Month: January 2020

Considering novel poly isolated mRNAs is whether they are protein coding or not. Human RNase k is a previously established protein belonging to a family conserved in all metazoans. The high conservation of all members along with the fact that the human enzyme is widely expressed in all tissues and developmental stages suggest a very important biological function that is currently under investigation. Human RNase k is represented by a single copy in the human genome and retains the pattern of gene organization that is preserved by almost all known members of the gene family, consisting of three exons interrupted by two introns. The human representative exhibits in vitro endoribonucleolytic activity cleaving preferentially ApG and ApU phosphodiester bonds, while it hydrolyzes UpU bonds at a lower rate. Reducing reagents and site-directed mutagenesis experiments showed that a disulfide bond between cysteine residues 6 and 69 is essential for the ribonucleolytic activity of the enzyme. The present work reports a human subtle alternative splicing event giving rise to a protein coding mRNA variant that lacks 4 nucleotides compared to the previously reported RNase k mRNA isoform. Cloning and expression analysis of this subtly alternatively spliced mRNA was achieved by the development of a modified hybrid selection approach which may provide a tool for the study of similar cases. RNase k-02 mRNA encodes the synthesis of an alternative protein isoform in human cell extracts which follows a cytoplasmic distribution. Subtle alternative splicing events are being observed in an everrising number of human genes, as depicted by the exponential accumulation of EST and high throughput RNA sequencing data. The impact of subtle alternative splicing on protein products depends on whether they are frame-shifting or frame-preserving with D denoting the sequence length between the tandem splice sites. Frame-preserving tandem sites result in the addition or absence of only a few aminoacids, leading to the production of similar protein isoforms which may bear different functional properties. For instance, a D3 subtle alternative splicing event occurring in ATN1 human gene product determines the cellular topology of the corresponding protein isoforms. At the donor site, alternative splicing events in which the distance between two splice sites is 4 nucleotides long are the most frequent. However, their frameshift effect often creates mRNA isoforms that are predicted as nonsense mediated decay targets and therefore these cases have not been systematically investigated. The methodological approach presented allowed us to achieve the cloning of a human RNase k mRNA variant that occurs after a D4 splicing event. This experimental strategy offers several advantages for the study of transcripts generated by alternative splicing compared to conventional RT-PCR approaches.

We now know that ubiquitination participates not only in the proteolytic function but also in many non-proteolytic reactions with crucial roles in cell metabolisms. For example, fluorescence ubiquitination-based cell cycle indicator enabled us to examine cell division within living cells by the Ub-proteasome system. In mammalian cells, there are dozens of E2s and several hundred E3s, and both define families of proteins displaying substrate specificity. However, there are only two E1 enzymes for the entire array of downstream reactions in mammals, Uba1 and Uba6. Uba1 encodes canonical E1. Previously, introduction and expression of epitope-tagged Uba1 cDNA constructs revealed that nuclear and cytoplasmic isoforms of Uba1 translate from first and second ATG codons: E1a, localized predominantly in the nucleus, and E1b, localized in the cytoplasm, respectively. To avoid confusion in terminology, we respectively refer to these two isoforms as Uba1A, defined here as the predominantly nuclear form of Uba1, and Uba1B, defined here as the cytoplasmic form of Uba1, instead of E1a and E1b. Uba6 is required to activate the E2Use1 both in vitro and in vivo and can also activate another ubiquitin-like modifier, FAT10. To identify genes responsible for the maintenance of chromosome integrity, Tsuji and colleagues isolated 25 temperaturesensitive mutants from hamster wild-type CHO-K1 cells. Using two of these mutants, we revealed that ts defects in RNA polymerase II and a protein involved in splicing gave rise both to chromosome instability and to cell cycle arrest. Another ts CHO-K1 mutant, tsTM3, exhibits chromosomal instability and cell-cycle arrest in the S to G2 phases with decreased DNA synthesis at the nonpermissive temperature, 39uC. Complementation tests with other mutants showed that tsTM3 did not complement with the Uba1-defective ts mutant ts85 and DNA replication-defective ts mutant ts131b, suggesting that these mutants harbor the same genetic defect. From 1980 to 1990, many ts mutants of Uba1 were isolated from several cell lines: ts85 of FM3A, ts20 of CHO, ts131b of FM3A, ts20 of Balb/c 3T3, tsBN75 of BHK21, tsFS20 of FM3A, and tsFT5 of FM3A. This unusually high incidence of Uba1 mutations was discussed in terms of Uba1 as a determinant of heat tolerance of cells and the fact that the Uba1 locus is located on the X chromosome. In regard to the connection between Uba1 and human disease, a recent study identified the association of pathogenic mutations in human UBA1 with an early-onset neurodegenerative disorder involving lower motor neurons. It provided evidence that the rare missense and synonymous mutations detected in exon 15 of UBA1 are associated with Xlinked spinal muscular atrophy. In the present study, to identify the mechanism underlying the tsTM3 phenotypes, we performed sequence analysis of the Uba1 gene and investigated the relation between a wild-type isoform of Uba1 tagged with green fluorescent protein and its localization.

If similar differences exist within or between cohorts of wild fishes, SSB may over estimate reproductive potential because differences between females in egg quality are not taken into account. This takes on added significance if transcriptomic fingerprints shift toward higher fertility profiles with increasing age as, despite evidence to the contrary, most current fishery management models consider many small females to be equivalent to a few larger ones if the ovarian biomass is comparable. Transcriptomic profiling may provide an expedient means to better assess the reproductive health of fisheries and other animal resources important to global food security and has far reaching implications in agriculture and in medical fields such as human assisted reproductive technology. We discovered that it is possible to predict the state of a biologically complex fitness trait, egg quality, with remarkable accuracy based solely on ovary gene expression profiles. The collective expression of a discrete suite of ovarian transcripts, constituting a transcriptomic fingerprint, involved specific, highlyconserved gene networks central to early embryonic development in all vertebrates. Our findings open the door to ‘transcriptomeassisted’ breeding and assessment of reproductive condition and value in farmed and possibly wild fishes and other vertebrates, possibly including human assisted reproductive technologies. The ovary transcriptome profiles and ANN analytical methods developed in the present study provide a foundation for future research into the effects of perturbations on egg transcriptome and quality, so that gene networks and molecular pathways most susceptible to a particular insult can be identified. Additionally, we propose that ANNs may be a superior alternative to linear-based analyses of gene expression data that can be used as a diagnostic tool, provided that the ANNs are trained with appropriate data. Light regulates a wide range of plant processes including seed germination, organ, cell and organelle differentiation, flowering and metabolism. The germination of a seed in the dark follows skotomorphogenesis. Upon exposure to light, seedlings go through photomorphogenesis that is marked by chlorophyll biosynthesis, differentiation of protoplastids into chloroplasts, the initiation of carbon assimilation, elongation and thickening of the hypocotyl, and the activation of the shoot apical meristem leading to the development of the first true leaves. Although the transition from skotomorphogenic to photomorphogenic growth has been well-documented in Arabidopsis, the complex gene networks at the genome level controlling this developmental transition in wheat are not well understood. In order to investigate and identify the complex transcriptional network associated with seedling photomorphogenesis in Einkorn wheat.

Among epigenetic aberrations, DNA methylation itself features a diverse presence. Recently, 5-hydroxymethylcytosine has been identified as a constituent of mammalian DNA and described as the sixth base of the genome. The loss of 5-hmC has been highlighted as a hallmark of melanoma by a single, remarkable study, whereas interesting clues as to the role of 5hydroxymethylcytosine are still emerging. In contrast to 5hmC, the importance of 5-methylcytosine in cancer cells is much more firmly established. Aberrant promoter DNA hypermethylation or localised methylation preferably occurs in CpG dinucleotide-dense regions, resulting in the down-regulation of the corresponding gene. It has recently become apparent that malignant melanomas feature hypermethylation, and currently more than 80 genes mainly in promoter regions are hypermethylated at a single-gene level. Taking a global view of the available data, the number of primary tumour samples involved in the studies and the frequency of positive results do not allow determining whether the hypermethylated genes described are appropriate for diagnosis or can be considered candidate therapeutic targets. Moreover, most of the data provided are derived from cell lines and estimated methylation values indirectly consisting of three steps: measuring mRNA or protein expression in cell lines, treating samples with a specific drug that acts against the process of methylation and measuring gene expression again. Nonetheless, powerful arguments have been presented in the literature that support direct experiments being less ambiguous; furthermore, most of the groups conducting direct measurements have applied candidate gene approaches. In addition to the rapid progress that has been made in studying promoter hypermethylation at the single-gene level, only two groups have attempted to conduct array-based experiments to identify the methylation pattern of thousands of gene promoters. Regrettably, one group has focused only on comparing the methylation level of primary invasive melanomas with benign melanocytes and has clearly identified a group of genes in a statistically powerful interpretation that can be used to discriminate naevi from melanomas based on their methylation signature. Another group has examined the short-term cultures of homogeneous stage III specimens. As no data are currently available regarding the methylation markers of diverse melanomas with different clinical behaviours, we performed a systematic comparison of localised methylation patterns among 42 primary melanomas using the Illumina Golden Gate Cancer Panel Bead Assay. We found differentially methylated CpGs altogether among melanoma subgroups and the majority of CpG sites were hypermethylated in melanomas that represent more favourable prognoses including a non-ulcerated tumour surface, superficial spreading histological subtype, nonmetastatic subgroup and smaller tumour thickness.

GABAergic dysfunctions and alterations in inhibitory circuits of the brain have been implicated in several mouse models of schizophrenia and related disorders ). We suggest that by influencing GABAergic signaling through vesicle trafficking, TMPAP might be linked to specific endophenotypes seen in neurological and neuropsychiatric disorders. Residents carry out much of daily patient care, while being learners at the same time. Therefore, patient care quality benefits from adequate supervision of residents. To assess adequate supervision in residency training, attending physicians are increasingly evaluated on their clinical teaching performance. Systems and tools for robust evaluation of teaching performance are available and high and low performing attending physicians can be identified. Research suggests that attending physicians who are younger, female and spend more time on teaching and conducting research, are more favorably evaluated on their teaching performance. In addition, which is in line with well documented personality research in the field of job performance and academic performance in medicine. Yet, personality research on teaching performance of attending physicians is limited and uses qualitative methods only. Understanding the plausible link between them can shed critical light on opportunities and policies regarding the development paths and career planning of attending physicians in medical education. What are personality traits and in what way could they affect teaching of attending physicians? Personality traits can be categorized in five comprehensive domains, called the Five Factor Model: conscientiousness, extraversion, emotional stability, agreeableness and openness. Conscientiousness refers to dependability and includes traits such as being responsible, organized, orderly and thorough. Employees who are conscientious take responsibility for their work, accomplishing their work tasks more thoroughly and orderly. Ultimately, university teachers who are orderly are better evaluated and physicians who are conscientious are thought to be good teachers in medicine. Higher levels of conscientiousness of attending physicians could positively influence their teaching in residency training, as training of residents is an important responsibility. Yet, the role of conscientiousness in clinical teaching practice remains unexplored. Extraversion means being sociable, talkative, outgoing and active. Extraverted people perform better in professions involving social interaction. Unlike research on the other four personality traits, research on the working mechanism of extraversion with respect to social interaction provided a neurobiological explanation. That is, extraverts have lower cortical activity than introverts. This makes extraverts seek to attain a higher level of arousal by increasing social activity, while the higher levels of cortical activity of introverts.