Molecular cloning, genomic characterization and over-expression of a novel gene, XRRA1, identified from human colorectal cancer cell HCT116(Clone2_XRR )and macaque testis

BACKGROUND: As part of our investigation into the genetic basis of tumor cell radioresponse, we have isolated several clones with a wide range of responses to X-radiation (XR) from an unirradiated human colorectal tumor cell line, HCT116. Using human cDNA microarrays, we recently identified a novel gene that was down-regulated by two-fold in an XR-resistant cell clone, HCT116(Clone2_XRR). We have named this gene as X-ray radiation resistance associated 1 (XRRA1) (GenBank BK000541). Here, we present the first report on the molecular cloning, genomic characterization and over-expression of the XRRA1 gene. RESULTS: We found that XRRA1 was expressed predominantly in testis of both human and macaque. cDNA microarray analysis showed three-fold higher expression of XRRA1 in macaque testis relative to other tissues. We further cloned the macaque XRRA1 cDNA (GenBank AB072776) and a human XRRA1 splice variant from HCT116(Clone2_XRR )(GenBank AY163836). In silico analysis revealed the full-length human XRRA1, mouse, rat and bovine Xrra1 cDNAs. The XRRA1 gene comprises 11 exons and spans 64 kb on chromosome 11q13.3. Human and macaque cDNAs share 96% homology. Human XRRA1 cDNA is 1987 nt long and encodes a protein of 559 aa. XRRA1 protein is highly conserved in human, macaque, mouse, rat, pig, and bovine. GFP-XRRA1 fusion protein was detected in both the nucleus and cytoplasm of HCT116 clones and COS-7 cells. Interestingly, we found evidence that COS-7 cells which over-expressed XRRA1 lacked Ku86 (Ku80, XRCC5), a non-homologous end joining (NHEJ) DNA repair molecule, in the nucleus. RT-PCR analysis showed differential expression of XRRA1 after XR in HCT116 clones manifesting significantly different XR responses. Further, we found that XRRA1 was expressed in most tumor cell types. Surprisingly, mouse Xrra1 was detected in mouse embryonic stem cells R1. CONCLUSIONS: Both XRRA1 cDNA and protein are highly conserved among mammals, suggesting that XRRA1 may have similar functions. Our results also suggest that the genetic modulation of XRRA1 may affect the XR responses of HCT116 clones and that XRRA1 may have a role in the response of human tumor and normal cells to XR. XRRA1 might be correlated with cancer development and might also be an early expressed gene

Cynomolgus monkey testicular cDNAs for discovery of novel human genes in the human genome sequence

BACKGROUND: In order to contribute to the establishment of a complete map of transcribed regions of the human genome, we constructed a testicular cDNA library for the cynomolgus monkey, and attempted to find novel transcripts for identification of their human homologues. RESULT: The full-insert sequences of 512 cDNA clones were determined. Ultimately we found 302 non-redundant cDNAs carrying open reading frames of 300 bp-length or longer. Among them, 89 cDNAs were found not to be annotated previously in the Ensembl human database. After searching against the Ensembl mouse database, we also found 69 putative coding sequences have no homologous cDNAs in the annotated human and mouse genome sequences in Ensembl. We subsequently designed a DNA microarray including 396 non-redundant cDNAs (with and without open reading frames) to examine the expression of the full-sequenced genes. With the testicular probe and a mixture of probes of 10 other tissues, 316 of 332 effective spots showed intense hybridized signals and 75 cDNAs were shown to be expressed very highly in the cynomolgus monkey testis, but not ubiquitously. CONCLUSIONS: In this report, we determined 302 full-insert sequences of cynomolgus monkey cDNAs with enough length of open reading frames to discover novel transcripts as human homologues. Among 302 cDNA sequences, human homologues of 89 cDNAs have not been predicted in the annotated human genome sequence in the Ensembl. Additionally, we identified 75 dominantly expressed genes in testis among the full-sequenced clones by using a DNA microarray. Our cDNA clones and analytical results will be valuable resources for future functional genomic studies

Collection of Macaca fascicularis cDNAs derived from bone marrow, kidney, liver, pancreas, spleen, and thymus

<p>Abstract</p> <p>Background</p> <p>Consolidating transcriptome data of non-human primates is essential to annotate primate genome sequences, and will facilitate research using non-human primates in the genomic era. <it>Macaca fascicularis </it>is a macaque monkey that is commonly used for biomedical and ecological research.</p> <p>Findings</p> <p>We constructed cDNA libraries of <it>Macaca fascicularis</it>, derived from tissues obtained from bone marrow, liver, pancreas, spleen, and thymus of a young male, and kidney of a young female. In total, 5'-end sequences of 56,856 clones were determined. Including the previously established cDNA libraries from brain and testis, we have isolated 112,587 cDNAs of <it>Macaca fascicularis</it>, which correspond to 56% of the curated human reference genes.</p> <p>Conclusion</p> <p>These sequences were deposited in the public sequence database as well as in-house macaque genome database <url>http://genebank.nibio.go.jp/qfbase/</url>. These data will become valuable resources for identifying functional parts of the genome of macaque monkeys in future studies.</p

Transient Spinal Segmental Myoclonus during Neuraxial Anesthesia: A Report of Two Cases

We report here two cases of transient spinal segmental myoclonus occurred during neuraxial anesthesia. One case was an 82-year-old woman, who was scheduled to undergo surgery for abdominal aortic aneurism, was indwelled epidural catheter the day before surgery. A quick, involuntary muscle jerk of the lower limbs began 30 minutes after giving a test dose bolus injection of mepivacaine via the epidural catheter. Despite removal of the epidural catheter, the involuntary muscle jerk persisted and ceased spontaneously five hours after injection of mepivacaine. The other case was a 33-year-old woman who underwent an emergency Caesarean section under a combined spinalepidural anesthesia. Involuntary muscle jerk of the upper limbs began 76 minutes after the intrathecal administration of hyperbaric bupivacaine during immediate post-operative period. The involuntary muscle contraction ceased four hours after injection of bupivacaine without any treatment. In both cases, no abnormal findings were detected in the spinal cord in MRI and/or CT. The involuntary muscle jerk observed in those two patients was diagnosed spinal segmental myoclonus by neurologists. The unusual effect of local anesthetics to the spinal cord neurons during neuraxial anesthesia was suspected as the cause of myoclonus. Although the spinal myoclonus caused by neuraxial anesthesia is quite rare and its etiology is unclear, anesthesiologists should be aware of this bizarre and stressful complication.本稿の要旨は日本麻酔科学会　中国・四国支部第50回学術集会（高松市，2013年）で発表した

Tactile Hypoesthesia Associated with Myofascial Trigger Points in Patients with Persistent Post-Mastectomy Pain: A Close Observation Study in A Case Series

Purpose: Numbness is frequently seen at or near the operated site in patients with persistent post-mastectomy pain (PPMP). Numbness including tactile hypoesthesia can be also seen in patients with myofascial pain. The purpose of this study was to observe relation between such tactile hypoesthesia and myofascial trigger points in patients with PPMP. Methods: We studied five female patients from 43 to 74 years old who consulted at our pain clinic for treatment of PPMP. The areas of tactile hypoesthesia and anesthesia, myofascial trigger points, as well as diffuse tenderness, were marked on the skin surface with delineating such areas and putting X mark on the site of myofascial trigger points using aqueous markers and recorded in photographs at every visit. Results: Tactile hypoesthesia and anesthesia were observed in five and four patients, respectively. Myofascial trigger points were identified in all the five patients and were located within, in the vicinity or near the same dermatomal levels of the hypoesthesia. A series of trigger point injections reduced NRS scores by more than 50% in all patients. The size of the area of tactile hypoesthesia was significantly reduced in association with the decrease of NRS scores in four patients. Conclusions: Our findings indicate that tactile hypoesthesia is closely associated with myofascial pain, and myofascial pain is one potential pathophysiological cause of prolongation and exacerbation of PPMP. Our results also indicate that treating them with trigger point injections are useful for alleviating PPMP

Immune Modulation by Telomerase-Specific Oncolytic Adenovirus Synergistically Enhances Antitumor Efficacy with Anti-PD1 Antibody

The clinical benefit of monotherapy involving immune checkpoint inhibitors (ICIs) such as anti-programmed death-1 antibody (PD-1 Ab) is limited to small populations. We previously developed a telomerase-specific oncolytic adenovirus, Telomelysin (OBP-301), the safety of which was confirmed in a phase I clinical study. Here, we examined the potential of OBP-502, an OBP-301 variant, as an agent for inducing immunogenic cell death (ICD) and synergistically enhancing the efficacy of OBP-502 with PD-1 Ab using CT26 murine colon cancer and PAN02 murine pancreatic cancer cell lines. OBP-502 induced the release of ICD molecules such as adenosine triphosphate (ATP) and high-mobility group box protein 1 (HMGB1) from CT26 and PAN02 cells, leading to recruitment of CD8-positive lymphocytes and inhibition of Foxp3-positive lymphocyte infiltration into tumors. Combination therapy involving OBP-502 intratumoral administration and PD-1 Ab systemic administration significantly suppressed the growth of not only OBP-502-treated tumors but also tumors not treated with OBP-502 (so-called abscopal effect) in CT26 and PAN02 bilateral subcutaneous tumor models, in which active recruitment of CD8-positve lymphocytes was observed even in tumors not treated with OBP-502. This combined efficacy was similar to that observed in a CT26 rectal orthotopic tumor model involving liver metastases. In conclusion, telomerase-specific oncolytic adenoviruses are promising candidates for combined therapies with ICIs

Rate of Evolution in Brain-Expressed Genes in Humans and Other Primates

Brain-expressed genes are known to evolve slowly in mammals. Nevertheless, since brains of higher primates have evolved rapidly, one might expect acceleration in DNA sequence evolution in their brain-expressed genes. In this study, we carried out full-length cDNA sequencing on the brain transcriptome of an Old World monkey (OWM) and then conducted three-way comparisons among (i) mouse, OWM, and human, and (ii) OWM, chimpanzee, and human. Although brain-expressed genes indeed appear to evolve more rapidly in species with more advanced brains (apes > OWM > mouse), a similar lineage effect is observable for most other genes. The broad inclusion of genes in the reference set to represent the genomic average is therefore critical to this type of analysis. Calibrated against the genomic average, the rate of evolution among brain-expressed genes is probably lower (or at most equal) in humans than in chimpanzee and OWM. Interestingly, the trend of slow evolution in coding sequence is no less pronounced among brain-specific genes, vis-à-vis brain-expressed genes in general. The human brain may thus differ from those of our close relatives in two opposite directions: (i) faster evolution in gene expression, and (ii) a likely slowdown in the evolution of protein sequences. Possible explanations and hypotheses are discussed

Hypofractionated stereotactic radiotherapy for acoustic neuromas: safety and effectiveness over 8 years of experience.

Little information is available about long-term outcomes of hypofractionated stereotactic radiotherapy (hypo-FSRT) for acoustic neuromas. In this study, the safety and effectiveness of hypo-FSRT for unilateral acoustic neuroma were reviewed over 8 years of experience at our institution

Identification of the sex-determining factor in the liverwort Marchantia polymorpha reveals unique evolution of sex chromosomes in a haploid system

半数体生物の性染色体上の性決定遺伝子を解明 --コケがもつ現生生物最古の起源の性染色体--. 京都大学プレスリリース. 2021-11-08.Sex determination is a central process for sexual reproduction and is often regulated by a sex determinant encoded on a sex chromosome. Rules that govern the evolution of sex chromosomes via specialization and degeneration following the evolution of a sex determinant have been well studied in diploid organisms. However, distinct predictions apply to sex chromosomes in organisms where sex is determined in the haploid phase of the life cycle: both sex chromosomes, female U and male V, are expected to maintain their gene functions, even though both are non-recombining. This is in contrast to the X-Y (or Z-W) asymmetry and Y (W) chromosome degeneration in XY (ZW) systems of diploids. Here, we provide evidence that sex chromosomes diverged early during the evolution of haploid liverworts and identify the sex determinant on the Marchantia polymorpha U chromosome. This gene, Feminizer, encodes a member of the plant-specific BASIC PENTACYSTEINE transcription factor family. It triggers female differentiation via regulation of the autosomal sex-determining locus of FEMALE GAMETOPHYTE MYB and SUPPRESSOR OF FEMINIZATION. Phylogenetic analyses of Feminizer and other sex chromosome genes indicate dimorphic sex chromosomes had already been established 430 mya in the ancestral liverwort. Feminizer also plays a role in reproductive induction that is shared with its gametolog on the V chromosome, suggesting an ancestral function, distinct from sex determination, was retained by the gametologs. This implies ancestral functions can be preserved after the acquisition of a sex determination mechanism during the evolution of a dominant haploid sex chromosome system