Alu-PCR Cloning of Genes Containing DNase I Hypersensitive site: Characterization of Differential Expression of Siglec-11 in Human Tumor Cells

• Main Author: 王碧玲
• Other Authors: Ming-ta Hsu
• Format: Others
• Language: zh-TW
• Published: 2003
• Online Access: http://ndltd.ncl.edu.tw/handle/87218356916077555733

碩士 === 國立陽明大學 === 生物化學研究所 === 91 === Our lab has previously identified nine novel DNase I hypersensitive clones using Inter-Alu PCR technology. In this work, I focused on the clone Hp-1-1.5, which contains a DNase I hypersensitive site in U937 cell line. It is located within the Intron 8 of a newly identified gene, Siglec-11. This gene is one of the Siglec family reported last year by Angata et al. Siglecs are sialic-acid-binding immnuglobulin-like lectins involved in cell-cell interactions and signaling functions in the haemopoietic, immune, nerve systems. Siglec-11 is expressed in macrophages in various human tissues. Siglec-11 interacts with protein-tyrosine phosphatase SHP-1 and SHP-2 upon tyrosine phosphorylation. Thus Siglec-11 may be involoved in regulating cells activation during immune and inflammatory response. In this work, we analyzed the relationship between DNase I hypersensitivity and Siglec-11 gene expression. To address whether Siglec-11 can be regulated by the hypersensitive site in Intron 8, we analyzed the DNaseI hypersensitivity pattern in intron 8 by specific PCR in U937 cells treated with PMA. DHS (DNase I hypersensitive site) in intron 8 lost its DNaseI hypersensitivity together with the expression of Siglec-11 upon induction of U937 differentiation by tratment with PMA. Siglec-11 was found to be expressed in most human tissues and differentiated cells, but not in tumor cell lines. We also noticed a new novel gene, Siglec-H, adjacent to Sigelc-11 but in opposite orientation has partial sequence homology to Siglec-11. RT-PCR analysis showed that Siglec-H was expressed in most tumor cell lines but not normal cells and differentiated cells. These data suggest that the expression of Siglec-H seems to be inversely correlated with that of Siglec-11. In conclusion, we showed that the expression of Siglec-11 is inversely correlated with the nuclease hypersensentivity in intron 8. This result suggests that the sequence in Intron 8 contains a silencer element. In addition, the expression of Siglec-H is mutually exclusive with Siglec-11. Together, theses two genes may serve as tumor markers.