title: Tethering sister centromeres to each other suggests the spindle checkpoint detects stretch within the kinetochore.
abstract: The spindle checkpoint ensures that newly born cells receive one copy of each chromosome by preventing chromosomes from segregating until they are all correctly attached to the spindle. The checkpoint monitors tension to distinguish between correctly aligned chromosomes and those with both sisters attached to the same spindle pole. Tension arises when sister kinetochores attach to and are pulled toward opposite poles, stretching the chromatin around centromeres and elongating kinetochores. We distinguished between two hypotheses for where the checkpoint monitors tension: between the kinetochores, by detecting alterations in the distance between them, or by responding to changes in the structure of the kinetochore itself. To distinguish these models, we inhibited chromatin stretch by tethering sister chromatids together by binding a tetrameric form of the Lac repressor to arrays of the Lac operator located on either side of a centromere. Inhibiting chromatin stretch did not activate the spindle checkpoint; these cells entered anaphase at the same time as control cells that express a dimeric version of the Lac repressor, which cannot cross link chromatids, and cells whose checkpoint has been inactivated. There is no dominant checkpoint inhibition when sister kinetochores are held together: cells expressing the tetrameric Lac repressor still arrest in response to microtubule-depolymerizing drugs. Tethering chromatids together does not disrupt kinetochore function; chromosomes are successfully segregated to opposite poles of the spindle. Our results indicate that the spindle checkpoint does not monitor inter-kinetochore separation, thus supporting the hypothesis that tension is measured within the kinetochore.
authors.affiliation: Molecular and Cellular Biology Department, Harvard University, Cambridge, Massachusetts, United States of America; FAS Center for Systems Biology, Harvard University, Cambridge, Massachusetts, United States of America.
authors.forename: Natalie J
authors.initials: NJ
authors.lastname: Nannas
authors.affiliation: Molecular and Cellular Biology Department, Harvard University, Cambridge, Massachusetts, United States of America; FAS Center for Systems Biology, Harvard University, Cambridge, Massachusetts, United States of America.
authors.forename: Andrew W
authors.initials: AW
authors.lastname: Murray
mesh_terms: D002503:Centromere / Q000235:genetics*; D002842:Chromatids / Q000235:genetics; D002843:Chromatin / Q000235:genetics; D020090:Chromosome Segregation / Q000235:genetics*; D002875:Chromosomes / Q000235:genetics; D006801:Humans; D018386:Kinetochores; D056968:Lac Repressors / Q000235:genetics; D059566:M Phase Cell Cycle Checkpoints / Q000235:genetics*; D008870:Microtubules / Q000235:genetics; D008938:Mitosis / Q000235:genetics; D012441:Saccharomyces cerevisiae; D008941:Spindle Apparatus / Q000235:genetics
chemical_list: D002843:Chromatin; D056968:Lac Repressors
references.citation: Chromosoma. 2008 Apr;117(2):123-35
references.pmid: 17989990
references.citation: Dev Cell. 2003 Nov;5(5):735-45
references.pmid: 14602074
references.citation: Science. 2009 Mar 6;323(5919):1350-3
references.pmid: 19150808
references.citation: Science. 2005 Jan 7;307(5706):130-3
references.pmid: 15637284
references.citation: J Cell Biol. 2009 Feb 9;184(3):373-81
references.pmid: 19193623
references.citation: Nat Rev Mol Cell Biol. 2012 Dec;13(12):789-803
references.pmid: 23175282
references.citation: Nature. 2013 May 2;497(7447):118-21
references.pmid: 23604256
references.citation: Curr Biol. 2001 Sep 18;11(18):1462-7
references.pmid: 11566107
references.citation: Science. 1996 Oct 11;274(5285):242-6
references.pmid: 8824188
references.citation: Cell. 1991 Aug 9;66(3):507-17
references.pmid: 1651171
references.citation: J Mol Biol. 2008 Nov 21;383(4):894-903
references.pmid: 18793650
references.citation: Nat Genet. 2007 Oct;39(10):1273-7
references.pmid: 17828265
references.citation: J Cell Biol. 1995 Aug;130(4):941-8
references.pmid: 7642709
references.citation: Cell. 1991 Aug 9;66(3):519-31
references.pmid: 1651172
references.citation: Cell. 2006 Dec 15;127(6):1179-91
references.pmid: 17174893
references.citation: Cell. 2002 Feb 8;108(3):317-29
references.pmid: 11853667
references.citation: Cell. 2007 Jan 12;128(1):101-14
references.pmid: 17218258
references.citation: Genes Dev. 2001 Dec 1;15(23):3118-29
references.pmid: 11731476
references.citation: Cell. 2000 Mar 17;100(6):619-33
references.pmid: 10761928
references.citation: EMBO J. 1993 May;12(5):1969-78
references.pmid: 8491189
references.citation: J Cell Biol. 2011 Apr 4;193(1):125-40
references.pmid: 21464230
references.citation: Cell. 2009 May 15;137(4):672-84
references.pmid: 19450515
references.citation: Annu Rev Biochem. 2007;76:563-91
references.pmid: 17362199
references.citation: Nat Rev Cancer. 2005 Oct;5(10):773-85
references.pmid: 16195750
references.citation: J Biochem. 2012 Nov;152(5):415-24
references.pmid: 22995986
references.citation: N Engl J Med. 2009 Jun 11;360(24):2556-62
references.pmid: 19516035
references.citation: J Cell Biol. 2009 Feb 9;184(3):383-90
references.pmid: 19188492
references.citation: Mol Cell Biol. 1987 Jul;7(7):2397-405
references.pmid: 3302676
references.citation: Curr Biol. 1996 Dec 1;6(12):1599-608
references.pmid: 8994824
references.citation: J Mol Biol. 1978 Aug 15;123(3):431-54
references.pmid: 211238
references.citation: Nature. 1995 Feb 16;373(6515):630-2
references.pmid: 7854422
references.citation: Nat Cell Biol. 2009 Sep;11(9):1116-20
references.pmid: 19684576
references.citation: J Cell Biol. 1998 Jun 1;141(5):1181-91
references.pmid: 9606210
references.citation: Annu Rev Cell Dev Biol. 2012;28:189-214
references.pmid: 22804579
references.citation: Cell. 2000 Jun 23;101(7):763-75
references.pmid: 10892747
references.citation: J Cell Sci. 2010 Mar 15;123(Pt 6):825-35
references.pmid: 20200228
references.citation: Nat Cell Biol. 2006 Jan;8(1):78-83
references.pmid: 16327780
references.citation: Chromosoma. 2008 Dec;117(6):521-33
references.pmid: 18677502
references.citation: Curr Biol. 2007 Nov 6;17(21):1837-46
references.pmid: 17980598
references.citation: Nat Cell Biol. 2000 Aug;2(8):492-9
references.pmid: 10934469