BUB1
Encyclopedia
Mitotic checkpoint serine/threonine-protein kinase BUB1 also known as BUB1 (budding uninhibited by benzimidazoles 1) is an enzyme
that in humans is encoded by the BUB1 gene
.
Bub1 is a serine/threonine protein kinase first identified in genetic screens of Saccharomyces cerevisiae
. The protein is bound to kinetochore
s and plays a key role in the establishment of the mitotic spindle checkpoint
and chromosome congression. The mitotic checkpoint kinase is evolutionary conserved in organisms as diverse as Saccharomyces cerevisiae and humans. Loss-of-function mutations or absence of Bub1 has been reported to result in aneuploidy
, chromosomal instability (CIN
) and premature senescence
.
(SAC) activation. The crystal structure of human Bub1 revealed the presence of a N-terminal tetratricopeptide
repeat (TPR) domain and a C-terminal kinase domain (residues 784–1085), adopting a canonical kinase fold with two lobes. The ATP
binding and the catalytic sites are located at the interface of the two lobes. The N-terminal extension contains three β-strands
and an α-helix
, wrapping around the N lobe of the kinase domain.
and S
phase of the cell cycle
, peaks at G2/M, and drops dramatically after mitosis. During prophase
it localizes as one of the first proteins to the outer kinetochore, a process generally implicated in correct mitotic timing and checkpoint response to spindle damage.
In eukaryotic cells the SAC serves as the central surveillance mechanism to ensure chromosomes are being passed on to the next generation in a reliable manner. Several components monitor correct bipolar attachment of microtubules to the kinetochore, presumably through detection of tension. Metaphase-to-anaphase
transition is halted by the SAC as long as single kinetochores lack bipolar microtubule attachment, implying the need for a highly sensitive signaling pathway. Bub1 was claimed to be the master regulator of SAC formation and signaling. At least thirteen other proteins (Mad1
, MAD2
, MAD3
/BubR1, BUB3
, Mps1
etc.) are part of the check point, among which many have been identified to interact with Bub1.
Upon activation of the SAC Bub1 directly phosphorylates APC
/C’s coactivator Cdc20
. This phosphorylation event is probably achieved in complex with Bub3, which itself has been subjected to prior phosphorylation by Bub1. The phosphorylation of Cdc20 ultimately leads to decreased activity of APC/C which determines the metaphase-to-anaphase transition. In turn APC/C, now in complex with Cdh1, also acts on Bub1 by priming it for degradation to exit mitosis.
In addition, kinetochore localization of Bub1 early during G2 or prophase is another aspect of SAC functioning. Bub1 is thought to serve as a platform recruiting other checkpoint and motor proteins as Mad1, Mad2, BubR1, CENP-E and PLK1
to the kinetochore. Indeed, recent data suggest that the primary role of Bub1 during SAC activity is not Cdc20 phosphorylation but rather recruitment of BubR1, Mad1 and Mad2.
Upon spindle damage Bub1 is also triggered to phosphorylate Mad1 leading to dissociation of the Mad1-Mad2 complex and thereby rendering Mad2 accessible for inhibition of Cdc20.
Bub1 generally protects sister chromatide cohesion by enhancing Shugoshin protein (Sgo1
) localization to the centromeric region. Through recruitment of the phosphatase PP2A Bub1 inhibits the action of PLK1, which removes Sgo1 from the centromere.
Contrarily PLK1 localization, as mentioned, also depends on the activity of Bub1. Studies in Xenopus extracts using RNAi or antibody depletion have indicated a crucial function of Bub1 in the organization of the inner centromere. Similarly to its role in kinetochore assembly, it recruits members of the chromosomal passenger complex (CPC) like Aurora B kinase
, Survivin
and INCENP
. Direct phosphorylation of INCENP by Bub1 has been observed.
RNAi mediated depletion of human Bub1 has indicated function in correct metaphase congression. Downstream targets identified are distinct kinetochore proteins as CENP-F
, MCAK
and the mentioned Sgo1.
Indications for possible Bub1 involvement in tumorigenesis also derive from animal experiments, where mice with reduced Bub1 expression showed an increase in tumor susceptibility. In vitro knockdown of Bub1 in p53
impaired cells (e.g. HeLa cells) caused aneuploidy. Whether aneuploidy alone is a sufficient driving cause during tumorigenesis or rather a mere consequence has been a matter of scientific debate.
is thereby inhibited via phosphorylation. Direct interaction between these two players has not been visualized so far, therefore molecules linking Bub1 and p73 are yet to be determined.
It has also been proposed that Bub1 binds p53 to prevent it from activating pro-apoptotic genes, therefore p53 is able to induce apoptosis
when Bub1 is depleted. However, an interaction between p53 and Bub1 has not yet been shown while p53 binding BubR1 has been reported.
Enzyme
Enzymes are proteins that catalyze chemical reactions. In enzymatic reactions, the molecules at the beginning of the process, called substrates, are converted into different molecules, called products. Almost all chemical reactions in a biological cell need enzymes in order to occur at rates...
that in humans is encoded by the BUB1 gene
Gene
A gene is a molecular unit of heredity of a living organism. It is a name given to some stretches of DNA and RNA that code for a type of protein or for an RNA chain that has a function in the organism. Living beings depend on genes, as they specify all proteins and functional RNA chains...
.
Bub1 is a serine/threonine protein kinase first identified in genetic screens of Saccharomyces cerevisiae
Saccharomyces cerevisiae
Saccharomyces cerevisiae is a species of yeast. It is perhaps the most useful yeast, having been instrumental to baking and brewing since ancient times. It is believed that it was originally isolated from the skin of grapes...
. The protein is bound to kinetochore
Kinetochore
The kinetochore is the protein structure on chromatids where the spindle fibers attach during cell division to pull sister chromatids apart....
s and plays a key role in the establishment of the mitotic spindle checkpoint
Spindle checkpoint
In order to preserve one cell's identity and its proper functioning, it is necessary to maintain constant the appropriate number of chromosomes after each cell division...
and chromosome congression. The mitotic checkpoint kinase is evolutionary conserved in organisms as diverse as Saccharomyces cerevisiae and humans. Loss-of-function mutations or absence of Bub1 has been reported to result in aneuploidy
Aneuploidy
Aneuploidy is an abnormal number of chromosomes, and is a type of chromosome abnormality. An extra or missing chromosome is a common cause of genetic disorders . Some cancer cells also have abnormal numbers of chromosomes. Aneuploidy occurs during cell division when the chromosomes do not separate...
, chromosomal instability (CIN
Chromosome instability syndrome
Chromosome instability syndromes are a group of inherited conditions associated with chromosomal instability and breakage. They often lead to an increased tendency to develop certain types of malignancies....
) and premature senescence
Senescence
Senescence or biological aging is the change in the biology of an organism as it ages after its maturity. Such changes range from those affecting its cells and their function to those affecting the whole organism...
.
Structure
Bub1p comprises a conserved N-terminal region, a central non-conserved region and a C-terminal serine/threonine kinase domain. The N-terminal region mediates binding of Hs-BUB1 to the mitotic kinetochore protein blinkin (a protein also commonly referred to as AF15q14). The latter interaction is essential for kinetochore localization of Bub1 and its function in cell cycle arrest induced by spindle assembly checkpointSpindle checkpoint
In order to preserve one cell's identity and its proper functioning, it is necessary to maintain constant the appropriate number of chromosomes after each cell division...
(SAC) activation. The crystal structure of human Bub1 revealed the presence of a N-terminal tetratricopeptide
Tetratricopeptide
The tetratricopeptide repeat is a structural motif. It consists in a degenerate 34 amino acid sequence motif identified in a wide variety of proteins. It is found in tandem arrays of 3–16 motifs, which form scaffolds to mediate protein–protein interactions and often the assembly of multiprotein...
repeat (TPR) domain and a C-terminal kinase domain (residues 784–1085), adopting a canonical kinase fold with two lobes. The ATP
Adenosine triphosphate
Adenosine-5'-triphosphate is a multifunctional nucleoside triphosphate used in cells as a coenzyme. It is often called the "molecular unit of currency" of intracellular energy transfer. ATP transports chemical energy within cells for metabolism...
binding and the catalytic sites are located at the interface of the two lobes. The N-terminal extension contains three β-strands
Beta sheet
The β sheet is the second form of regular secondary structure in proteins, only somewhat less common than the alpha helix. Beta sheets consist of beta strands connected laterally by at least two or three backbone hydrogen bonds, forming a generally twisted, pleated sheet...
and an α-helix
Alpha helix
A common motif in the secondary structure of proteins, the alpha helix is a right-handed coiled or spiral conformation, in which every backbone N-H group donates a hydrogen bond to the backbone C=O group of the amino acid four residues earlier...
, wrapping around the N lobe of the kinase domain.
Subcellular location
In humans Bub1 accumulates gradually during G1G1 phase
The G1 phase is a period in the cell cycle during interphase, before the S phase. For many cells, this phase is the major period of cell growth during its lifespan. During this stage new organelles are being synthesized, so the cell requires both structural proteins and enzymes, resulting in great...
and S
S phase
S-phase is the part of the cell cycle in which DNA is replicated, occurring between G1 phase and G2 phase. Precise and accurate DNA replication is necessary to prevent genetic abnormalities which often lead to cell death or disease. Due to the importance, the regulatory pathways that govern this...
phase of the cell cycle
Cell cycle
The cell cycle, or cell-division cycle, is the series of events that takes place in a cell leading to its division and duplication . In cells without a nucleus , the cell cycle occurs via a process termed binary fission...
, peaks at G2/M, and drops dramatically after mitosis. During prophase
Prophase
Prophase, from the ancient Greek πρό and φάσις , is a stage of mitosis in which the chromatin condenses into a highly ordered structure called a chromosome in which the chromatin becomes visible. This process, called chromatin condensation, is mediated by the condensin complex...
it localizes as one of the first proteins to the outer kinetochore, a process generally implicated in correct mitotic timing and checkpoint response to spindle damage.
Function
The protein kinase Bub1 possesses versatile and distinct functions during the cell cycle, mainly in the SAC and chromosome alignment during metaphase. The protein’s interaction network currently identified is similarly complex (see Figure 1).In eukaryotic cells the SAC serves as the central surveillance mechanism to ensure chromosomes are being passed on to the next generation in a reliable manner. Several components monitor correct bipolar attachment of microtubules to the kinetochore, presumably through detection of tension. Metaphase-to-anaphase
Anaphase
Anaphase, from the ancient Greek ἀνά and φάσις , is the stage of mitosis or meiosis when chromosomes move to opposite poles of the cell....
transition is halted by the SAC as long as single kinetochores lack bipolar microtubule attachment, implying the need for a highly sensitive signaling pathway. Bub1 was claimed to be the master regulator of SAC formation and signaling. At least thirteen other proteins (Mad1
Mad1
Mitotic spindle assembly checkpoint protein MAD1 is a protein that in humans is encoded by the MAD1L1 gene.MAD1L1 is also known as Human Accelerated Region 3. It may therefore have played a key role in differentiating Humans from Apes.-Interactions:...
, MAD2
MAD2
MAD2 is an essential spindle checkpoint protein. The spindle checkpoint system is a regulatory system that restrains progression through the metaphase-to-anaphase transition. The Mad2 gene was first identified in the yeast S. cerevisiae in a screen for genes which when mutated would confer...
, MAD3
BUB1B
Mitotic checkpoint serine/threonine-protein kinase BUB1 beta is an enzyme that in humans is encoded by the BUB1B gene.-Interactions:BUB1B has been shown to interact with AP2B1, HDAC1, BUB3, MAD2L1, Gamma-synuclein, BRCA2 and CDC20.-Further reading:...
/BubR1, BUB3
BUB3
Mitotic checkpoint protein BUB3 is a protein that in humans is encoded by the BUB3 gene.Bub3 is a protein involved with the regulation of the Spindle Assembly Checkpoint ; though BUB3 is non-essential in yeast, it is essential in higher eukaryotes...
, Mps1
RPS27
40S ribosomal protein S27 is a protein that in humans is encoded by the RPS27 gene.-Further reading:...
etc.) are part of the check point, among which many have been identified to interact with Bub1.
Upon activation of the SAC Bub1 directly phosphorylates APC
Anaphase-promoting complex
Anaphase-Promoting Complex, also called cyclosome , is an E3 ubiquitin ligase that marks target cell cycle proteins for degradation by the 26S proteasome. The APC/C is a large complex of 11–13 subunit proteins, including a cullin and RING subunit much like SCF...
/C’s coactivator Cdc20
CDC20
The cell-division cycle protein 20 is an essential regulator of cell division that is encoded by the CDC20 gene in humans. To the best of current knowledge its most important function is to activate the anaphase promoting complex , a large 11-13 subunit complex that initiates chromatid separation...
. This phosphorylation event is probably achieved in complex with Bub3, which itself has been subjected to prior phosphorylation by Bub1. The phosphorylation of Cdc20 ultimately leads to decreased activity of APC/C which determines the metaphase-to-anaphase transition. In turn APC/C, now in complex with Cdh1, also acts on Bub1 by priming it for degradation to exit mitosis.
In addition, kinetochore localization of Bub1 early during G2 or prophase is another aspect of SAC functioning. Bub1 is thought to serve as a platform recruiting other checkpoint and motor proteins as Mad1, Mad2, BubR1, CENP-E and PLK1
PLK1
Serine/threonine-protein kinase PLK1, also known as polo-like kinase 1 or serine/threonine-protein kinase 13 , is an enzyme that in humans is encoded by the PLK1 gene.- Structure :...
to the kinetochore. Indeed, recent data suggest that the primary role of Bub1 during SAC activity is not Cdc20 phosphorylation but rather recruitment of BubR1, Mad1 and Mad2.
Upon spindle damage Bub1 is also triggered to phosphorylate Mad1 leading to dissociation of the Mad1-Mad2 complex and thereby rendering Mad2 accessible for inhibition of Cdc20.
Bub1 generally protects sister chromatide cohesion by enhancing Shugoshin protein (Sgo1
SGOL1
Shugoshin-like 1 is a protein that in humans is encoded by the SGOL1 gene.-Further reading:...
) localization to the centromeric region. Through recruitment of the phosphatase PP2A Bub1 inhibits the action of PLK1, which removes Sgo1 from the centromere.
Contrarily PLK1 localization, as mentioned, also depends on the activity of Bub1. Studies in Xenopus extracts using RNAi or antibody depletion have indicated a crucial function of Bub1 in the organization of the inner centromere. Similarly to its role in kinetochore assembly, it recruits members of the chromosomal passenger complex (CPC) like Aurora B kinase
Aurora B kinase
Aurora B kinase is a protein that functions in the attachment of the mitotic spindle to the centromere.In cancerous cells, over-expression of these enzymes causes unequal distribution of genetic information, creating aneuploid cells, a hallmark of cancer....
, Survivin
Survivin
Survivin, also called baculoviral inhibitor of apoptosis repeat-containing 5 or BIRC5, is a protein that, in humans, is encoded by the BIRC5 gene....
and INCENP
INCENP
Inner centromere protein is a protein that in humans is encoded by the INCENP gene.In mammalian cells, two broad groups of centromere-interacting proteins have been described: constitutively binding centromere proteins and 'passenger' proteins...
. Direct phosphorylation of INCENP by Bub1 has been observed.
RNAi mediated depletion of human Bub1 has indicated function in correct metaphase congression. Downstream targets identified are distinct kinetochore proteins as CENP-F
CENPF
Centromere protein F is a protein that in humans is encoded by the CENPF gene.-Further reading:...
, MCAK
KIF2C
Kinesin-like protein KIF2C is a protein that in humans is encoded by the KIF2C gene.-Further reading:...
and the mentioned Sgo1.
Implications in cancer
Disturbed mitotic checkpoints are a common feature of many human cancers. More precisely, mutations in the spindle checkpoint can lead to chromosomal instability and aneuploidy, a feature present in over 90% of all solid tumors. Loss-of-function mutations or reduced gene expression of Bub1 have been identified in several human tumors as colon, esophageal, gastric, breast cancer and melanoma. A correlation between Bub1 expression levels and the localization of tumors along with their severity was found. For instance, low Bub1 expression levels resulted in more sarcomas, lymphomas and lung tumors, whereas higher ones caused sarcomas and tumors in the liver. Moreover, Bub1 has been identified as a target of the large T antigen of the SV-40 virus, possibly contributing to its potential for oncogenic transformation.Indications for possible Bub1 involvement in tumorigenesis also derive from animal experiments, where mice with reduced Bub1 expression showed an increase in tumor susceptibility. In vitro knockdown of Bub1 in p53
P53
p53 , is a tumor suppressor protein that in humans is encoded by the TP53 gene. p53 is crucial in multicellular organisms, where it regulates the cell cycle and, thus, functions as a tumor suppressor that is involved in preventing cancer...
impaired cells (e.g. HeLa cells) caused aneuploidy. Whether aneuploidy alone is a sufficient driving cause during tumorigenesis or rather a mere consequence has been a matter of scientific debate.
Link to caspase-independent mitotic death (CIMD)
Recently Bub1 has been identified as a negative regulator of CIMD. Depletion of Bub1 results in increased CIMD in order to avoid aneuploidy caused by reduced SAC functioning. The transcriptional activity of p73P73
p73 is a protein related to the p53 tumor protein. Because of its structural resemblance to p53, it has also been considered a tumor suppressor. It is involved in cell cycle regulation, and induction of apoptosis. Like p53, p73 is characterized by the presence of different isoforms of the protein...
is thereby inhibited via phosphorylation. Direct interaction between these two players has not been visualized so far, therefore molecules linking Bub1 and p73 are yet to be determined.
It has also been proposed that Bub1 binds p53 to prevent it from activating pro-apoptotic genes, therefore p53 is able to induce apoptosis
Apoptosis
Apoptosis is the process of programmed cell death that may occur in multicellular organisms. Biochemical events lead to characteristic cell changes and death. These changes include blebbing, cell shrinkage, nuclear fragmentation, chromatin condensation, and chromosomal DNA fragmentation...
when Bub1 is depleted. However, an interaction between p53 and Bub1 has not yet been shown while p53 binding BubR1 has been reported.