Rearrangement of T cell receptor genes Tcra and Tcrd and its regulatory mechanism / 中华微生物学和免疫学杂志

The high diversity of T cell receptors (TCRs) is the basis for recognizing antigens, playing an essential role in adaptive immunity. TCR diversity is generated from V(D)J rearrangement during the thymocyte development in the thymus. Standing out from the four TCR genes, Tcra and Tcrd genes are characterized by locating at the same locus and sharing specific V genes. Hence, their rearrangement and regulation have a certain particularity. Previous studies mainly focused on cis-regulatory elements and trans-acting factors regulating the Tcra/ Tcrd rearrangement. However, recent progress has shown that chromatin spatial organization plays an essential role in antigen receptor gene rearrangement. Chromatin organization proteins, such as CTCF-Cohesin, are involved in regulating rearrangement and enhancing the diversity of TCR repertoire by loop extrusion. Recombinase RAG also scans chromatin of antigen receptor genes for rearrangement. This review described the progress in the rearrangement of Tcra and Tcrd genes and the possible regulatory mechanism, especially the influence of the chromatin spatial organization.

Identification and analysis of anovel variant of TRAPPC2 in a X-linked spondyloepiphyseal dysplasia tarda pedigree / 中华骨科杂志

Objective:To provide experimental evidence for genetic counseling and prenatal diagnosis by analyzing the clinical characteristics, screening and identification of the function of suspicious variants in a X-1inked spondyloepiphyseal dysplasia tarda (SEDT) family.Methods:The family members' medical history, general physical examination, femur, spine X-ray examination were collected. Peripheral blood samples of the family members were collected and DNA was extracted from these samples. Sequencing clinical whole exons of proband DNA by targeted gene high-throughput sequencing method, then analysis sequencing data. The suspicious mutation was confirmed in pedigree members by PCR and Sanger sequencing. Reverse transcription polymerase chain reaction (RT-PCR) experiments of total RNA from blood lymphocytes were performed. The amplification of exons 3 and 4 of the pathogenic gene were amplified and identified by agarose gel. The expression of the pathogenic gene was also detected.Results:Three affected males of the family were diagnosed with SEDT according to their clinical and radiological features. A nonsense mutation in the transport protein particle complex subunit 2 ( TRAPPC2) gene NM_001011658: c.91A>T (p.K31*) was found in the proband using whole exome sequencing. This variation was also detected in his cousin, but not in non-phenotypic members of the family. The RT-PCR result for amplification of exon 3 and 4 of peripheral blood lymphocytes was the same as those of normal controls, indicating that the mutation did not affect the splicing of transcripts. qPCR results showed that the transcriptional expression of TRAPPC2 in patients was significantly lower than that in family normal controls and normal people controls. Conclusion:Identification of the novel nonsense mutation (c.91A>T) in the SEDT family enables early patients screening, carrier detection, genetic counseling, prenatal diagnosis, and clinical prevention and treatment. The detailed genotype/phenotype descriptions contribute to the SEDT mutation spectrum. The study of the function of TRAPPC2 mutation will help to further elucidate the role of sedlin in cartilage.

Analysis of MYO7A gene mutation in a family with non-syndromic autosomal recessive deafness / 中华医学遗传学杂志

Objective@#To explore the genetic basis for a family with non-syndromic autosomal recessive deafness.@*Methods@#The proband and her parents were subjected to physical and audiological examinations. With genomic DNA extracted from peripheral blood samples, next-generation sequencing was carried out using a panel for deafness genes. Suspected mutation was validated by Sanger sequencing and qPCR analysis of her parents.@*Results@#The proband presented bilateral severe sensorineural hearing loss at three days after birth. Her auditory threshold was 110-120 dBnHL but with absence of vestibular and retinal symptoms. Her brother also had deafness but her parents were normal. No abnormality was found upon physical examination of her family members, while audiological examination showed no middle ear or retrocochlear diseases. Next-generation sequencing identified compound heterozygous mutations of the MYO7A gene, including a previously known c. 462C>A (p. Cys154Ter) and a novel EX43_46 Del, which were respectively derived from her mother and father.@*Conclusion@#The compound heterozygous mutations of the MYO7A gene probably underlie the disease in this family. Our findings has enriched the mutation spectrum for non-syndromic autosomal recessive deafness 2.

The improvewment of DNA library construction in non-crosslinked chromatin immunoprecipitation coupled with next-generation sequencing / 南方医科大学学报

OBJECTIVE@#To optimize DNA library construction in non-crosslinked chromatin immunoprecipitation coupled with next-generation sequencing (Native ChIP-seq) to obtain high-quality Native ChIP-seq data.@*METHODS@#Human nasopharyngeal carcinoma HONE1 cell lysate was digested with MNase for release of the nucleosomes, and the histone-DNA complexes were immunoprecipitated with specific antibodies. The protein component in the precipitate was digested with proteinase K followed by DNA purification; the DNA library was constructed for sequence analysis.@*RESULTS@#Compared with the conventional DNA library construction, Tn5 transposase method allowed direct enrichment of the target DNA after Tn5 fragmentation, which was simple, time-saving and more efficient. The IGV visualized map showed that the information obtained by the two library construction methods was consistent. The sequencing data obtained by the two methods revealed more signal enrichment with Tn5 transposase library construction than with the conventional approach. H3K4me3 ChIP results showed a good reproducibility after Tn5 transposase library construction with a signal-to-noise ratio above 50%.@*CONCLUSIONS@#Tn5 transposase method improves the efficiency of DNA library construction and the results of subsequent sequence analysis, and is especially suitable for detecting histone modification in the DNA to provide a better technical option for epigenetic studies.

Effect of the chemoprotectant tempol on anti-tumor activity of cisplatin / 南方医科大学学报

OBJECTIVE@#To investigate the effect of the chemoprotectant tempol on the anti-tumor activity of cisplatin (DDP).@*METHODS@#The cellular toxicity of tempol in human colon cancer SW480 cells and mouse colon cancer CT26 cells were evaluated using MTT and cell counting kit-8 assays. CalcuSyn software analysis was used to determine the interaction between tempol and DDP in inhibition of the cell viability. A subcutaneous homograft mouse model of colon cancer was established. The mice were randomly divided into control group, tempol group, cisplatin group and tempol + DDP treatment group with intraperitoneal injections of the indicated agents. The tumor size, body weight and lifespan of the mice were measured, and HE staining was used to analyze the cytotoxic effect of the agents on the kidney and liver. Immunohistochemistry and Western blotting were performed to detect the expression of Bax and Bcl2 in the tumor tissue, and TUNEL staining was used to analyze the tumor cell apoptosis. The level of reactive oxygen species (ROS) in the tumor tissue was determined using flow cytometry.@*RESULTS@#Tempol showed inhibitory effects on the viability of SW480 and CT26 cells. CalcuSyn software analysis showed that tempol had a synergistic anti-tumor effect with DDP (CI < 1). In the homograft mouse model, tempol treatment alone did not produce obvious anti-tumor effect. HE staining showed that the combined use of tempol and DDP alleviated DDP-induced fibrogenesis in the kidneys, but tempol also reduced the anti-tumor activity of DDP. Compared with the mice treated with DDP alone, the mice treated with both tempol and DDP had a significantly larger tumor size ( < 0.01) and a shorter lifespan ( < 0.05). Tempol significantly reversed DDP-induced expression of Bax and Bcl2 in the tumor tissue and tumor cell apoptosis ( < 0.001), and obviously reduced the elevation of ROS level in the tumor tissue induced by DDP treatment ( < 0.05).@*CONCLUSIONS@#Tempol can attenuate the anti-tumor effect of DDP while reducing the side effects of DDP. Caution must be taken and the risks and benefits should be carefully weighed when considering the use of tempol as an anti-oxidant to reduce the toxicities of DDP.

Prenatal diagnosis of partial trisomy 3q in a fetus / 中华医学遗传学杂志

OBJECTIVE@#To carry out prenatal diagnosis for a fetus with ultrasonographic abnormality.@*METHODS@#Chromosomal karyotyping and array comparative genomic hybridization (array-CGH) analysis were applied for the diagnosis. Peripheral blood samples were also taken from the parents for chromosome karyotyping analysis.@*RESULTS@#The fetal karyotype showed additional material of unknown-origin attached to Yq. Array CGH analysis confirmed that the material was derived from 3q22.1q29. The father was found to carry a balanced translocation 46, X, t(Y;3)(q12;q23) (which was diagnosed as 46,XY,Y≥18 elsewhere), whilst the mother was found to be normal.@*CONCLUSION@#3q partial trisomy may present as malformation of multiple systems. Combination of chromosome karyotyping and array-CGH can provide reliable diagnosis for fetuses with abnormalities by ultrasonography.

Analysis of MYO7A gene mutation in a family with non-syndromic autosomal recessive deafness / 中华医学遗传学杂志

OBJECTIVE@#To explore the genetic basis for a family with non-syndromic autosomal recessive deafness.@*METHODS@#The proband and her parents were subjected to physical and audiological examinations. With genomic DNA extracted from peripheral blood samples, next-generation sequencing was carried out using a panel for deafness genes. Suspected mutation was validated by Sanger sequencing and qPCR analysis of her parents.@*RESULTS@#The proband presented bilateral severe sensorineural hearing loss at three days after birth. Her auditory threshold was 110-120 dBnHL but with absence of vestibular and retinal symptoms. Her brother also had deafness but her parents were normal. No abnormality was found upon physical examination of her family members, while audiological examination showed no middle ear or retrocochlear diseases. Next-generation sequencing identified compound heterozygous mutations of the MYO7A gene, including a previously known c.462C>A (p. Cys154Ter) and a novel EX43_46 Del, which were respectively derived from her mother and father.@*CONCLUSION@#The compound heterozygous mutations of the MYO7A gene probably underlie the disease in this family. Our findings has enriched the mutation spectrum for non-syndromic autosomal recessive deafness 2.

Correction to: Gene activation in human cells using CRISPR/Cpf1-p300 and CRISPR/Cpf1-SunTag systems

In the original publication the Supplementary Material and Fig. 2 are incorrect. The correct version is provided in this correction article. The text HBG2 appearing in the article should be read as HBG1.

A miRNA-HERC4 pathway promotes breast tumorigenesis by inactivating tumor suppressor LATS1

The E3 ligase HERC4 is overexpressed in human breast cancer and its expression levels correlated with the prognosis of breast cancer patients. However, the roles of HERC4 in mammary tumorigenesis remain unclear. Here we demonstrate that the knockdown of HERC4 in human breast cancer cells dramatically suppressed their proliferation, survival, migration, and tumor growth in vivo, while the overexpression of HERC4 promoted their aggressive tumorigenic activities. HERC4 is a new E3 ligase for the tumor suppressor LATS1 and destabilizes LATS1 by promoting the ubiquitination of LATS1. miRNA-136-5p and miRNA-1285-5p, expression of which is decreased in human breast cancers and is inversely correlated with the prognosis of breast cancer patients, are directly involved in suppressing the expression of HERC4. In summary, we discover a miRNA-HERC4-LATS1 pathway that plays important roles in the pathogenesis of breast cancer and represents new therapeutic targets for human breast cancer.