Chromosomal Proteins, Non-Histone ; chemistry ; genetics ; metabolism ; HeLa Cells ; Histones ; chemistry ; genetics ; metabolism ; Humans ; Protein Domains

Humans ; Carcinoma, Squamous Cell/pathology* ; Chromosomal Proteins, Non-Histone/genetics* ; Ear, Middle/pathology* ; Poly-ADP-Ribose Binding Proteins ; Nuclear Proteins

OBJECTIVE: To explore the clinical and genetic features of a child with autosomal dominant mental retardation type 40 (MRD40) due to variant of the CHAMP1 gene. METHODS: Clinical characteristics of the child were analyzed. Genetic testing was carried out by low-depth high-throughput and whole genome copy number variant sequencing (CNV-seq) and whole exome sequencing (WES). A literature review was also carried out for the clinical phenotype and genetic characteristics of patients with MRD40 due to CHAMP1 gene variants. RESULTS: The child, a 11-month-old girl, has presented with intellectual and motor developmental delay. CNV-seq revealed no definite pathogenic variants. WES has detected the presence of a heterozygous c.1908C>G (p.Y636*) variant in the CHAMP1 gene, which was carried by neither parent and predicted to be pathogenic. Literature review has identified 33 additional children from 12 previous reports. All children had presented with developmental delay and mental retardation, and most had dystonia (94.1%), delayed speech and/or walking (85.2%, 82.4%) and ocular abnormalities (79.4%). In total 26 variants of the CHAMP1 gene were detected, with all nonsense variants being of loss-of-function type, located in exon 3, and de novo in origin. CONCLUSION The heterozygous c.1908C>G (p.Y636*) variant of the CHAMP1 gene probably underlay the WRD40 in this child. Genetic testing should be considered for children featuring global developmental delay, mental retardation, hypertonia and facial dysmorphism.
Humans ; Intellectual Disability/genetics* ; Genetic Testing ; Phenotype ; Exome Sequencing ; Heterozygote ; Mutation ; Chromosomal Proteins, Non-Histone/genetics* ; Phosphoproteins/genetics*

Animals ; Chromosomal Proteins, Non-Histone ; chemistry ; genetics ; metabolism ; Humans ; Models, Molecular ; Mutation ; Transcription Factors ; chemistry ; genetics ; metabolism

Mammalian cells are frequently at risk of DNA damage from both endogenous and exogenous sources. Accordingly, cells have evolved the DNA damage response (DDR) pathways to monitor and assure the integrity of their genome. In cells, the intact and effective DDR is essential for the maintenance of genomic stability and it acts as a critical barrier to suppress the development of cancer in humans. Two central kinases for the DDR pathway are ATM and ATR, which can phosphorylate and activate many downstream proteins for cell cycle arrest, DNA repair, or apoptosis if the damages are irreparable. In the last several years, we and others have made significant progress to this field by identifying BRIT1 (also known as MCPH1) as a novel key regulator in the DDR pathway. BRIT1 protein contains 3 breast cancer carboxyl terminal (BRCT) domains which are conserved in BRCA1, MDC1, 53BP1, and other important molecules involved in DNA damage signaling, DNA repair, and tumor suppression. Our in vitro studies revealed BRIT1 to be a chromatin-binding protein required for recruitment of many important DDR proteins (ATM, MDC1, NBS1, RAD51, BRCA2) to the DNA damage sites. We recently also generated the BRIT1 knockout mice and demonstrated its essential roles in homologous recombination DNA repair and in maintaining genomic stability in vivo. In humans, BRIT1 is located on chromosome 8p23.1, where loss of hetero-zigosity is very common in many types of cancer. In this review, we will summarize the novel roles of BRIT1 in DDR, describe the relationship of BRIT1 deficiency with cancer development, and also discuss the use of synthetic lethality approach to target cancers with HR defects due to BRIT1 deficiency.
Animals ; Chromosomal Proteins, Non-Histone/genetics/metabolism/*physiology ; DNA Damage/genetics/*physiology ; DNA Repair/genetics/*physiology ; Humans ; Mice ; Models, Biological ; Neoplasms/*genetics ; Nerve Tissue Proteins/genetics/metabolism/*physiology

Mammalian cells are frequently at risk of DNA damage from both endogenous and exogenous sources. Accordingly, cells have evolved the DNA damage response (DDR) pathways to monitor and assure the integrity of their genome. In cells, the intact and effective DDR is essential for the maintenance of genomic stability and it acts as a critical barrier to suppress the development of cancer in humans. Two central kinases for the DDR pathway are ATM and ATR, which can phosphorylate and activate many downstream proteins for cell cycle arrest, DNA repair, or apoptosis if the damages are irreparable. In the last several years, we and others have made significant progress to this field by identifying BRIT1 (also known as MCPH1) as a novel key regulator in the DDR pathway. BRIT1 protein contains 3 breast cancer carboxyl terminal (BRCT) domains which are conserved in BRCA1, MDC1, 53BP1, and other important molecules involved in DNA damage signaling, DNA repair, and tumor suppression. Our in vitro studies revealed BRIT1 to be a chromatin-binding protein required for recruitment of many important DDR proteins (ATM, MDC1, NBS1, RAD51, BRCA2) to the DNA damage sites. We recently also generated the BRIT1 knockout mice and demonstrated its essential roles in homologous recombination DNA repair and in maintaining genomic stability in vivo. In humans, BRIT1 is located on chromosome 8p23.1, where loss of hetero-zigosity is very common in many types of cancer. In this review, we will summarize the novel roles of BRIT1 in DDR, describe the relationship of BRIT1 deficiency with cancer development, and also discuss the use of synthetic lethality approach to target cancers with HR defects due to BRIT1 deficiency.
Animals ; Chromosomal Proteins, Non-Histone/genetics/metabolism/*physiology ; DNA Damage/genetics/*physiology ; DNA Repair/genetics/*physiology ; Humans ; Mice ; Models, Biological ; Neoplasms/*genetics ; Nerve Tissue Proteins/genetics/metabolism/*physiology

Objective: To investigate the clinicopathological, immunohistochemical and molecular genetic characteristics of gastric carcinoma with NTRK-rearrangement/amplification. Methods: The clinicopathological data of gastric carcinoma cases with NTRK-rearrangement/amplification diagnosed from January 2011 to September 2020 at the Nanjing Drum Tower Hospital, the Affiliated Hospital of Nanjing University Medical School, China, were collected. The clinicopathological, immunophenotypic and molecular pathological features were analyzed. The relevant literature was reviewed. Results: There were 4 cases of gastric carcinoma with NTRK-rearrangement/amplification. All 4 patients were male, aged 57-67 years (average, 63 years). Tumor sizes ranged from 3.5 to 5.2 cm (average, 4.8 cm). All tumors were in the antrum. All 4 patients underwent radical gastrectomy and were followed up after the surgery. Morphologically, all tumors showed histological features with enteroblastic-differentiated gastric carcinoma. Tumor cells showed predominantly tubular/papillary architecture, with conspicuous vesicular nuclei and pale staining or transparent cytoplasm. Immunohistochemistry showed pan-TRK expression in all cases, with various degrees of positivity in the cytoplasm. All cases were subject to NTRK1/2/3 detection using fluorescence in situ hybridization. There were NTRK translocations in 2 cases and NTRK amplifications in 2 cases. These cases were further verified by RNAseq next generation sequencing which confirmed that NTRK1 gene translocation (TPM3-NTRK1) and NTRK2 gene translocation (NTRK2-SMCHD1) occurred in two cases, respectively. Conclusions: NTRK mutation occurs less frequently in gastric cancer. In this study, the cases mainly occur in the antrum. The morphology has the characteristics of enteroblastic differentiation. The tumors have unique histological, immunophenotypic and molecular characteristics, which require much attention from pathologists to effectively guide clinicians to choose the best treatment.
Humans ; Male ; Female ; Receptor, trkA/genetics* ; Stomach Neoplasms/surgery* ; In Situ Hybridization, Fluorescence ; Biomarkers, Tumor/genetics* ; Translocation, Genetic ; Carcinoma ; Oncogene Proteins, Fusion/genetics* ; Chromosomal Proteins, Non-Histone/genetics*

OBJECTIVETo explore the role of centromere protein H (CENP-H) in the proliferation of human gastric cancer cells.

METHODSRT-PCR and Western blot analysis were employed to examine the mRNA and protein expressions of CENP-H in 7 human gastric cancer cell lines and immortalized human gastric epithelial cells (GES-1). The cells were infected with the retrovirus vectors pMSCV-CENP-H or CENP-H-RNAi to establish stable cell lines with high CENP-H expression or CENP-H expression interference. MTT assay and colony formation assay were used to examine the changes in the cell proliferation after the infection.

RESULTSCENP-H was over-expressed in gastric cancer cell lines AGS, BGC823, SGC-7901, MKN45, HGC27, MGC-803 and MKN28 at both mRNA and protein levels. The established AGS/CENP-H cell line with increased CENP-H expression showed enhanced proliferative activity, while the cell line MGC-803/CENP-H-RNAi with CENP-H expression interference showed an obviously lowered proliferation ability.

CONCLUSIONCENP-H promotes the proliferation of human gastric cancer cells, suggesting its important role in the occurrence and development of gastric cancer.

Cell Line, Tumor ; Cell Proliferation ; Chromosomal Proteins, Non-Histone ; genetics ; metabolism ; Humans ; RNA, Messenger ; genetics ; metabolism ; Real-Time Polymerase Chain Reaction ; Stomach Neoplasms ; metabolism ; pathology

OBJECTIVE: To construct a eukaryotic expression vector of bromodomain-containing protein 7 (BRD7) with deletion of bromodomain (BRD7△brd) using the homologous recombination and reverse PCR amplification techniques. METHODS: The linear DNA fragments of bromodomain-deleted mutation of BRD7 (pIRES2-EGFP- 3Flag/BRD7△brd) were amplified by one pair of reverse PCR primers using high-fidelity enzyme, and then these fragments were transformed into E.coli to obtain the eukaryotic expression vector expressing BRD7△brd protein based on homologous recombination and plasmid cyclization. RESULTS: Bromodomain-deleted clones were identified by digestion with restrictive enzymes, and then the sequence and protein expression were further confirmed by sequencing and Western blot assays. The results suggest that pIRES2-EGFP-3Flag/BRD7△brd was successfully constructed. CONCLUSION We establish a simple and quick method to construct plasmids with pIRES2-EGFP- 3Flag/BRD7△brd using reverse PCR amplification and homologous recombination techniques. We also found that the concentration of template in PCR reaction system is one of the critical factors that affect the rate of homologous recombination. Of all, this improved technique could be widely used in the construction of gene mutations.
Chromosomal Proteins, Non-Histone ; genetics ; Escherichia coli ; genetics ; Homologous Recombination ; Humans ; Mutation ; Plasmids ; Polymerase Chain Reaction ; methods ; Sequence Deletion ; Transfection ; methods

Animals ; Autoantigens ; genetics ; metabolism ; physiology ; Centromere Protein A ; Chromosomal Instability ; Chromosomal Proteins, Non-Histone ; genetics ; metabolism ; physiology ; Gene Expression Regulation, Neoplastic ; Humans ; Kinetochores ; metabolism ; Mitosis ; physiology ; Rectal Neoplasms ; genetics ; metabolism ; pathology ; Spindle Apparatus ; metabolism