Increase of Pro-opiomelanocortin mRNA Prior to Tyrosinase, Tyrosinase-Related Protein 1, Dopachrome Tautomerase, Pmel-17/gp100, and P-Protein mRNA in Human Skin After Ultraviolet B Irradiation

In ultraviolet-induced tanning, the protein levels of various gene products critical for pigmentation (including tyrosinase and tyrosinase-related protein-1) are increased in response to ultraviolet B irradiation, but changes in mRNA levels of these factors have not been investigated in vivo. We have established an in situ hybridization technique to investigate mRNA levels of pro-opiomelanocortin, tyrosinase, tyrosinase-related protein-1, dopachrome tautomerase, P-protein, Pmel-17/gp100, and microphthalmia-associated transcription factor, and have analyzed the changes in mRNA levels in the ultraviolet B-exposed skin in vivo. The right or left forearm of each volunteer was irradiated with ultraviolet B, and skin biopsies were obtained at 2 and 5 d postirradiation. mRNA level of pro- opiomelanocortin was increased 2 d after ultraviolet B irradiation, and returned to a near-basal level after 5 d, whereas the mRNA levels of tyrosinase, tyrosinase-related protein-1, dopachrome tautomerase, P-protein, and Pmel-17/gp100 showed some or no increase at 2 d, but were significantly increased 5 d after ultraviolet B irradiation. Microphthalmia-associated transcription factor mRNA was slightly increased on days 2 and 5 after ultraviolet B irradiation. Our results suggest that the mechanism of the tanning response of human skin may involve the transcriptional regulation of certain pigmentary genes, and that pro-opiomelanocortin-derived melanocortins such as α-melanocyte-stimulating hormone and adrenocorticotropic hormone may play a part in regulating these genes in vivo

Construction and characterization of a BAC library from a gynogenetic channel catfish Ictalurus punctatus

A bacterial artificial chromosome (BAC) library was constructed by cloning HindIII-digested high molecular weight DNA from a gynogenetic channel catfish, Ictalurus punctatus, into the vector pBeloBAC11. Approximately 53 500 clones were arrayed in 384-well plates and stored at -80°C (CCBL1), while clones from a smaller insert size fraction were stored at -80°C without arraying (CCBL2). Pulsed-field gel electrophoresis of 100 clones after NotI digestion revealed an average insert size of 165 kb for CCBL1 and 113 kb for CCBL2. Further characterization of CCBL1 demonstrated that 10% of the clones did not contain an insert. CCBL1 provides a 7.2-fold coverage of the channel catfish haploid genome. PCR-based screening demonstrated that 68 out of 74 unique loci were present in the library. This represents a 92% chance to find a unique sequence. These libraries will be useful for physical mapping of the channel catfish genome, and identification of genes controlling major traits in this economically important species

不揮発性分子のレーザー蒸発/レーザー分光の装置開発

第4回分子科学討論会, 2010年9月14日-17日, 大阪大学豊中キャンパス(大阪), 4A1

A BAC-based physical map of the Nile tilapia genome

BACKGROUND: Cichlid fishes, particularly tilapias, are an important source of animal protein in tropical countries around the world. To support selective breeding of these species we are constructing genetic and physical maps of the tilapia genome. Physical maps linking collections of BAC clones are a critical resource for both positional cloning and assembly of whole genome sequences. RESULTS: We constructed a genome-wide physical map of the tilapia genome by restriction fingerprinting 35,245 bacterial artificial chromosome (BAC) clones using high-resolution capillary polyacrylamide gel electrophoresis. The map consists of 3,621 contigs and is estimated to span 1.752 Gb in physical length. An independent analysis of the marker content of four contigs demonstrates the reliability of the assembly. CONCLUSION: This physical map is a powerful tool for accelerating genomic studies in cichlid fishes, including comparative mapping among fish species, long-range assembly of genomic shotgun sequences, and the positional cloning of genes underlying important phenotypic traits. The tilapia BAC fingerprint database is freely available at

Neural Sources of Vagus Nerve Stimulation–Induced Slow Cortical Potentials

[Objectives] This study investigated neuronal sources of slow cortical potentials (SCPs) evoked during vagus nerve stimulation (VNS) in patients with epilepsy who underwent routine electroencephalography (EEG) after implantation of the device. [Materials and Methods] We analyzed routine clinical EEG from 24 patients. There were 5 to 26 trains of VNS during EEG. To extract SCPs from the EEG, a high-frequency filter of 0.2 Hz was applied. These EEG epochs were averaged and used for source analyses. The averaged waveforms for each patient and their grand average were subjected to multidipole analysis. Patients with at least 50% seizure frequency reduction were considered responders. Findings from EEG analysis dipole were compared with VNS responses. [Results] VNS-induced focal SCPs whose dipoles were estimated to be located in several cortical areas including the medial prefrontal cortex, postcentral gyrus, and insula, with a significantly higher frequency in patients with a good VNS response than in those with a poor response. [Conclusions] This study suggested that some VNS-induced SCPs originating from the so-called vagus afferent network are related to the suppression of epileptic seizures

Hyper-expansion of large DNA segments in the genome of kuruma shrimp, Marsupenaeus japonicus

<p>Abstract</p> <p>Background</p> <p>Higher crustaceans (class Malacostraca) represent the most species-rich and morphologically diverse group of non-insect arthropods and many of its members are commercially important. Although the crustacean DNA sequence information is growing exponentially, little is known about the genome organization of Malacostraca. Here, we constructed a bacterial artificial chromosome (BAC) library and performed BAC-end sequencing to provide genomic information for kuruma shrimp (<it>Marsupenaeus japonicus</it>), one of the most widely cultured species among crustaceans, and found the presence of a redundant sequence in the BAC library. We examined the BAC clone that includes the redundant sequence to further analyze its length, copy number and location in the kuruma shrimp genome.</p> <p>Results</p> <p>Mj024A04 BAC clone, which includes one redundant sequence, contained 27 putative genes and seemed to display a normal genomic DNA structure. Notably, of the putative genes, 3 genes encode homologous proteins to the inhibitor of apoptosis protein and 7 genes encode homologous proteins to white spot syndrome virus, a virulent pathogen known to affect crustaceans. Colony hybridization and PCR analysis of 381 BAC clones showed that almost half of the BAC clones maintain DNA segments whose sequences are homologous to the representative BAC clone Mj024A04. The Mj024A04 partial sequence was detected multiple times in the kuruma shrimp nuclear genome with a calculated copy number of at least 100. Microsatellites based BAC genotyping clearly showed that Mj024A04 homologous sequences were cloned from at least 48 different chromosomal loci. The absence of micro-syntenic relationships with the available genomic sequences of <it>Daphnia </it>and <it>Drosophila </it>suggests the uniqueness of these fragments in kuruma shrimp from current arthropod genome sequences.</p> <p>Conclusions</p> <p>Our results demonstrate that hyper-expansion of large DNA segments took place in the kuruma shrimp genome. Although we analyzed only a part of the duplicated DNA segments, our result suggested that it is difficult to analyze the shrimp genome following normal analytical methodology. Hence, it is necessary to avoid repetitive sequence (such as segmental duplications) when studying the other unique structures in the shrimp genome.</p

Immunohistochemical Ki67 after short-term hormone therapy identifies low-risk breast cancers as reliably as genomic markers

Background: The purpose of this study was to test whether immunohistochemical (IHC) Ki67 levels after short-term preoperative hormone therapy (post-Ki67) predict similar numbers of patients with favorable prognoses as genomic markers. Results: Thirty paired cases (60 samples) were enrolled in this study. Post-Ki67 levels were significantly lower than pre-treatment Ki67 levels (P < 0.001). Post-Ki67 predicted more low-risk cases (83.3%, 25/30) than pre-genomic surrogate signature(GSS) (66.7%: 20/30), but the difference in predictive power was not significant (P = 0.233). Proliferation (MKI67, STK15, Survivin, CCNB1, and MYBL2) and estrogen (ER, PGR, BCL2, and SCUBE2) related signatures were significantly downregulated after therapy (P < 0.001 and 0.041, respectively). Materials and Methods: Core needle biopsy specimens of primary breast cancer were collected at Okayama University Hospital from hormone receptor-positive and human epidermal growth factor 2-negative patients that subsequently received two weeks of neoadjuvant hormone therapy. Paired post-treatment specimens from surgical samples were also collected. IHC Ki67 levels and GSS were compared between pre- and post-hormone treatment samples. Changes of gene expression pattern in short-term hormone therapy were also assessed. Conclusions: IHC based post-Ki67 levels may have distinct predictive power compared with the naïve IHC Ki67. Future studies with larger cohorts and longer follow-up periods may be needed to validate our results