Transcript 슬라이드 1
Promoter activity of LTR element of the human FPRL2 gene Hong-seok HA, Jae-Won HUH, Dae-Soo KIM1, Heui-Soo KIM * Division of Biological Sciences, College of Natural Sciences, Pusan National University. 1PBBRC, Interdisciplinary Research Program of Bioinformatics, College of Natural Sciences, Pusan National University. HTTP://WWW.PRIMATE.OR.KR ABSTRACT INTRODUCTION LTR: Long Terminal Repeat The human genome is estimated to consist of approximately 8% human endogenous retroviruses (HERVs) and related sequences. FPRL2 (fomyl peptide receptor-like 2) gene has a solitary LTR (long terminal repeat). The LTR is located between first exon and promoter region of the FPRL2 gene. The FPRL2 gene containing LTR element was expressed in various human tissues except fetal brain and cerebellum. The LTR element was detected in hominoid, Old World monkeys, and New World monkeys except for common marmoset, whereas LINE (long interspersed repetitive element) and SINE (short interspersed repetitive element) elements were detected in prosimian (ring-tailed lemur) and common marmoset. We also examined promoter activity of the LTR element in FPRL2 gene, and discussed its biological role. Taken together, the insertion of retroelements into primate genome could have different biological roles during primate evolution. . MATERIALS & METHODS Bioinformatics Luciferase Assay Transfac 6.0 Real Time RT-PCR Bisulfite Sequencing PCR Genomic DNA PCR & Gene cloning LINE: Long Integrated nuclear element SINE: Short Integrated nuclear element SINE 13% Other region 16% Retroelement HERV: Human Endogenous Retro-Virus RNA intermediate LINE 20% - LTR element + LTR element Retroposon - env Gene-related Sequence 36% HERV element 8% Retrotransposon - RT DNA element 3% Pseudogene Coding sequence 1% + RT LTR SINE ORF1 ORF2 LTR Yeast Ty1/copia/truncated HERVs 3% LTR P LTR Human THE1 Poly(A) Human Alu Retroelements have been subjected to many amplification and transposition events resulting in a widespread distribution of complete or partial retroviral sequences throughout the human genome. The human genome comprises approximately 8% of the human endogenous retroviruses (HERVs) and other long terminal repeat (LTR)–like elements . Most HERVs seem to have entered the genome between 10 and 50 million years ago, and they comprise over 200 distinct groups and subgroups . Expression of retroelements can influence the outcome of infections in different ways that can be either beneficial or detrimental to the host. A function of the multiple copy families, scattered throughout the genome, has been reported regulatory functions on the gene expression of nearby located genes . A small minority of such sequences has acquired a role in regulating gene expression, and some of these may be related to differences between individuals, and to expression of disease. . + env Retrovirus LINE LTR P ORF1 ORF2 L1 Poly(A) gag pol env LTR Full-length HERVs/exogenous retrovirus REFERENCES 1. Maksakova IA, Mager DL. 2005. Transcriptional regulation of early transposon elements, an active family of mouse long terminal repeat re t rot ra nsposons . J Vi ro . . 2. Yang D, Chen Q, Gertz B, He R, Phulsuksombati M, Ye RD, Oppenheim JJ. 2002. Human dendritic cells express functional formyl peptide receptor-like-2 (FPRL2) throughout maturation. J Leukoc Biol. . RESULTS & DISCUSSION Fig. 1. The genomic structure of FPRL2 gene including LTR54 element. Exons were represented by solid box with the exon numbers. Arrows indicate the primer location. The LTR54 element was integrated into the FPRL2 gene with the antisense orientation on human chromosome 19q13.41 Fig. 4. PCR analysis for the presence of LTR54-derived promoter region of FPRL2 gene using the various primate genomic DNAs (A). Primates showed PCR products that were cloned and sequenced (B,C). Fig. 2. Real Time RT-PCR analysis of LTR54 derived transcript from different human tissues. Fig. 3. Luciferase reporter gene assay for LTR54-derived promoter of FPRL2 gene in transient transfected HCT116. Relative activity of luciferase assay for pGL2-hFPRL2-LTR54 in forward and reverse orientation or the pGL2 basic vector was indicated as schematic diagram. Results are expressed as ratios of the luciferase activity to that of the promoterless pGL2 reporter plasmid. Fig 5. Comparison of FPRL2 gene family in silico analysis; gene location (A), comparison of mRNA structure using PIP maker program(B) , alignment of amino acid (C)