Similar articles

Abstract: Objective: To analyze the genomic structure of SNC6, a progesterone-receptor associated protein gene and its regulatory elements in its 5'-flanking region. Methods: Genomic sequence from GenBank database (accession number: Z98048) covering the whole SNC6 gene was used to analyze the genomic structure of SNC6 and design primers for PCR amplification of its 5'-flanking region. A 1894 bp fragment of the 5'-flanking region (−1814 to +75) was cloned by PCR using genomic DNA from a healthy donor peripheral blood lymphocyte as template. This fragment, as well as 3 shorter derivative fragments (1423 bp, 632 bp and 416 bp, which correspond to −1344 to +75, −552 to +75 and −337 to +75 respectively), were subcloned into pGL2 series luciferase reporter vectors. These constructs were introduced into colorectal cancer cell line SW620 for transient expression of reporter gene and luciferase activities were measured. Results: The genomic structure analysis showed there are 12 exons for SNC6 gene, which spans 32017 bp (nt71529 to nt39513 in Z98048 sequence). All transfected SW620 cells with the above 5-flanking region-containing constructs showed luciferase activities. The highest luciferase activities were measured in transfected cells with vectors containing 1894 bp fragments, and the lowest luciferase activities were measured in transfected cells with vectors containing 416 bp fragments. Luciferase activities were higher in transfected cells with vectors containing 632 bp fragments than that in transfected cells with vectors containing 1423 bp fragments. Conclusion: The basic transcription-promoting element (promoter) for SNC6 expression resides between 0 to −337, and two transcription-enhancing elements (enhancer) resides between −337 to −552 and −1344 to −1814, whereas one transcription-inhibiting element (silencer) exists between −552 to −1344.

[1] Cai,X., Zhang,Y., Geng,L., et al., 1997. Assignment of a novel colorectal cancer-asociated gene HSU17714 gene to human chromosome band 22q13 by in situ hybridization. Chin. J. Oncol., 19(3):177-179(in Chinese, with English abstract).

[2] Cao,J., Cai,X., Zheng,L., et al., 1997. Characterization of colorectal-cancer-related cDNA clones obtained by subtractive hybridization screening. J. Cancer Res. Clin. Oncol., 123:447-451.

[3] DeRisi,J., Penland,L., Brown,P.O., et al., 1996. Use of a cDNA microarray to analyse gene expression pattern in human cancer. Nat. Genet., 14(4):457-460.

[4] Geng,L., Cai,X., Cao,J., et al., 1999. Cloning the 5'-end fragment of ST13 cDNA by nested PCR. Chin. J. Med. Genet., 16(3):174-176.

[5] Kurian,K.M., Watson,C.J., Wyllie,A.H., et al., 1999. DNA chip technology. J. Pathol., 187(3):267-271.

[6] Liang,P., Pardee,A.B., 1992. Diffenential display of eukaryotic messenger RNA by means of the polymerase chain reaction. Science., 257(5072):967-971.

[7] Mo,Y., Zheng,S., Shen,D., et al., 1996. Differential expression of HSU17714 gene in colorectal cancer and normal colonic mucosa. Chin. J. Oncol., 18(4):241-243(in Chinese, with English abstract).

[8] Prapapanich,V., Chen,S., Nair,S.C., et al., 1996. Molecular cloning of human p48, a transient component of progesterone receptor complexes and an hsp70-binding protein. Mol. Endocrinol., 10:420-431.

[9] Sagerstrom,C.G., Sun,B.I., Sive,H.L., 1997. Subtractive cloning: past, present, and future. Annu. Rev. Biochem., 66:751-783.

[10] Velculescu,V.E., Zhang,L., Volgelstein, B., et al., 1995. Serial analysis of gene expression. Science., 270(5235):484-487.

[11] Zheng,S., Cai,X., Cao,J., et al., 1997. Screening and identification of down-regulated genes in colorectal carcinoma by subtractive hybridization: a method to identify putative tumor suppressor genes. Chin. Med. J., 110(7):543-547.

[12] Zheng,S., Cai,X., Cao,J., et al., 1997. Application of subtractive hybridization in screening for colorectal cancer negatively related genes. Natl. Med. J. Chin., 77(4):256-259(in Chinese, with English abstract).