Identification of a tumor suppressor gene using chromosome engineering

Many tumor suppressor genes are involved in the multistep process of neoplastic development. However, the signaling mechanism that underlies the development of tumors has not yet been completely elucidated. Therefore, Discovery of a novel tumor suppressor gene plays a crucial role in our understanding of the development and progression of malignant tumors. Chromosome engineering technique that base on Microcell-mediated chromosome transfer （MMCT），which can be generally used to the introduction of a single chromosome to a variety of tumor cells, is one of effective approach for mapping and identification of tumor suppressor genes. We have identified paired-like homeodomain 1（PITX1）gene as a novel telomerase negative regulatory factor that inhibit the expression of telomerase reverse transcriptase （TERT）using chromosome engineering technique. Here, we describe a unique strategy from mapping to identification of tumor suppressor gene by using MMCT approach

長鎖非コードRNA による染色体ドメインレベルの遺伝子発現制御

　ヒトゲノムの解読により, タンパクをコードするmRNAは全RNAのわずか1.5％に過ぎず, 残りの98％はタンパクをコードしていない非コードRNA(non-coding RNA: ncRNA)で占めていることが明らかとなった. それに加え, 近年, ENCODEプロジェクトにおいて, 長鎖非コードRNA(long non-coding RNA: lncRNA)は, ヒトの〜9000ものゲノム領域から転写されていた1). これまでlncRNAには, がん抑制遺伝子Ink4遺伝子座の発現制御に関わるANRIL, ゲノムインプリンティングに関わるKCNQ1OT1/LIT1, Air, HOX遺伝子の転写抑制に関わるHOTAIRなどが知られている. 最近では, 代謝の再プログラミング化に関与するlncRNAとしてlincRNA-p21が見出された. このように, lncRNAもまた機能性RNAとして様々な生体制御機構に重要な役割を担っている.　本稿では, lncRNAの中でもクロマチン上に集積(コーティング)することで染色体全体あるいは, 局所的にヘテロクロマチン化を誘導するユニークな遺伝子発現制御に関わるXistおよびKCNQ1OT1/LIT1に着目し, 最新の知見から遺伝子発現制御や疾患との関わりについて考察する

Regulation of functional KCNQ1OT1 lncRNA by β-catenin.

Long noncoding RNAs (lncRNAs) have been implicated in many biological processes through epigenetic mechanisms. We previously reported that KCNQ1OT1, an imprinted antisense lncRNA in the human KCNQ1 locus on chromosome 11p15.5, is involved in cis-limited silencing within an imprinted KCNQ1 cluster. Furthermore, aberration of KCNQ1OT1 transcription was observed with a high frequency in colorectal cancers. However, the molecular mechanism of the transcriptional regulation and the functional role of KCNQ1OT1 in colorectal cancer remain unclear. Here, we show that the KCNQ1OT1 transcriptional level was significantly increased in human colorectal cancer cells in which β-catenin was excessively accumulated in the nucleus. Additionally, overexpression of β-catenin resulted in an increase in KCNQ1OT1 lncRNA-coated territory. On the other hand, knockdown of β-catenin resulted in significant decrease of KCNQ1OT1 lncRNA-coated territory and an increase in the mRNA expression of the SLC22A18 and PHLDA2 genes that are regulated by KCNQ1OT1. We showed that β-catenin can promote KCNQ1OT1 transcription through direct binding to the KCNQ1OT1 promoter. Our evidence indicates that β-catenin signaling may contribute to development of colorectal cancer by functioning as a novel lncRNA regulatory factor via direct targeting of KCNQ1OT1

Localization of an hTERT repressor region on human chromosome 3p21.3 using chromosome engineering

Telomerase is a ribonucleoprotein enzyme that synthesizes telomeric DNA. The reactivation of telomerase activity by aberrant upregulation/expression of its catalytic subunit hTERT is a major pathway in human tumorigenesis. However, regulatory mechanisms that control hTERT expression are largely unknown. Previously, we and others have demonstrated that the introduction of human chromosome 3, via microcell-mediated chromosome transfer (MMCT), repressed transcription of the hTERT gene. These results suggested that human chromosome 3 contains a regulatory factor(s) involved in the repression of hTERT. To further localize this putative hTERT repressor(s), we have developed a unique experimental approach by introducing various truncated chromosome 3 regions produced by a novel chromosomal engineering technology into the renal cell carcinoma cell line (RCC23 cells). These cells autonomously express ectopic hTERT (exohTERT) promoted by a retroviral LTR promoter in order to permit cellular division after repression of endogenous hTERT. We found a telomerase repressor region located within a 7-Mb interval on chromosome 3p21.3. These results provide important information regarding hTERT regulation and a unique method to identify hTERT repressor elements

黒色腫細胞において、miR-19bはPITX1 mRNAを標的としてhTERT mRNAの発現を制御する

Purpose: This article sheds light onto the increasing problem of product returns, which is exacerbated by growing e-commerce. Many retailers and academics are oblivious to the nature and scale of this challenge. Interdisciplinary research is needed to develop supporting theory, and cross-functional teams are required to implement measures addressing economic, ecological and social sustainability issues. Design/methodology/approach: The initial project adopted a multi-case study approach, whereby returns processes were mapped, vulnerabilities identified and a returns cost calculator was developed. Findings: Product returns processes are usually complicated, prone to internal and external fraud, inefficient and lack sustainability. They can generate considerable losses to the business, especially as returns data are often not systematically collected, monitored or reported to senior management. There are important implications for strategic and operational management, namely the need to develop a concept for Lean returns systems. Originality/value: Product returns are a unique and understudied but growing field in academic research, with only few publications over the last two decades. Yet the phenomenon is causing increasing problems in business and society. Robust solutions could achieve great financial and non-financial impacts.</p

Possible Relationship Between MYBL1 Alterations and Specific Primary Sites in Adenoid Cystic Carcinoma: A Clinicopathological and Molecular Study of 36 Cases

[Background] Adenoid cystic carcinoma (ACC) is a relatively rare malignant neoplasm that occurs in salivary glands and various other organs. Recent studies have revealed that a significant proportion of ACCs harbor gene alterations involving MYB or MYBL1 (mostly fusions with NFIB) in a mutually-exclusive manner. However, its clinical significance remains to be well-established. [Methods] We investigated clinicopathological and molecular features of 36 ACCs with special emphasis on the significance of MYBL1 alterations. Reverse-transcription polymerase-chain reaction (RT-PCR) and fluorescence in-situ hybridization (FISH) were performed to detect MYB/MYBL1-NFIB fusions and MYBL1 alterations, respectively. Immunohistochemistry was performed to evaluate MYB expression in the tumors. The results were correlated with clinicopathological profiles of the patients. [Results] RT-PCR revealed MYB-NFIB and MYBL1-NFIB fusions in 10 (27.8%) and 7 (19.4%) ACCs, respectively, in a mutually-exclusive manner. FISH for MYBL1 rearrangements was successfully performed in 11 cases, and the results were concordant with those of RT-PCR. Immunohistochemically, strong MYB expression was observed in 23 (63.9%) tumors, none of which showed MYBL1 alterations. Clinicopathologically, a trend of a better disease-specific survival was noted in patients with MYBL1 alterations than in those with MYB-NFIB fusions and/or strong MYB expression; however, the difference was not significant. Interestingly, we found tumors with MYBL1 alterations significantly frequently occurred in the mandibular regions (P = 0.012). Moreover, literature review revealed a similar tendency in a previous study. [Conclusion] Our results suggest that there are some biological or etiological differences between ACCs with MYB and MYBL1 alterations. Moreover, the frequent occurrence of MYBL1-associated ACC in the mandibular regions suggests that MYB immunohistochemistry is less useful in diagnosing ACCs arising in these regions. Further studies are warranted to verify our findings

Dysregulation of Gene Expression in the Artificial Human Trisomy Cells of Chromosome 8 Associated with Transformed Cell Phenotypes

A change in chromosome number, known as aneuploidy, is a common characteristic of cancer. Aneuploidy disrupts gene expression in human cancer cells and immortalized human epithelial cells, but not in normal human cells. However, the relationship between aneuploidy and cancer remains unclear. To study the effects of aneuploidy in normal human cells, we generated artificial cells of human primary fibroblast having three chromosome 8 (trisomy 8 cells) by using microcell-mediated chromosome transfer technique. In addition to decreased proliferation, the trisomy 8 cells lost contact inhibition and reproliferated after exhibiting senescence-like characteristics that are typical of transformed cells. Furthermore, the trisomy 8 cells exhibited chromosome instability, and the overall gene expression profile based on microarray analyses was significantly different from that of diploid human primary fibroblasts. Our data suggest that aneuploidy, even a single chromosome gain, can be introduced into normal human cells and causes, in some cases, a partial cancer phenotype due to a disruption in overall gene expression

Studies of Tumor Suppressor Genes via Chromosome Engineering

The development and progression of malignant tumors likely result from consecutive accumulation of genetic alterations, including dysfunctional tumor suppressor genes. However, the signaling mechanisms that underlie the development of tumors have not yet been completely elucidated. Discovery of novel tumor-related genes plays a crucial role in our understanding of the development and progression of malignant tumors. Chromosome engineering technology based on microcell-mediated chromosome transfer (MMCT) is an effective approach for identification of tumor suppressor genes. The studies have revealed at least five tumor suppression effects. The discovery of novel tumor suppressor genes provide greater understanding of the complex signaling pathways that underlie the development and progression of malignant tumors. These advances are being exploited to develop targeted drugs and new biological therapies for cancer