients. Knockdown of circWHSC1 suppressed HCC cell proliferation and metastasis in vitro and restrained tumorigenesis in vivo. CircWHSC1 functioned as the sponge for miR-142-3p, which directly targeted HOXA1. Inhibition of miR-142-3p ameliorated the effects of circWHSC1 knockdown on HCC cell proliferation and metastasis. Moreover, miR-142-3p overexpression restrained the growth and motility of HCC cells, with HOXA1 elevation reversing the impacts. Additionally, circWHSC1 was increased in HCC patients' serum and might be a diagnostic indicator for HCC. CircWHSC1 played a tumour-promoting role in HCC by elevating HOXA1 through sponging miR-142-3p.

WHSC1/NSD2 regulates immune infiltration in prostate cancer

BackgroundImmunotherapy in prostate cancer (PCa) lags behind the progresses obtained in other cancer types partially because of its limited immune infiltration. Tumor-resident immune cells have been detected in the prostate,...

WHSC1 Promotes Cell Proliferation, Migration, and Invasion in Hepatocellular Carcinoma by Activating mTORC1 Signaling.

BackgroundWolf-Hirschhorn syndrome candidate gene-1 (WHSC1) plays key regulatory roles in cancer development and progression. However, its specific functions and potential mechanisms of action remain to be described in hepatocellular carcinoma (HCC).Materials and MethodsWHSC1 expression in HCC was evaluated using The Cancer Genome Atlas and verified in HCC tissues and cell lines using qRT-PCR, Western blotting, and immunohistochemistry. Functional assays were performed to explore the role of WHSC1 in HCC progression. Immunoprecipitation-mass spectrometry, co-immunoprecipitation, immunofluorescence, and immunohistochemistry were conducted to evaluate the interaction between WHSC1 and prolyl 4-hydroxylase subunit beta (P4HB). Pathway enrichment was performed using gene set enrichment analysis.ResultsWHSC1 was markedly overexpressed in HCC tissues and cell lines. The level of expression was strongly associated with adverse clinicopathological characteristics. Survival analyses revealed that WHSC1 upregulation predicted poor overall survival and higher recurrence rates in patients with HCC. Functional studies revealed that WHSC1 significantly stimulated HCC proliferation, migration, and invasion in vitro and in vivo. WHSC1 was shown to interact with P4HB to stimulate P4HB expression and subsequently activate mTOR1 signaling.ConclusionWe determined the oncogenic role of WHSC1 in HCC, via P4HB interaction, which activates mTOR1 signaling, and identified WHSC1 as a promising therapeutic target for HCC.

WHSC1 promotes wnt/β-catenin signaling in a FoxM1-dependent manner facilitating proliferation, invasion and epithelial-mesenchymal transition in breast cancer

Objectives: Wolf-Hirschhorn syndrome candidate gene-1 (WHSC1) is highly expressed in various malignant tumors. We investigated the correlation and regulatory pathway of WHSC1 in the progression of breast cancer (BC).Methods: The expression and distribution of WHSC1 in the BC tissues and cell lines were determined by quantitative reverse transcription-polymerase chain reaction (qRT-PCR) and immunohistochemical staining. Spearman correlation analysis demonstrated the correlation between WHSC1 high expression level and the clinical characteristics of BC patients. The effects of WHSC1 on the proliferation, apoptosis, migration and invasion of BC cells were analyzed by cell transfection, MTT, colony formation, scratch assay, and transwell. Furthermore, the expression of Forkhead box M1 (FoxM1) and the location of β-catenin were detected by qRT-PCR and western blot.Results: Firstly, WHSC1 expression was up-regulated in BC tissues and cell lines. The high expression of WHSC1 in BC is associated with the tumor size (p = 0.027), metastasis (p = 0.018) and pathological stages (p = 0.025) of the BC patients. The knockdown of WHSC1 inhibited the growth, proliferation migration, invasion and EMT of BC cell lines. Furthermore, WHSC1 could promote the expression of FoxM1 in BC cells and tissues. WHSC1 enhanced the expression of FoxM1, and promoted the nuclear localization of β-catenin, and thus activated the downstream genes expression of Wnt/β-catenin signaling pathway to regulate the development of BC.Conclusion: In summary, our study elucidates the correlation and specific regulatory mechanism between WHSC1 and the progression of BC, thus implying that WHSC1 may function as molecular diagnosis, prognosis and molecular targeted therapy of BC.

Acute NelfA knockdown restricts compensatory gene expression and precipitates ventricular dysfunction during cardiac hypertrophy

Coordinated functional balance of negative and positive transcription complexes maintain and accommodate gene expression in hearts during quiescent and hypertrophic conditions, respectively. Negative elongation factor (Nelf) complex has been implicated in RNA polymerase II (pol II) pausing, a widespread regulatory transcriptional phenomenon observed across the cardiac genome. Here, we examine the role of NelfA aka, Wolf-Hirschhorn syndrome candidate 2 (Whsc2), a critical component of the negative elongation complex in hearts undergoing pressure-overload induced hypertrophy. Alignment of high-resolution genome-wide occupancy data of NelfA, Pol II, TFIIB and H3k9ac from control and hypertrophied hearts reveal that NelfA associates with active gene promoters. High NelfA occupancy is seen at promoters of essential and cardiac-enriched genes, expressed under both quiescent and hypertrophic conditions. Conversely, de novo NelfA recruitment is observed at inducible gene promoters with pressure overload, accompanied by significant increase in expression of these genes with hypertrophy. Interestingly, change in promoter NelfA levels correlates with the transcript output in hypertrophied hearts compared to Sham, suggesting NelfA might be playing a critical role in the regulation of gene transcription during cardiac hypertrophy. In vivo knockdown of NelfA (siNelfA) in hearts subjected to pressure-overload results in early ventricular dilatation and dysfunction, associated with decrease in expression of inducible and cardiac-enriched genes in siNelfA hypertrophied compared to control hypertrophied hearts. In accordance, in vitro knockdown of NelfA in cardiomyocytes showed no change in promoter pol II, however significant decrease in in-gene and downstream pol II occupancy was observed. These data suggest an inhibited pol II progression in transcribing and inducible genes, which reflects as a decrease in transcript abundance of these genes. These results indicate that promoter NelfA occupancy is essential for pol II –dependent transcription. Therefore, we conclude that NelfA is required for active transcription and gene expression during cardiac hypertrophy.

A Scalable Platform for Producing Recombinant Nucleosomes with Codified Histone Methyltransferase Substrate Preferences

Wolf-Hirschhorn Syndrome Candidate 1 (WHSC1; also known as NSD2) is a SET domain-containing histone lysine methyltransferase. A chromosomal translocation occurs in 15–20% of multiple myeloma patients and is associated with increased production of WHSC1 and poor clinical prognosis. To define the substrate requirements of NSD2, we established a platform for the large-scale production of recombinant polynucleosomes, based on authentic human histone proteins, expressed in E. coli, and complexed with linearized DNA. A brief survey of methyltransferases whose substrate requirements are recorded in the literature yielded expected results, lending credence to the fitness of our approach. This platform was readily ‘codified’ with respect to both position and extent of methylation at histone 3 lysines 18 and 36 and led to the conclusion that the most readily discernible activity of NSD2 in contact with a nucleosome substrate is dimethylation of histone 3 lysine 36. We further explored reaction mechanism, and conclude a processive, rather than distributive mechanism best describes the interaction of NSD2 with intact nucleosome substrates. The methods developed feature scale and flexibility and are suited to thorough pharmaceutical-scale drug discovery campaigns.

WHSC1 acts as a prognostic indicator and functions as an oncogene in cervical cancer.

Purpose: Wolf–Hirschhorn syndrome candidate 1 (WHSC1) is an epigenetic modifier, considered to play a driving role in oncogenesis. However, very little is known about the roles of WHSC1 and its prognostic impacts in cervical cancer. This study aimed to investigate the role of WHSC1 in the prognosis of cervical cancer and explore the effect of WHSC1 on proliferation, migration, and invasion of cervical cancer cells and angiogenesis in human umbilical vein endothelial cells (HUVECs).Methods: We evaluated the expression levels of WHSC1 in cervical cancer samples and relevant cell lines by immunohistochemistry, real-time quantitative PCR, and Western blot. In vitro, Cell Counting Kit-8 and transwell assays were used to investigate the viability and migration of C33A cells, and a tube formation assay was used to study the effect of WHSC1 on angiogenesis in HUVECs.Results: WHSC1 was overexpressed in cervical cancer tissues and cells, and correlated with the FIGO stage and differentiation. WHSC1 knockdown inhibited proliferation, suppressed migration and invasion in endothelial nitric oxide synthase (eNOS)-overexpressing C33A cells, and inhibited angiogenesis in HUVECs.Conclusion: WHSC1 may be a poor prognostic indicator of cervical cancer and a potential novel therapeutic target for the same. WHSC1 may participate in the regulation of cervical cancer progression through the eNOS signaling pathway.

Wolf–Hirschhorn Syndrome Candidate 1 (whsc1) Functions as a Tumor Suppressor by Governing Cell Differentiation

Wolf–Hirschhorn syndrome candidate 1 (WHSC1) is a histone 3 lysine 36 (H3K36) specific methyltransferase that is frequently deleted in Wolf–Hirschhorn syndrome (WHS). Whsc1 is also found mutated in a subgroup of B-cell derived malignant diseases by genomic translocation or point mutation, both of which resulted in hyperactivity of WHSC1 mediated H3K36 methylation and uncontrolled cell proliferation, suggesting that whsc1 functions as an oncogene. However, here we provided evidences to show that whsc1 also has tumor suppressor functions. We used zebrafish as an in vivo model and generated homozygous whsc1 mutant lines via clustered regularly interspaced short palindromic repeats-associated protein Cas9 (CRISPR/Cas9) technology. Then western-blot (WB) and immunofluorescence (IF) were performed to analysis the expression level of H3K36Me2 and H3K36Me3, and we identified the diseased tissue via hematoxylin–eosin (HE) staining, IF staining or immunohistochemistry (IHC). Whsc1 lose-of-function led to significant decrease in di- and tri-methylation of H3K36. A series of WHS related phenotypes were found in whsc1−/− zebrafish, including growth retardation, neural development defects and heart failure. In addition, loss of function of whsc1 led to defects in the development of swim bladder, possibly through the dis-regulation of key genes in swim bladder organogenesis and inhibition of progenitor cell differentiation, which was correlated with its expression in this organ during embryonic development. At later stage, these whsc1−/− zebrafishes are inclined to grow tumors in the swim bladder. Our work suggested that whsc1 may function as a tumor suppressor by governing progenitor cell differentiation.

Wolf-Hirschhorn Syndrome Candidate 1 Is Necessary for Correct Hematopoietic and B Cell Development

Immunodeficiency is one of the most important causes of mortality associated with Wolf-Hirschhorn syndrome (WHS), a severe rare disease originated by a deletion in chromosome 4p. The WHS candidate 1 (WHSC1) gene has been proposed as one of the main genes responsible for many of thealterations in WHS, but its mechanism of actionis still unknown. Here, we present invivo genetic evidence showing that Whsc1 plays an important role at several points of hematopoietic development. Particularly, our results demonstrate that both differentiation and function of Whsc1-deficient B cells are impaired at several key developmental stages due to profound molecular defects affecting B cell lineage specification, commitment, fitness, and proliferation, demonstrating a causal role for WHSC1 in the immunodeficiency of WHS patients.

Wolf-Hirschhorn syndrome candidate 1-like 1 epigenetically regulates nephrin gene expression.

Altered expression of nephrin underlies the pathophysiology of proteinuria in both congenital and acquired nephrotic syndrome. However, the epigenetic mechanisms of nephrin gene regulation remain elusive. Here, we show that Wolf-Hirschhorn syndrome candidate 1-like 1 long form (WHSC1L1-L) is a novel epigenetic modifier of nephrin gene regulation. WHSC1L1-L was associated with histone H3K4 and H3K36 in human embryonic kidney cells. WHSC1L1-L gene was expressed in the podocytes, and functional protein product was detected in these cells. WHSC1L1-L was found to bind nephrin but not other podocyte-specific gene promoters, leading to its inhibition/suppression, abrogating the stimulatory effect of WT1 and NF-κB. Gene knockdown of WHSC1L1-L in primary cultured podocytes accelerated the transcription of nephrin but not CD2AP. An in vivo zebrafish study involving the injection of Whsc1l1 mRNA into embryos demonstrated an apparent reduction of nephrin mRNA but not podocin and CD2AP mRNA. Immunohistochemistry showed that both WHSC1L1-L and nephrin emerged at the S-shaped body stage in glomeruli. Immunofluorescence and confocal microscopy displayed WHSC1L1 to colocalize with trimethylated H3K4 in the glomerular podocytes. Chromatin immunoprecipitation assay revealed the reduction of the association of trimethylated H3K4 at the nephrin promoter regions. Finally, nephrin mRNA was upregulated in the glomerulus at the early proteinuric stage of mouse nephrosis, which was associated with the reduction of WHSC1L1. In conclusion, our results demonstrate that WHSC1L1-L acts as a histone methyltransferase in podocytes and regulates nephrin gene expression, which may in turn contribute to the integrity of the slit diaphragm of the glomerular filtration barrier.