The centromere and kinetochore Flashcards
what is a centromere?
in three short parts
- A constricted region on a chromosome that joins sister chromatids
- The site where kinetochore is formed
- Specialised fragment of DNA, which allows sister chromatids to segregate
what is a kinetochore?
in three parts
- A multiprotein complex that forms at the centromere
- Specialised structure which allows sister chromatids to segregate during cell division
- Site on a chromosome where microtubules attach
how is centromeric chromatin different from euchromatin or heterochromatin?
nucelosomes of centromeric chromatin contain CENP-A (CenH) (centromeric marker)
CENP-A containing arrays are more condensed than canonical ones
(unqie set of histone markers, presence of centromeric proteins)
what is CENP-As main role?
defines the position of the centromere
whats the difference in association of CCAN and kinetochores with centromeres
cell cycle position
CCAN components are at centromeres throughout the cell cycle
Kinetochore components are at centromeres only during mitosis
what are the three major functions of the kinetochore?
- Capturing microtubules to form a connection between chromosomes and mitotic spindles
- Identifying incorrect attachments and repairing them
- Harnessing the force to generate movement of chromosomes during anaphase
when and where are kinetochores and CCAN complex organised
Kinetochores are assembled on centromeric chromatin in the beginning of mitosis
CCAN complex organised on CENPA containing nucelosomes and are present throughout the cell cycle
CCan is on cenpa nuclesomes, on top of which is the kmn network
what is the structural core of the kinetochore and what are its components?
the KMN network
three complexes: * KNL1/Spc105 complex * Mis12 complex * Ndc80 complex
the human KMN network is connected to the centromere via 2 separate pathways involving what proteins
centromere/kinetochore genes are misregulated in many cancers, what can overexpression of these genes indicate in the clinical status of cancer?
correlate with increased levels of genomic instability and several specific adverse tumour properties,
prognostically poor patient for survival for breast and lung cancers, especially early-stage tumours
levels of the overexpression can help to forecast patient response to adjuvant chemotherapy or radiotherapy
what are the two major proteins that help load CENP-A onto the DNA
MIS18 (marks where new CENPA is going to be loaded)
HJURP (directly participates in loading)
How is bi-polar attachment of chromosomes achieved?
the trial and error approach
Attachment of only one kinetochore is unstable – it is removed.
All attachments are unstable except for the bipolar. - Lasts for longer
Incorrect attachments are not stable and do not last.
Correct attachment becomes ‘locked’ in space
how are incorrect attachments repaired?
the chromosome passenger complex (CPC) aka Aurora B complex
when kinetochores are not properly attached, the tension is low the outer kinetochore is closer to the inner centromere where CPC is localised. Aurora B phosphorylates Ndc80 protein weakening interaction with the microtubules which are then released.
when the kinetochore is properly attached to microtubules, the high tension removes Ndc80 from the reach of Aurora B kinase and the attachments becomes stable
kinetochore has spring like properties that move away or towards the centromere under higher or lower tension
how are incorrect attachments repaired?
the chromosome passenger complex (CPC) aka Aurora B complex
when kinetochores are not properly attached, the tension is low the outer kinetochore is closer to the inner centromere where CPC is localised. Aurora B phosphorylates Ndc80 protein weakening interaction with the microtubules which are then released.
when the kinetochore is properly attached to microtubules, the high tension removes Ndc80 from the reach of Aurora B kinase and the attachments becomes stable
kinetochore has spring like properties that move away or towards the centromere under higher or lower tension
what are the functions for each of the Aurora kinases A B and C?
- Aurora A kinase is localised primarily to centrosomes and it controls centrosomal activities, e.g. mitotic spindle formation
- Aurora B kinase is a component of CPC and its localisation changes from inner-centromeric to microtubules of the central spindle and midzone. It participates in chromosome condensation, segregation and cytokinesis
- Aurora C is involved in meiosis
what happens witch Aurora B depletion in cells?
Not so well aligned at the metaphase plate and chromosome progression is already limited
inhibitors targetting aurora kinases are examples of what types of drugs used in cancer therapy
anti-mitotic drugs
what happens when we counteract aurora kinase overexpression in cancer cells?
some cancer cells get addicted to the overexpession so when we decrease it the cell suffer
aurora kinase inhibition leads to cell death by apoptosis
- aurora A inhibition -> monopolar spindle, prolonged mitosis or perturbed prometaphase then sever aneuploidy and so cell deaath
- aurora B inhibition -> unaligned chromosomes then premature mitotic exit and polyploidy and so cell death
how do anti-mitotic drugs work in cancer therapy?
Because cancer cells proliferate uncontrollable, anti-mitotic drugs should stop cells from dividing, as a consequence, stop the tumour from further growth. Anti-mitotic drugs are expected to arrest cell cycle for extended period of time
give three examples of targets for anti-mitotic drugs
- microtubules, as major components of mitotic spindle (stabilisers, de-stabilisers)
- kinesins, as major regulators of microtubule dynamicity
- mitotic kinases, as major regulators of cell cycle and cell division (CDKs, PLKs, Aurora kinases, Wee 1 kinases)
what proteins make up the cohesin complex?
SMC subunits = SMC1 and SMC3
non-SMC subunits = SCC3 and RAD21
when is the cohesin complex loaded and unloaded from the chromosomes?
It is loaded on chromosomes during G1 phase, after DNA replication it holds sister chromatids together.
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Its release from chromosome arms in prophase coincides with the axial compression of chromosomes during mitosis
Along with CTCF it defines borders of chromatin units during interphase
when is the cohesin complex loaded and unloaded from the chromosomes?
It is loaded on chromosomes during G1 phase, after DNA replication it holds sister chromatids together.
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Its release from chromosome arms in prophase coincides with the axial compression of chromosomes during mitosis
Along with CTCF it defines borders of chromatin units during interphase
what are the functions of cohesin in mitosis?
- Sister chromatid cohesion (at centromeres)
- Holding together sister centrioles
what are the function of cohesins in meiosis?
- Pairing of homologous chromosomes during meiosis
- Assembly of the axes of synaptonemal complex in meiosis
- Coordination of sister kinetochores during first meiotic division
what are the functions of cohesin during interphase?
- Sister chromatids cohesion (entire chromatin)
- Repair of DNA breaks
- Assembly of DNA replication factories during S phase
- Regulation of transcription
- Organisation of chromatin loops and TADs
describe the cohesin cycle?
In vertebrates cohesin is loaded onto DNA just after mitosis in a “non-cohesive” form. (no strands to hold together). Cohesin loading factor Scc2 is required for this step.
Sister chromatids created after dna replication in s phase - Eco1 (Esco1/Esco2) acetylate cohesin during S phase to establish “cohesive” cohesin that holds sister chromatids together
Cohesin is removed completely from chromatin during cell division (M phase)
removal of cohesin that enables chromosomes to segregate occurs in two waves, what are they?
Prophase pathway - Mitotic kinases + Wapl
(removes all but the centromereic cohesin because it is protected by two proteins)
Metaphase pathway - Separase
(removes all the rest)
what are the two major prerequisites of the metaphase to anaphase transition?
what are both of these events triggered and regulated by?
- inactivation of Cdk1
- activation of Separase
triggered by the degredation of regulatory proteins via the proteasome pathway - by Anaphase-promoting complex/ cyclosome (APC/C)
regulated by the spindle assembly checkpoint (SAC)
how does the spindle assembly checkpoint work?
Unattached kinetochores generate “STOP” signal, which blocks activity of APC/C. This signal is a complex of 4 proteins that is called Mitotic Checkpoint Complex (MCC). It consists of BubR1, Bub3, Mad2 and Cdc20, and binds directly APC/C and inhibits it.
When all kinetochores become attached to microtubules of the mitotic spindle, the MCC is no longer produced and APC/C becomes active. it is then able to ubiquitilate cyclin B1 and securin (which leads to inactivation of cdk1 and activation of separase) - the cell can now exit mitosis
what mitotic kinase is repsonsible for sensing the attachments of MTs and KTs
Mps1
what is a common feature of all SAC components
they are recruited to unattached kinetochores but not to properly attached ones
what is the point of the spindle assembly checkpoint?
SAC activity gives a cell more time to establish proper MT-KT attachments. Without this extra time cells may proceed into anaphase before these attachments are made.(which would result in aneuploidy)
what is the point of the spindle assembly checkpoint?
SAC activity gives a cell more time to establish proper MT-KT attachments. Without this extra time cells may proceed into anaphase before these attachments are made.(which would result in aneuploidy)
why should aurora b be considered as part of SAC?
Aurora B activity is needed for proper function of bona fide SAC components
(for example, Aurora B inhibition prevents the recruitment of Mps1 to kinetochores)
- By participating in the error correction mechanism it generates unattached kinetochores that are recognised by SAC
- It directly participates in SAC
what’s the most frequently mutated component of cohesin complex?
the STAG2 (SA2, Scc3-types) subunit
what function of cohesin does not involve DNA?
holding centrioles in centrosomes together
what are the similarities and differences in the centromere and centromere separation involving cohesin degredation and cleavage?
give some examples of antimitotic drugs already used in cancer treatment
one for each target
AT9283 & Alisertib - aurora A/B inhibtion,
Taxanes and Vinca alkaloids - microtubules/mitotic spindle,
ispinesib - kinesin spindle protein or
MK-1775 - Wee1 inhibition.
To target cdks there are four inhihibtors, pablociclib, ribociclib, abemaciclib, flavopiridol
