Normal human chromosome 11 suppresses tumorigenicity of human cervical tumor cell line SiHa

Abstract

We examined the ability of human chromosome 11 derived from normal fibroblast cells to suppress the tumorigenicity of SiHa cells, a human cervical tumor cell line. Using DNA transfection, the human chromosome was tagged with a selectable marker (the pSV2neo gene, which encodes resistance to the antibiotic G418), transferred to mouse A9 cells by cell hybridization and microcell transfer techniques, and then transferred to SiHa cells by microcell transfer. These procedures resulted in the appearance of 15 independent, G418-resistant clones, 5 of which had one or two extra copies of an intact human chromosome 11. In situ chromosomal hybridization of these clones with the pSV2neo plasmid revealed the presence of a neo-tagged human chromosome 11 in all of the five SiHa-microcell hybrids. Two SiHa-microcell hybrids that contained a single copy of neo-tagged human chromosome 12 were also isolated by the same methods. The tumorigenicities of SiHa clones with one or two extra copies of chromosome 11 (SiHa-11) were suppressed; four of the five SiHa-11 clones formed no tumors in nude mice, whereas both parental SiHa cells and SiHa cells with an extra chromosome 12 formed tumors within 30 d. One SiHa-11 cell clone formed a single tumor 90 d after injection. This rare tumor had lost one copy of chromosome 11 and rapidly formed tumors when reinjected. These results indicate that the introduction of a single copy of normal human chromosome 11, but not chromosome 12, suppresses the tumorigenicity of SiHa cells, indicating the presence on human chromosome 11 of a putative tumor-suppressor gene (or genes) for human cervical tumors.

Aberrant promoter methylation and silencing of the POU2F3 gene in cervical cancer

“…However, chromosome complementation experiments suggest that alteration of additional tumor suppressor proteins likely contributes to cervical carcinogenesis. Microcell transfer of a partial copy of chromosome 11 suppresses tumorigenicity in CC cell lines (Saxon et al, 1986;Koi et al, 1989). POU2F3 lies within a significant region of LOH at 11q23.3 in CC, and is therefore a candidate tumor suppressor gene that could be inactivated by both chromosomal loss (Rader et al, 1996;Huettner et al, 1998;O'Sullivan et al, 2001;Zhang et al, 2005) and aberrant methylation of its promoter.…”

Epigenetic reprogramming of the human H19 gene in mouse embryonic cells does not erase the primary parental imprint

“…A9 hybrids containing a paternal or maternal human chromosome 11 tagged with pSV2bsr were constructed as described previously (Koi et al 1989) and were maintained with selective medium containing 3 mg/mL of blasticidin S (BS). Mouse m5S cells, an immortalized near-diploid cell line (Sasaki & Kodama 1987), were kindly provided by Dr Masao M. Sasaki and were maintained in alpha minimum essential medium (MEM, Gibco) supplemented with 10% foetal calf serum (FCS).…”

“…Introduction of human chromosome 11 into mouse recipient cells was performed via microcell-mediated chromosome transfer (Koi et al 1989). Microcells purified from 1 × 10 8 donor A9 cells containing a paternal or maternal human chromosome 11 were fused with m5S, P19 and OTF9-63 cells using 47% polyethylene glycol (PEG) (M r 1000, Baker).…”

“…Microcell-hybrid clones were selected with 3 mg/mL of BS for 2-3 weeks. Cell-cell hybridization was performed essentially as described previously (Koi et al 1989). In brief, equal numbers of BS-resistant OTF9 hybrids and G418-resistant A9 cells were mixed and incubated for 1 day.…”

Inactivation of CHEK1 and EI24 is associated with the development of invasive cervical carcinoma: Clinical and prognostic implications

“…studies in SiHa and HeLa cell lines 13,14 suggest the presence of putative TSGs at this locus. However, the deleted regions are quite broad, and there is ambiguity among the investigators regarding localization of consensus deleted regions.…”