Chromosome Fusions Triggered by piRNAs in the Ciliate Oxytricha trifallax

Chromosome fusions, common in cancer cells, result from telomere loss at chromosome ends (Liu, Cheng, & Hsieh, 2009). Complicated to grasp mechanistically, these fusions may result in aberrant gene products as well as prevent deterioration and cleavage of chromosome isoforms (Wang et al., submitted). Currently, the ciliate Oxytricha trifallax holds the record for the most fragments during the fragmentation process in conjugation. Oxytricha is capable of separating into an 8-gene nanochromosome (Swart et al., 2013). The research presented in this thesis discovers the robustness of piRNA injection to induce chromosome fusions and the influential role that these small oligonucleotides play in the origin of multi-gene chromosomes. O. trifallax has been pushed to form a 5-gene isoform via triggering the fusion of three adjacent chromosomes from a single MIC locus by microinjections of 27 nt piRNAs. Fusions assayed by PCR showed a viable triple fusion of the MAC DNA of contigs 11682.0, 20527.0, and 16348.0; however, fusions assayed by Southern analysis only confirmed a double fusion of contigs 20527.0 and 16348.0 when probed with contig20527.0. Nonetheless, the linkage of 5 genes was accomplished representing novel research for the strength of piRNAs in ciliate systems such as Oxytricha. Therefore, piRNAs act as mediators in the linkage of adjacent genes in MIC DNA while protecting DNA sequences from aberrant breakages or deletions

Chromosome fusions triggered by noncoding RNA

Chromosomal fusions are common in normal and cancer cells and can produce aberrant gene products that promote transformation. The mechanisms driving these fusions are poorly understood, but recurrent fusions are widespread. This suggests an underlying mechanism, and some authors have proposed a possible role for RNA in this process. The unicellular eukaryote Oxytricha trifallax displays an exorbitant capacity for natural genome editing, when it rewrites its germline genome to form a somatic epigenome. This developmental process provides a powerful model system to directly test the influence of small noncoding RNAs on chromosome fusion events during somatic differentiation. Here we show that small RNAs are capable of inducing chromosome fusions in four distinct cases (out of four tested), including one fusion of three chromosomes. We further show that these RNA-mediated chromosome fusions are heritable over multiple sexual generations and that transmission of the acquired fusion is associated with endogenous production of novel piRNA molecules that target the fused junction. We also demonstrate the capacity of a long noncoding RNA (lncRNA) to induce chromosome fusion of two distal germline loci. These results underscore the ability of short-lived, aberrant RNAs to act as drivers of chromosome fusion events that can be stably transmitted to future generations

Characterization of the hepatitis B virus DNA detected in urine of chronic hepatitis B patients

Abstract Background Detection of human hepatitis B virus (HBV) DNA in the urine of patients with chronic hepatitis B infection (CHB) has been reported previously, suggesting urine could provide a potential route of horizontal HBV transmission. However, it is not clear whether the HBV DNA detected in urine is indeed full-length, infectious viral DNA. The aim of this study is to assess the potential infectivity of urine from patients with CHB and to correlate HBV DNA detection in urine with clinical parameters, such as serum viral load and HBeAg status. Methods Urine from 60 CHB patients with serum viral loads ranging from undetectable to 108 IU/mL were analyzed for HBV DNA and serum immune markers. HBV DNA was detected from total urine DNA and size-fractionated urine DNA (separated into ≤1 kb and > 1 kb fractions) by PCR analysis of six regions of the HBV genome. Results Twenty-seven of 59 (45.7%) patients with HBV serum viral load (≥20 IU/mL) contained at least 20 copies per mL of fragmented HBV DNA in urine detected in at least 1 of the 6 PCR assay regions. Only one patient contained HBV DNA detected by all six regions, and was found to have evidence of blood in the urine. Sixteen of 25 urine samples with high viral load (> 105 IU/mL) and 11 of 34 urine samples with low viral load (< 105 IU/mL) contained detectable HBV DNA. Twelve of 27 (44.44%) patients with detectable HBV DNA in urine were HBeAg positive, and only 5 of these HBeAg positive patients were in the group of 33 (15.15%) patients with no detectable HBV DNA in urine. By Fishers’ exact test, HBV DNA in urine is significantly associated with high serum viral load (P = 0.0197) and HBeAg (P = 0.0203). Conclusions We conclude that urine from CHB patients with healthy kidney function should not contain full-length HBV DNA, and therefore should not be infectious

Additional file 1: of Characterization of the hepatitis B virus DNA detected in urine of chronic hepatitis B patients

Urine DNA analysis by qPCR assays targeting multiple locations in the HBV genome. Raw data for Fig. 2. (XLSX 18 kb