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The interrelationship between DRIM gene expression and cytogenetic and phenotypic characteristics in human breast tumor cell lines.

Goodison S, Viars C, Grazzini M, Urquidi V - BMC Genomics (2003)

Bottom Line: DRIM encodes a protein comprising 2785 amino acids with significant homology to a protein in yeast and C. elegans, but little else is currently known about its function or pattern of expression.We confirmed higher expression of DRIM mRNA in the non-metastatic MDA-MB-435 clone NM2C5, relative to its metastatic counterpart M4A4, but this appeared to be due to the presence of an extra copy of the DRIM gene in the cell line's genome.The other non-metastatic cell lines in the series (T47D MCF-7, SK-BR-3 and ZR-75-1) contained either 3 or 4 chromosomal copies of DRIM gene.

View Article: PubMed Central - HTML - PubMed

Affiliation: UCSD Cancer Center and Department of Pathology, University of California, San Diego, La Jolla, CA, USA. sgoodison@ucsd.edu

ABSTRACT

Background: In order to facilitate the identification of genes involved in the metastatic phenotype we have previously developed a pair of cell lines from the human breast carcinoma cell line MDA-MB-435, which have diametrically opposite metastatic potential in athymic mice. Differential display analysis of this model previously identified a novel gene, DRIM (down regulated in metastasis), the decreased expression of which correlated with metastatic capability. DRIM encodes a protein comprising 2785 amino acids with significant homology to a protein in yeast and C. elegans, but little else is currently known about its function or pattern of expression. In a detailed analysis of the DRIM gene locus we quantitatively evaluated gene dosage and the expression of DRIM transcripts in a panel of breast cell lines of known metastatic phenotype.

Results: Fluorescent in situ hybridization (FISH) analyses mapped a single DRIM gene locus to human chromosome 12q23~24, a region of conserved synteny to mouse chromosome 10. We confirmed higher expression of DRIM mRNA in the non-metastatic MDA-MB-435 clone NM2C5, relative to its metastatic counterpart M4A4, but this appeared to be due to the presence of an extra copy of the DRIM gene in the cell line's genome. The other non-metastatic cell lines in the series (T47D MCF-7, SK-BR-3 and ZR-75-1) contained either 3 or 4 chromosomal copies of DRIM gene. However, the expression level of DRIM mRNA in M4A4 was found to be 2-4 fold higher than in unrelated breast cells of non-metastatic phenotype.

Conclusions: Whilst DRIM expression is decreased in metastatic M4A4 cells relative to its non-metastatic isogenic counterpart, neither DRIM gene dosage nor DRIM mRNA levels correlated with metastatic propensity in a series of human breast tumor cell lines examined. Collectively, these findings indicate that the expression pattern of the DRIM gene in relation to the pathogenesis of breast tumor metastasis is more complex than previously recognized.

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Spectral karyotyping (SKY) classification of representative metaphases of MDA-MB-435 subclones M4A4 (upper panel) and NM2C5 (lower panel). Chromosome material involved in translocations are indicated on the figure in some cases. Each cell line has 2 normal chromosome 12's. NM2C5 cells contain a marker chromosome consisting of material from chromosomes 12 and 15 (see chromosome 15 box, lower panel).
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Figure 3: Spectral karyotyping (SKY) classification of representative metaphases of MDA-MB-435 subclones M4A4 (upper panel) and NM2C5 (lower panel). Chromosome material involved in translocations are indicated on the figure in some cases. Each cell line has 2 normal chromosome 12's. NM2C5 cells contain a marker chromosome consisting of material from chromosomes 12 and 15 (see chromosome 15 box, lower panel).

Mentions: The DRIM gene was discovered through differential display analysis of the MDA-MB-435 subclones M4A4 and NM2C5 [4]. In order to investigate whether gene copy number was a factor in this differential expression, the karyotypes of these two cell lines were compared using G-banding in combination with Spectral karyotyping (SKY) analyses. In particular, the complement and the integrity of chromosome 12 material was evaluated. Spectral karyotyping [6] is a multi-color FISH procedure capable of classifying all chromosomes in a single hybridization experiment. Using this technique it was revealed that both the metastatic M4A4, and the non-metastatic NM2C5 cell lines are hyperdiploid (modal number of 56–57) and contain multiple chromosomal aberrations (Figure 3). A number of signature markers were common to both lines (Figure 3) and were present in all metaphase spreads analyzed for each cell line. Detailed comparisons of multiple images allowed clarification of almost all marker chromosomes in these two cell lines. Both cell lines contained 2 copies of apparently normal chromosome 12. However, one specific chromosomal rearrangement, found only in the non-metastatic cell line NM2C5, consisted of a non-reciprocal translocation involving chromosome 12 material (Figure 2). This (12;15) translocation occurred in 100% of NM2C5 cells examined, but was never present in M4A4. Confirmatory FISH and G-banding evaluations identified this translocation as t(12;15)(q22;q26.1). No other chromosomal rearrangements were found to be specific to the NM2C5 cell line. FISH analysis revealed that the extra chromosome 12 material present in NM2C5 cells was composed of 12q sequences including a third copy of the DRIM locus (Figure 3). Further analysis revealed that in 40% of M4A4 cells there was a deletion in the p-arm of one chromosome 12. However, this would not affect the DRIM gene located on the q-arm, which appeared normal in all cases. DRIM-specific FISH analyses confirmed that M4A4 cells contain only 2 DRIM loci located on 2 normal chromosomes 12 (Figure 4).


The interrelationship between DRIM gene expression and cytogenetic and phenotypic characteristics in human breast tumor cell lines.

Goodison S, Viars C, Grazzini M, Urquidi V - BMC Genomics (2003)

Spectral karyotyping (SKY) classification of representative metaphases of MDA-MB-435 subclones M4A4 (upper panel) and NM2C5 (lower panel). Chromosome material involved in translocations are indicated on the figure in some cases. Each cell line has 2 normal chromosome 12's. NM2C5 cells contain a marker chromosome consisting of material from chromosomes 12 and 15 (see chromosome 15 box, lower panel).
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Related In: Results  -  Collection

Show All Figures
getmorefigures.php?uid=PMC222913&req=5

Figure 3: Spectral karyotyping (SKY) classification of representative metaphases of MDA-MB-435 subclones M4A4 (upper panel) and NM2C5 (lower panel). Chromosome material involved in translocations are indicated on the figure in some cases. Each cell line has 2 normal chromosome 12's. NM2C5 cells contain a marker chromosome consisting of material from chromosomes 12 and 15 (see chromosome 15 box, lower panel).
Mentions: The DRIM gene was discovered through differential display analysis of the MDA-MB-435 subclones M4A4 and NM2C5 [4]. In order to investigate whether gene copy number was a factor in this differential expression, the karyotypes of these two cell lines were compared using G-banding in combination with Spectral karyotyping (SKY) analyses. In particular, the complement and the integrity of chromosome 12 material was evaluated. Spectral karyotyping [6] is a multi-color FISH procedure capable of classifying all chromosomes in a single hybridization experiment. Using this technique it was revealed that both the metastatic M4A4, and the non-metastatic NM2C5 cell lines are hyperdiploid (modal number of 56–57) and contain multiple chromosomal aberrations (Figure 3). A number of signature markers were common to both lines (Figure 3) and were present in all metaphase spreads analyzed for each cell line. Detailed comparisons of multiple images allowed clarification of almost all marker chromosomes in these two cell lines. Both cell lines contained 2 copies of apparently normal chromosome 12. However, one specific chromosomal rearrangement, found only in the non-metastatic cell line NM2C5, consisted of a non-reciprocal translocation involving chromosome 12 material (Figure 2). This (12;15) translocation occurred in 100% of NM2C5 cells examined, but was never present in M4A4. Confirmatory FISH and G-banding evaluations identified this translocation as t(12;15)(q22;q26.1). No other chromosomal rearrangements were found to be specific to the NM2C5 cell line. FISH analysis revealed that the extra chromosome 12 material present in NM2C5 cells was composed of 12q sequences including a third copy of the DRIM locus (Figure 3). Further analysis revealed that in 40% of M4A4 cells there was a deletion in the p-arm of one chromosome 12. However, this would not affect the DRIM gene located on the q-arm, which appeared normal in all cases. DRIM-specific FISH analyses confirmed that M4A4 cells contain only 2 DRIM loci located on 2 normal chromosomes 12 (Figure 4).

Bottom Line: DRIM encodes a protein comprising 2785 amino acids with significant homology to a protein in yeast and C. elegans, but little else is currently known about its function or pattern of expression.We confirmed higher expression of DRIM mRNA in the non-metastatic MDA-MB-435 clone NM2C5, relative to its metastatic counterpart M4A4, but this appeared to be due to the presence of an extra copy of the DRIM gene in the cell line's genome.The other non-metastatic cell lines in the series (T47D MCF-7, SK-BR-3 and ZR-75-1) contained either 3 or 4 chromosomal copies of DRIM gene.

View Article: PubMed Central - HTML - PubMed

Affiliation: UCSD Cancer Center and Department of Pathology, University of California, San Diego, La Jolla, CA, USA. sgoodison@ucsd.edu

ABSTRACT

Background: In order to facilitate the identification of genes involved in the metastatic phenotype we have previously developed a pair of cell lines from the human breast carcinoma cell line MDA-MB-435, which have diametrically opposite metastatic potential in athymic mice. Differential display analysis of this model previously identified a novel gene, DRIM (down regulated in metastasis), the decreased expression of which correlated with metastatic capability. DRIM encodes a protein comprising 2785 amino acids with significant homology to a protein in yeast and C. elegans, but little else is currently known about its function or pattern of expression. In a detailed analysis of the DRIM gene locus we quantitatively evaluated gene dosage and the expression of DRIM transcripts in a panel of breast cell lines of known metastatic phenotype.

Results: Fluorescent in situ hybridization (FISH) analyses mapped a single DRIM gene locus to human chromosome 12q23~24, a region of conserved synteny to mouse chromosome 10. We confirmed higher expression of DRIM mRNA in the non-metastatic MDA-MB-435 clone NM2C5, relative to its metastatic counterpart M4A4, but this appeared to be due to the presence of an extra copy of the DRIM gene in the cell line's genome. The other non-metastatic cell lines in the series (T47D MCF-7, SK-BR-3 and ZR-75-1) contained either 3 or 4 chromosomal copies of DRIM gene. However, the expression level of DRIM mRNA in M4A4 was found to be 2-4 fold higher than in unrelated breast cells of non-metastatic phenotype.

Conclusions: Whilst DRIM expression is decreased in metastatic M4A4 cells relative to its non-metastatic isogenic counterpart, neither DRIM gene dosage nor DRIM mRNA levels correlated with metastatic propensity in a series of human breast tumor cell lines examined. Collectively, these findings indicate that the expression pattern of the DRIM gene in relation to the pathogenesis of breast tumor metastasis is more complex than previously recognized.

Show MeSH
Related in: MedlinePlus