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Securin is not required for chromosomal stability in human cells.

Pfleghaar K, Heubes S, Cox J, Stemmann O, Speicher MR - PLoS Biol. (2005)

Bottom Line: Here we show that, after hSecurin knockout through homologous recombination, chromosome losses are only a short, transient effect.This was unexpected, as the securin loss resulted in a persisting reduction of the sister-separating protease separase and inefficient cleavage of the cohesin subunit Scc1.We propose that human cells possess efficient mechanisms to compensate for the loss of genes involved in chromosome segregation.

View Article: PubMed Central - PubMed

Affiliation: Institute of Human Genetics, Technical University Munich, Munich, Germany.

ABSTRACT
Abnormalities of chromosome number are frequently observed in cancers. The mechanisms regulating chromosome segregation in human cells are therefore of great interest. Recently it has been reported that human cells without an hSecurin gene lose chromosomes at a high frequency. Here we show that, after hSecurin knockout through homologous recombination, chromosome losses are only a short, transient effect. After a few passages hSecurin(-/-) cells became chromosomally stable and executed mitoses normally. This was unexpected, as the securin loss resulted in a persisting reduction of the sister-separating protease separase and inefficient cleavage of the cohesin subunit Scc1. Our data demonstrate that securin is dispensable for chromosomal stability in human cells. We propose that human cells possess efficient mechanisms to compensate for the loss of genes involved in chromosome segregation.

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hSecurin−/− Cells Regain Chromosomal Stability Quickly after hSecurin Knockout by Homologous Recombination: Summary of M-FISH Analysis of hSecurin−/− Cells at Different Passages(A–D) Graphic summary of M-FISH data from hSecurin−/− cells at passages 2 (A), 3 (B), 8 (C), and 12 (D). At each passage point 20 or 30 metaphase spreads were painted by M-FISH and analyzed for alterations of chromosome structure and number. Loss of a single copy of a given chromosome is marked in red, loss of both copies is marked in crimson, and gain of a single chromosome is marked in green. Rows indicate the analyzed metaphase spreads (m1 to m30 or m20); columns indicate the chromosome number (1–22 and X).(E) Graphic representation of the percentages of metaphase spreads with chromosomal copy number aberrations at different passages for the series of experiments shown in (A–D) (blue line) and for a repeat experiment (purple line).(F) M-FISH karyotype of a passage 12 hSecurin−/− cell, showing that the karyotype is identical to that of the parent cell line HCT116 (for details, see text).
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pbio-0030416-g001: hSecurin−/− Cells Regain Chromosomal Stability Quickly after hSecurin Knockout by Homologous Recombination: Summary of M-FISH Analysis of hSecurin−/− Cells at Different Passages(A–D) Graphic summary of M-FISH data from hSecurin−/− cells at passages 2 (A), 3 (B), 8 (C), and 12 (D). At each passage point 20 or 30 metaphase spreads were painted by M-FISH and analyzed for alterations of chromosome structure and number. Loss of a single copy of a given chromosome is marked in red, loss of both copies is marked in crimson, and gain of a single chromosome is marked in green. Rows indicate the analyzed metaphase spreads (m1 to m30 or m20); columns indicate the chromosome number (1–22 and X).(E) Graphic representation of the percentages of metaphase spreads with chromosomal copy number aberrations at different passages for the series of experiments shown in (A–D) (blue line) and for a repeat experiment (purple line).(F) M-FISH karyotype of a passage 12 hSecurin−/− cell, showing that the karyotype is identical to that of the parent cell line HCT116 (for details, see text).

Mentions: In an initial step, metaphase spreads of the hSecurin−/− cell line were karyotyped by multiplex fluorescence in situ hybridization (M-FISH) at different passages (Figure 1). For passages 2 and 3, we confirmed the loss of numerous chromosomes in the majority of analyzed cells (Figure 1A and 1B). About 60% (12/20) of metaphase spreads showed losses of at least one chromosome. Surprisingly however, the high rate of chromosome losses in the hSecurin−/− cell line had almost vanished by passage 8 (Figure 1C), when chromosome losses were noted in only 10% (2/20) of cells. By passage 12, we observed no chromosome losses (Figure 1D). In the latter two analyses, merely one metaphase spread each had a gain of a single chromosome (Figure 1C and 1D).


Securin is not required for chromosomal stability in human cells.

Pfleghaar K, Heubes S, Cox J, Stemmann O, Speicher MR - PLoS Biol. (2005)

hSecurin−/− Cells Regain Chromosomal Stability Quickly after hSecurin Knockout by Homologous Recombination: Summary of M-FISH Analysis of hSecurin−/− Cells at Different Passages(A–D) Graphic summary of M-FISH data from hSecurin−/− cells at passages 2 (A), 3 (B), 8 (C), and 12 (D). At each passage point 20 or 30 metaphase spreads were painted by M-FISH and analyzed for alterations of chromosome structure and number. Loss of a single copy of a given chromosome is marked in red, loss of both copies is marked in crimson, and gain of a single chromosome is marked in green. Rows indicate the analyzed metaphase spreads (m1 to m30 or m20); columns indicate the chromosome number (1–22 and X).(E) Graphic representation of the percentages of metaphase spreads with chromosomal copy number aberrations at different passages for the series of experiments shown in (A–D) (blue line) and for a repeat experiment (purple line).(F) M-FISH karyotype of a passage 12 hSecurin−/− cell, showing that the karyotype is identical to that of the parent cell line HCT116 (for details, see text).
© Copyright Policy
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC1287505&req=5

pbio-0030416-g001: hSecurin−/− Cells Regain Chromosomal Stability Quickly after hSecurin Knockout by Homologous Recombination: Summary of M-FISH Analysis of hSecurin−/− Cells at Different Passages(A–D) Graphic summary of M-FISH data from hSecurin−/− cells at passages 2 (A), 3 (B), 8 (C), and 12 (D). At each passage point 20 or 30 metaphase spreads were painted by M-FISH and analyzed for alterations of chromosome structure and number. Loss of a single copy of a given chromosome is marked in red, loss of both copies is marked in crimson, and gain of a single chromosome is marked in green. Rows indicate the analyzed metaphase spreads (m1 to m30 or m20); columns indicate the chromosome number (1–22 and X).(E) Graphic representation of the percentages of metaphase spreads with chromosomal copy number aberrations at different passages for the series of experiments shown in (A–D) (blue line) and for a repeat experiment (purple line).(F) M-FISH karyotype of a passage 12 hSecurin−/− cell, showing that the karyotype is identical to that of the parent cell line HCT116 (for details, see text).
Mentions: In an initial step, metaphase spreads of the hSecurin−/− cell line were karyotyped by multiplex fluorescence in situ hybridization (M-FISH) at different passages (Figure 1). For passages 2 and 3, we confirmed the loss of numerous chromosomes in the majority of analyzed cells (Figure 1A and 1B). About 60% (12/20) of metaphase spreads showed losses of at least one chromosome. Surprisingly however, the high rate of chromosome losses in the hSecurin−/− cell line had almost vanished by passage 8 (Figure 1C), when chromosome losses were noted in only 10% (2/20) of cells. By passage 12, we observed no chromosome losses (Figure 1D). In the latter two analyses, merely one metaphase spread each had a gain of a single chromosome (Figure 1C and 1D).

Bottom Line: Here we show that, after hSecurin knockout through homologous recombination, chromosome losses are only a short, transient effect.This was unexpected, as the securin loss resulted in a persisting reduction of the sister-separating protease separase and inefficient cleavage of the cohesin subunit Scc1.We propose that human cells possess efficient mechanisms to compensate for the loss of genes involved in chromosome segregation.

View Article: PubMed Central - PubMed

Affiliation: Institute of Human Genetics, Technical University Munich, Munich, Germany.

ABSTRACT
Abnormalities of chromosome number are frequently observed in cancers. The mechanisms regulating chromosome segregation in human cells are therefore of great interest. Recently it has been reported that human cells without an hSecurin gene lose chromosomes at a high frequency. Here we show that, after hSecurin knockout through homologous recombination, chromosome losses are only a short, transient effect. After a few passages hSecurin(-/-) cells became chromosomally stable and executed mitoses normally. This was unexpected, as the securin loss resulted in a persisting reduction of the sister-separating protease separase and inefficient cleavage of the cohesin subunit Scc1. Our data demonstrate that securin is dispensable for chromosomal stability in human cells. We propose that human cells possess efficient mechanisms to compensate for the loss of genes involved in chromosome segregation.

Show MeSH
Related in: MedlinePlus