Identification and characterization of a novel group of natural antisense transcripts from RNA1.2 gene locus of human cytomegalovirus / 中华医学杂志(英文版)

Background@#Natural anti-sense transcripts (NATs), which are transcribed from the complementary DNA strand of annotated genes, exert regulatory function of gene expression. Increasing studies recognized anti-sense transcription widespread throughout human cytomegalovirus (HCMV) genome, whereas the anti-sense transcription of RNA1.2 gene locus has never been investigated. In this study, the transcription of the RNA1.2 anti-sense strand was investigated in clinically isolated HCMV strain.@*Methods@#Strand-specific high-through RNA-sequencing (RNA-seq) was performed to find possible anti-sense transcripts (ASTs). For analyzing and visualization of RNA-seq data sets, Integrative Genomics Viewer software was applied. To confirm these possibilities, Northern blotting and rapid amplification of cDNA ends (RACE) were used.@*Results@#Transcription of the opposite strand of RNA1.2 gene locus was detected by RNA-sequencing using RNAs extracted from human embryonic lung fibroblasts infected with HCMV clinical isolate HAN. At least three HCMV NATs, named RNA1.2 AST 1, RNA1.2 AST2, and RNA1.2 AST3, were characterized by Northern blotting and RACE analyses. These RNA1.2 ASTs orientated from the complementary strand of RNA1.2 locus during the late phase of HCMV infection. The 5′- and 3′-termini of these transcripts were located within the opposite sequence of the predicted RNA1.2 gene.@*Conclusion@#A cluster of novel NATs was transcribed from the opposite sequence of the HCMV RNA1.2 gene region.

Identification and characterization of a novel group of natural anti-sense transcripts from RNA1.2 gene locus of human cytomegalovirus / 中华医学杂志(英文版)

BACKGROUND@#Natural anti-sense transcripts (NATs), which are transcribed from the complementary DNA strand of annotated genes, exert regulatory function of gene expression. Increasing studies recognized anti-sense transcription widespread throughout human cytomegalovirus (HCMV) genome, whereas the anti-sense transcription of RNA1.2 gene locus has never been investigated. In this study, the transcription of the RNA1.2 anti-sense strand was investigated in clinically isolated HCMV strain.@*METHODS@#Strand-specific high-through RNA-sequencing (RNA-seq) was performed to find possible anti-sense transcripts (ASTs). For analyzing and visualization of RNA-seq data sets, Integrative Genomics Viewer software was applied. To confirm these possibilities, Northern blotting and rapid amplification of cDNA ends (RACE) were used.@*RESULTS@#Transcription of the opposite strand of RNA1.2 gene locus was detected by RNA-sequencing using RNAs extracted from human embryonic lung fibroblasts infected with HCMV clinical isolate HAN. At least three HCMV NATs, named RNA1.2 AST 1, RNA1.2 AST2, and RNA1.2 AST3, were characterized by Northern blotting and RACE analyses. These RNA1.2 ASTs orientated from the complementary strand of RNA1.2 locus during the late phase of HCMV infection. The 5'- and 3'-termini of these transcripts were located within the opposite sequence of the predicted RNA1.2 gene.@*CONCLUSION@#A cluster of novel NATs was transcribed from the opposite sequence of the HCMV RNA1.2 gene region.

Retrospective analysis of clinical features and prognostic factors for pathological fracture in 39 patients with extremity osteosarcoma / 肿瘤

Objective: To identify the clinical features and prognostic factors in relation to extremity osteosarcoma with pathological fracture. Methods: The clinical data from 244 patients with extremity osteosarcoma between October 2003 and October 2011 were retrospectively reviewed. These patients were divided into two groups: patients with pathological fracture group and the patients without pathological fracture group. The differences in the clinical features of the two groups were analyzed. The survival analysis was performed using Kaplan-Meier method. For patients with pathological fracture, a univariate analysis (log-rank) was used to determine the prognostic factors related to the survival, and a COX proportional hazards regression model was used to identify the independent prognostic factors. Results: A higher proportion of patients with larger tumor size (P = 0.012), humeral osteosarcoma (P = 0.004) or local recurrence (P = 0.002) was observed in patients with pathological fracture. Additionally, more patients with pathological fracture received an amputation surgery, as compared with patients without pathological fracture (P = 0.032). The median survival time of patients with pathological fracture was significantly shorter than that of patients without pathological fracture (16 vs 21 months, P = 0.006). The univariate analysis showed that the significant prognosis-related factors were the tumor size, Enneking's surgical staging, KPS (Karnofsky performance status) score, cycles of adjuvant chemotherapy, local recurrence and metastasis (P < 0.05). The multivariate analysis revealed that the factors of KPS score, cycles of adjuvant chemotherapy and metastasis were the independent prognostic factors of extremity osteosarcoma with pathological fracture. Conclusion: Compared with the patients without pathological fracture, a higher proportion of patients receiving amputation surgery or having larger tumor size, humeral osteosarcoma or local recurrence was observed in patients with pathological fracture, and the prognosis of these patients was poor. The KPS score, cycles of adjuvant chemotherapy and metastasis were independent prognostic factors of extremity osteosarcoma with pathological fracture. Copyright © 2012 by TUMOR.

Sequence conservation of human cytomegalovirus UL140 open reading frame in clinical strains / 中国医学科学杂志(英文版)

<p><b>OBJECTIVE</b>To investigate the variability of human cytomegalovirus (HCMV) UL140 open reading frame (ORF) in clinical strains, and to explore the relationship between the variability of UL140 ORF and different symptoms of HC-MV infection.</p><p><b>METHODS</b>HCMV UL140 ORF was amplified by polymerase chain reaction and sequenced selectedly in 30 clinical strains.</p><p><b>RESULTS</b>UL140 ORF of all clinical strains was amplified successfully. Compared with that of Toledo strain, the nucleotide and amino acid sequence identities among all strains were 96.5%-100.0% and 95.2%-100.0%, respectively. All of the nucleotide changes were substitutions. The post-translational modification sites were conserved. The result of phylogenetic tree showed that the strains did not cluster according to different clinical symptoms.</p><p><b>CONCLUSION</b>HCMV UL140 ORF in clinical strains is highly conserved, which may play an important role in HC-MV infection.</p>

Polymorphism of human cytomegalovirus UL134 gene in low-passage clinical isolates / 中国当代儿科杂志

<p><b>OBJECTIVE</b>Human cytomegalovirus (HCMV) displays genetic polymorphisms. Nineteen open reading frames (ORFs, UL133-UL151) found in the Toledo strain of HCMV and other low-passage clinical isolates may be essential for viral infection. This study aimed to analyze the polymorphism of HCMV UL134 gene in clinical isolates and explore the relationship between the polymorphism and HCMV infection.</p><p><b>METHODS</b>PCR was performed to amplify entire UL134 region in 32 clinical isolates, which had been proven as HCMV-DNA positive by FQ-PCR. PCR products were sequenced.</p><p><b>RESULTS</b>All of the 32 isolates were amplified and sequenced successfully. HCMV UL134 gene was highly conserved in the clinical isolates. UL134 ORF and its predicted protein in the clinical strains displayed 96.4%-98.3% nucleotide identity and 92.7%-94.9% amino acid identity respectively compared to those in the Toledo strain. A new posttranslational modification site, sulfationcamp (SUL) site, was found in UL134 protein of all of the clinical isolates except 35j.</p><p><b>CONCLUSIONS</b>HCMV UL134 gene in clinical isolates was highly conserved. No substantial relation was found between UL134 gene and HCMV infectious diseases.</p>

Sequence variability of human cytomegalovirus UL143 in low-passage clinical isolates / 中华医学杂志(英文版)

<p><b>BACKGROUND</b>Human cytomegalovirus (HCMV) infects a number of organs and tissues in vivo. The different symptoms and tissue tropisms of HCMV infection perhaps result from genetic polymorphism. A new region of DNA containing at least 19 open reading frames (ORFs) (denoted UL133 to 151) was found in the low-passage HCMV clinical strain, Toledo, and several other low-passage clinical isolates, but not present in the HCMV laboratory strain, AD169. One of these genes, UL143, was studied to explore the sequence variability of UL143 ORF in HCMV clinical isolates and examine the possible association between gene variability and the outcome of HCMV infection.</p><p><b>METHODS</b>The UL143 gene of the strains obtained from suspected congenitally HCMV-infected infants was amplified by polymerase chain reaction (PCR) and sequenced.</p><p><b>RESULTS</b>Nineteen sequences of the strains were divided into 2 major groups, G(1) (n = 16) and G(2) (n = 3). All of the sequences had frame-shift mutation compared to Toledo. Nucleotide polymorphisms conferred substantial amino acid substitutions when compared with Toledo. All 16 UL143 putative proteins of the strains in G(1) had a new myristylation site and loss of two PKC sites owing to missense mutations. No convincing relationships were observed between the presence of HCMV disease and the UL143 sequence group.</p><p><b>CONCLUSIONS</b>HCMV-UL143 existed in low passage isolates. Sequence variability caused by frame-shift mutation was found in all HCMV clinical strains. No obvious linkage was observed between UL143 polymorphisms and the outcome of suspected congenital HCMV infection.</p>

High variability of human cytomegalovirus UL150 open reading frame in low-passaged clinical isolates / 中国医学科学杂志(英文版)

<p><b>OBJECTIVE</b>To investigate the polymorphism of human cytomegalovirus (HCMV) UL150 open reading frame (ORF) in low-passaged clinical isolates, and to study the relationship between the polymorphism and different pathogenesis of congenital HCMV infection.</p><p><b>METHODS</b>PCR was performed to amplify the entire HCMV UL150 ORF region of 29 clinical isolates, which had been proven containing detectable HCMV-DNA using fluorescence quantitative PCR. PCR amplification products were sequenced directly, and the data were analyzed.</p><p><b>RESULTS</b>Totally 25 among 29 isolates were amplified, and 18 isolates were sequenced successfully. HCMV UL150 ORF sequences derived from congenitally infected infants were high variability. The UL150 ORF in all 18 clinical isolates shifted backward by 8 nucleotides leading to frame-shift, and contained a single nucleotide deletion at nucleotide position 226 compared with that of Toledo strain. The nucleotide diversity was 0.1% to 6.8% and the amino acid diversity was 0.2% to 19.2% related to Toledo strain. However, the nucleotide diversity was 0.1% to 6.4% and amino acid diversity was 0.2% to 8.3% by compared with Merlin strain. Compared with Toledo, 4 new cysteine residues and 13 additional posttranslational modification sites were observed in UL150 putative proteins of clinical isolates. Moreover, the UL150 putative protein contained an additional transmembrane helix at position of 4-17 amino acid related to Toledo.</p><p><b>CONCLUSION</b>HCMV UL150 ORF and deduced amino acid sequences of clinical strains are hypervariability. No obvious linkage between the polymorphism and different pathogenesis of congenital HCMV infection is found.</p>