Yun-Ji Kim

Selection of internal reference genes for SYBR green qRT-PCR studies of rhesus monkey (Macaca mulatta) tissues

BackgroundThe rhesus monkey (Macaca mulatta) is a valuable and widely used model animal for biomedical research. However, quantitative analyses of rhesus gene expression profiles under diverse experimental conditions are limited by a shortage of suitable internal controls for the normalization of mRNA levels. In this study, we used a systematic approach for the selection of potential reference genes in the rhesus monkey and compared their suitability to that of the corresponding genes in humans.ResultsEight housekeeping genes (HKGs) (GAPDH, SDHA, ACTB, RPL13A, RPL32, UBA52, PGK1Y, and YWHAZ) from rhesus monkeys and humans were selected to test for normalization of expression levels in six different tissue types (brain, colon, kidney, liver, lung, and stomach). Their stability and suitability as reference genes were validated by geNorm, NormFinder and BestKeeper programs. Intriguingly, RPL13A and RPL32 were selected as ideal reference genes only in rhesus monkeys.ConclusionThe results clearly indicated the necessity of using different reference genes for normalization of expression levels between rhesus monkeys and humans in various tissues.

Molecular characterization of the DYX1C1 gene and its application as a cancer biomarker

PurposeDYX1C1 has three alternatively spliced transcripts. Therefore, we expect that alternative transcripts of DYX1C1 are used as a biomarker to detect specific cancer.MethodsRT-PCR analysis is conducted in order to detect expression of the DYX1C1 gene and the PCR products were analyzed using the Image J program to compare the expression levels of each transcript.ResultsWe found one of the transcripts was directly associated with an HERV-H LTR element that could be translated into protein sequence. Four new alternative transcripts were identified by RT-PCR analysis with various human tissue samples including 10 normal and adjacent tumor tissue sets. Semi-quantitative RT-PCR analysis showed the transcriptional activity of V3 and V2 was higher in tumor than in normal tissue samples, especially in the colorectal tissue samples.ConclusionOur results indicated that alternatively spliced transcript variants of the DYX1C1 gene could be used as cancer biomarkers to detect colorectal cancer.

The Whole-Genome and Transcriptome of the Manila Clam (Ruditapes philippinarum)

Abstract The manila clam, Ruditapes philippinarum, is an important bivalve species in worldwide aquaculture including Korea. The aquaculture production of R. philippinarum is under threat from diverse environmental factors including viruses, microorganisms, parasites, and water conditions with subsequently declining production. In spite of its importance as a marine resource, the reference genome of R. philippinarum for comprehensive genetic studies is largely unexplored. Here, we report the de novo whole-genome and transcriptome assembly of R. philippinarum across three different tissues (foot, gill, and adductor muscle), and provide the basic data for advanced studies in selective breeding and disease control in order to obtain successful aquaculture systems. An approximately 2.56 Gb high quality whole-genome was assembled with various library construction methods. A total of 108,034 protein coding gene models were predicted and repetitive elements including simple sequence repeats and noncoding RNAs were identified to further understanding of the genetic background of R. philippinarum for genomics-assisted breeding. Comparative analysis with the bivalve marine invertebrates uncover that the gene family related to complement C1q was enriched. Furthermore, we performed transcriptome analysis with three different tissues in order to support genome annotation and then identified 41,275 transcripts which were annotated. The R. philippinarum genome resource will markedly advance a wide range of potential genetic studies, a reference genome for comparative analysis of bivalve species and unraveling mechanisms of biological processes in molluscs. We believe that the R. philippinarum genome will serve as an initial platform for breeding better-quality clams using a genomic approach.

Quantitative analysis of alternative transcripts of human PCDH11X/Y genes

The Protocadherin 11X/Y (PCDH11X/Y) gene pair has been proposed as a carrier of the variation relating to cerebral asymmetry and psychosis on the ground that the Y gene was generated by duplication at 6 million years (close to the chimpanzee–human separation) and there is a case for an X/Y determinant of cerebral asymmetry. The present article investigated the patterns of alternative splicing and expression of the PCDH11X/Y genes. Twelve alternative transcripts of PCDH11X/Y genes were presently identified by in silico analysis. To investigate the biological roles of alternative transcripts of PCDH11X/Y genes, the transcripts were analyzed by real‐time reverse transcription‐polymerase chain reaction amplification. A total of 31 normal tissues including 11 different regions of human brain were used to investigate a wide spectrum of expression profiles. Dominant expression patterns were identified in several tissues (Tx1‐fetal liver; Tx3‐adult brain; Tx4‐adult brain and kidney; Tx5‐bone marrow; Ty1‐fetal brain; Ty2‐adrenal gland). Tx4 transcripts showed specific expression patterns in olfactory tissues. The findings can guide functional investigation of neuropsychiatric disorders.

Alternative Splicing and Its Impact as a Cancer Diagnostic Marker

Most genes are processed by alternative splicing for gene expression, resulting in the complexity of the transcriptome in eukaryotes. It allows a limited number of genes to encode various proteins with intricate functions. Alternative splicing is regulated by genetic mutations in cis-regulatory factors and epigenetic events. Furthermore, splicing events occur differently according to cell type, developmental stage, and various diseases, including cancer. Genome instability and flexible proteomes by alternative splicing could affect cancer cells to grow and survive, leading to metastasis. Cancer cells that are transformed by aberrant and uncontrolled mechanisms could produce alternative splicing to maintain and spread them continuously. Splicing variants in various cancers represent crucial roles for tumorigenesis. Taken together, the identification of alternative spliced variants as biomarkers to distinguish between normal and cancer cells could cast light on tumorigenesis.

Radiation-Induced Human Endogenous Retrovirus (HERV)-R env Gene Expression by Epigenetic Control

It is commonly accepted that ionizing radiation induces genomic instability by changes in genomic structure, epigenetic regulation and gene expression. Human endogenous retroviruses (HERV)-R also are often differentially expressed between normal and disease tissues under unstable genomic conditions and are implicated in the pathogenesis of several human diseases. To understand the influence of ionizing radiation on HERV-R expression, we performed quantitative reverse transcription-polymerase chain reaction (RT-PCR) analyses using γ-irradiated normal human cells. Compared to nonirradiated cells, HERV-R expression was up-regulated in γ-irradiated cells. The regulatory mechanism of HERV-R expression in irradiated cells was investigated by methylation analyses of HERV-R 5′LTRs and treatment with garcinol. These data indicated that the up-regulated transcription of HERV-R may be regulated by radiation-induced epigenetic changes induced by histone modification, and thus could be of great importance for understanding the relationship between radiation-induced biological effects and transposable elements.

Cooperative exonization of MaLR and AluJo elements contributed an alternative promoter and novel splice variants of RNF19

The RNF19 protein, which contains RING-finger and IBR motifs, acts as an E3 ubiquitin ligase localized to Lewy bodies. RNF19 is located on human chromosome 8q22.2, has a 4.4-kb transcript, and is ubiquitously expressed in various tissues. Here, we identified an alternative RNF19 promoter region and alternative RNF19 transcripts derived from MaLR (mammalian apparent LTR-retrotransposon) and AluJo elements. Comparative analyses indicated human-specific expression of the MaLR- and AluJo-related transcripts. From the expression analysis of 72 tissue samples including human normal, tumor, and primate tissues, three different isoforms (V1, V2, and V3) of MaLR-derived transcripts were identified. Quantitative RT-PCR analysis showed a dominant expression pattern of the V2 MaLR-derived transcript. A reporter gene assay for MaLR element promoter activity indicated that pGL2-RNF19/MaLR in the forward orientation is capable of driving luciferase gene expression in Cos7 and HCT116 cells. These findings suggest that RNF19 has acquired a new promoter and alternative exons via continuous retrotransposition events of MaLR and AluJo elements during mammalian and primate evolution, respectively.

Alu-related transcript of TJP2 gene as a marker for colorectal cancer.

Identifying diagnostic markers is very important in cancer biology. Transposable elements (TEs) account for 45% of the human genome and are usually silent in normal tissues. Contrary to this, in cancer tissues, TEs have been found to be expressed specifically with genomic instability. Here, we investigated the transcripts related to TEs using bioinformatic tools and RT-PCR analysis in tumor and adjacent normal tissues as well as in cancer cell lines. From this analysis, we identified the Alu-related transcript of TJP2 gene (TJP2-Alu transcript) that was differentially expressed between tumor and normal tissues. Its expression was higher in colon cancer cell lines and tumor tissues of colorectal cancer patients. These results could contribute to a better understanding of the involvement of TEs in human colorectal cancer, and to evaluating their potential as diagnostic markers for colorectal cancer.

Ionic conduction and structural phase transitions of Na2SO4 doped with various impurities

Abstract The metastable room-temperature phase III of Na 2 SO 4 crystal transforms from III→I on heating and I→II→III on cooling. Phase II exists in a narrow temperature interval (less than 7°C). To obtain better microscopic understandings about the existence and nature of the intermediate phase II, electrical conductivity and Raman spectrum were measured for mixed crystals of Na 2 (SO 4 ) 1− x (SeO 4 ) x and Na 2(1− x ) Ag 2 x SO 4 with x ≤0.1. On cooling, the intermediate phase II is stabilized in a wider temperature interval with the incorporation of guest ions. In the case of Na 1.8 Ag 0.2 SO 4 , phase II persists as wide as ∼59°C. On heating, phase II, which is otherwise forbidden on heating in pure Na 2 SO 4 crystal, reappears when x is large.

Chimpanzee-specific endogenous retrovirus generates genomic variations in the chimpanzee genome.

Endogenous retroviruses (ERVs), eukaryotic transposable elements, exist as proviruses in vertebrates including primates and contribute to genomic changes during the evolution of their host genomes. Many studies about ERVs have focused on the elements residing in the human genome but only a few studies have focused on the elements which exist in non-human primate genomes. In this study, we identified 256 chimpanzee-specific endogenous retrovirus copies (PtERVs: Pan troglodyte endogenous retroviruses) from the chimpanzee reference genome sequence through comparative genomics. Among the chimpanzee-specific ERV copies, 121 were full-length chimpanzee-specific ERV elements while 110 were chimpanzee-specific solitary LTR copies. In addition, we found eight potential retrotransposition-competent full-length chimpanzee-specific ERV copies containing an intact env gene, and two of them were polymorphic in chimpanzee individuals. Through computational analysis and manual inspection, we found that some of the chimpanzee-specific ERVs have propagated via non-classical PtERV insertion (NCPI), and at least one of the PtERVs may have played a role in creating an alternative transcript of a chimpanzee gene. Based on our findings in this study, we state that the chimpanzee-specific ERV element is one of the sources of chimpanzee genomic variations, some of which might be related to the alternative transcripts in the chimpanzee population.