MAD1
Encyclopedia
Mad1 is a non-essential protein in yeast which has a function in the spindle assembly checkpoint (SAC).
This checkpoint monitors chromosome attachment to spindle microtubules and prevents cells from starting anaphase until the spindle is built up. The name Mad refers to the observation that mutant cells are mitotic arrest deficient (MAD) during microtubule depolymerization. Mad1 recruits the anaphase inhibitor Mad2
to unattached kinetochores and is essential for Mad2-Cdc20
complex formation in vivo but not in vitro. In vivo, Mad1 acts as a competitive inhibitor
of the Mad2-Cdc20 complex.
Mad1 is phosphorylated by Mps1 which then leads together with other activities to the formation of the mitotic checkpoint complex (MCC). Thereby it inhibits the activity of the anaphase-promoting complex/cyclosome (APC/C). Homolog’s of Mad1 are conserved in eukaryotes from yeast to mammals.
and MAD3 genes. They are conserved in all eukaryotes and are involved in a pathway that is active in prometaphase
to prevent the premature separation of sister chromatids and constitute the so called spindle assembly checkpoint(SAC). This checkpoint monitors the status of chromosome attachment to the mitotic spindle and inhibits the metaphase
to anaphase
transition by preventing the activation of the anaphase-promoting complex
/cyclosome (APC/C), and thereby the degradation
of cell cycle
regulators . Mad1 is in this pathway accumulated at unattached kinetochores and acts as a sensor for unattached kinetochores in this machinery.
to unattached kinetochores (Fig. 2) and induces mitotic arrest signal amplification. There is a pool of free cytoplasmic Mad2 in its inactive open conformation called o-MAD2. When bound to Mad1, Mad2 adopts an active conformation
called closed (c-Mad2) and forms a heterotetramer of two Mad1 and two c-Mad2 units. The heterotetramer of Mad1–c-Mad2 is very stable and works as a catalytic receptor
for free cytoplasmic o-Mad2. Free o-Mad2 binds to this receptor and changes its conformation to the active closed form. This second c-MAD2 is transferred to Cdc20
with yet unknown mechanism and forms Cdc20–c-Mad2 complex. This complex is an essential component of mitotic checkpoint complex MCC
. MCC binds and inhibits APC
/C and therefore arrests progression through mitosis .
. Mps1 phosphorylates Mad1 both in vitro and in vivo and is thought to regulate Mad1 and Mad2 localization to kinetochores and their interaction dynamics
. BUB1
is the other kinase that recruits Mad1 to kinetochores and activates it if a kinetochore is unattached .
If a kinetochore is attached to spindle, SAC inhibitor p31comet inhibits Mad1 mediated conformational rearrangement of Mad2 and prevents Mad2 from binding to Cdc20 .
with a characteristic rod shape
in 1995. Chrystal structures followed soon. Then in 2002 the crystal structure of human Mad1 in complex with human Mad2 forming a tetramer was published (Fig. 1). Due to experimental limitations the structure only shows Mad1 residues 484 - 584. Elongated Mad1 monomers are tightly held together by a parallel coiled-coil involving the N-terminal alpha helices. The Mad1 chains point away from the coiled-coil towards their Mad2 ligands forming two sub-complexes with Mad2. The segment between alpha helices 1 and 2 contains the Mad2 binding domain (Fig. 3). The first part of this binding domain is flexible and adopts different conformations giving rise to an asymmetric complex. In their work, employing thermodynamic studies, Sironi et al.
show that Mad1 functions such as to slow down the rate of Mad2-Cdc20
complex formation and therefore acts as a competitive inhibitor in vivo. Furthermore the authors suggest, the Mad1-Mad2 binding sites are buried inside the structure perhaps rendering the binding sites inaccessible for Cdc20 binding. Mad1-Mad2 binding is unusual in that the Mad2 C-terminal folds over Mad1. The authors therefore conclude that an unperturbed Mad1-Mad2 complex will not release Mad2 requiring a novel, so far poorly understood, mechanism of conformational change .
This checkpoint monitors chromosome attachment to spindle microtubules and prevents cells from starting anaphase until the spindle is built up. The name Mad refers to the observation that mutant cells are mitotic arrest deficient (MAD) during microtubule depolymerization. Mad1 recruits the anaphase inhibitor 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...
to unattached kinetochores and is essential for Mad2-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...
complex formation in vivo but not in vitro. In vivo, Mad1 acts as a competitive inhibitor
of the Mad2-Cdc20 complex.
Mad1 is phosphorylated by Mps1 which then leads together with other activities to the formation of the mitotic checkpoint complex (MCC). Thereby it inhibits the activity of the anaphase-promoting complex/cyclosome (APC/C). Homolog’s of Mad1 are conserved in eukaryotes from yeast to mammals.
Introduction
In the early 90s, yeast genes were identified which mutations resulted in a defect in mitotic arrest in response to microtubule disassembly (mitotic arrest deficient genes - MAD genes). This cells showed during division no mitotic arrest in the presence of microtubule polymerization inhibitors and were therefore not able to delay cell division. The genes identified included the MAD1, MAD2MAD2
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...
and MAD3 genes. They are conserved in all eukaryotes and are involved in a pathway that is active in prometaphase
Prometaphase
Prometaphase is the phase of mitosis following prophase and preceding metaphase, in eukaryotic somatic cells. In Prometaphase, The nuclear envelope breaks into fragments and disappears. The tiny nucleolus inside the nuclear envolope, also dissolves. Microtubules emerging from the centrosomes at the...
to prevent the premature separation of sister chromatids and constitute the so called spindle assembly checkpoint(SAC). This checkpoint monitors the status of chromosome attachment to the mitotic spindle and inhibits the metaphase
Metaphase
Metaphase, from the ancient Greek μετά and φάσις , is a stage of mitosis in the eukaryotic cell cycle in which condensed & highly coiled chromosomes, carrying genetic information, align in the middle of the cell before being separated into each of the two daughter cells...
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 by preventing the activation of the anaphase-promoting complex
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...
/cyclosome (APC/C), and thereby the degradation
Chemical decomposition
Chemical decomposition, analysis or breakdown is the separation of a chemical compound into elements or simpler compounds. It is sometimes defined as the exact opposite of a chemical synthesis. Chemical decomposition is often an undesired chemical reaction...
of 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...
regulators . Mad1 is in this pathway accumulated at unattached kinetochores and acts as a sensor for unattached kinetochores in this machinery.
Function
Eukaryotic cells show a mitotic arrest in the presence of microtubule polymerization inhibitors. A spindle assembly checkpoint monitors the status of the spindle and links the metaphase-anaphase transition to proper bipolar attachment of all kinetochores to the mitotic spindle. The spindle assembly checkpoint inhibits the activity of the anaphase promoting complex by preventing degradation of downstream effectors, which otherwise lead to anaphase onset and exit from mitosis. Depletion of Mad1 leads to the loss of SAC function. Mad1 localises predominantly at unattached kinetochores and triggers mitotic arrest in case of a single unattached kinetochore. Mad1 recruits the important SAC component Mad2MAD2
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...
to unattached kinetochores (Fig. 2) and induces mitotic arrest signal amplification. There is a pool of free cytoplasmic Mad2 in its inactive open conformation called o-MAD2. When bound to Mad1, Mad2 adopts an active conformation
Chemical structure
A chemical structure includes molecular geometry, electronic structure and crystal structure of molecules. Molecular geometry refers to the spatial arrangement of atoms in a molecule and the chemical bonds that hold the atoms together. Molecular geometry can range from the very simple, such as...
called closed (c-Mad2) and forms a heterotetramer of two Mad1 and two c-Mad2 units. The heterotetramer of Mad1–c-Mad2 is very stable and works as a catalytic receptor
Receptor (biochemistry)
In biochemistry, a receptor is a molecule found on the surface of a cell, which receives specific chemical signals from neighbouring cells or the wider environment within an organism...
for free cytoplasmic o-Mad2. Free o-Mad2 binds to this receptor and changes its conformation to the active closed form. This second c-MAD2 is transferred to 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...
with yet unknown mechanism and forms Cdc20–c-Mad2 complex. This complex is an essential component of mitotic checkpoint complex MCC
MCC
MCC may refer to:* Marylebone Cricket Club, the home of cricket in the United Kingdom - Business-related topics:* Merchant Category Code, a code assigned to companies accepting credit cards.- Business and corporations:...
. MCC binds and inhibits APC
APC
-Biology:* Adenomatous polyposis coli, a type of colon cancer caused by a defect APC-protein due to mutations in the APC-gene* Antigen-presenting cell in medicine/immunology* Activated protein C, an anti-coagulant and anti-inflammatory protein...
/C and therefore arrests progression through mitosis .
Regulation
There are two upstream checkpoint kinases implicated in regulating Mad1 function through phosphorylationPhosphorylation
Phosphorylation is the addition of a phosphate group to a protein or other organic molecule. Phosphorylation activates or deactivates many protein enzymes....
. Mps1 phosphorylates Mad1 both in vitro and in vivo and is thought to regulate Mad1 and Mad2 localization to kinetochores and their interaction dynamics
Dynamics
Dynamics may refer to:-Physics and engineering:* Dynamics , the time evolution of physical processes** Aerodynamics, the study of gases in motion...
. BUB1
BUB1
Mitotic checkpoint serine/threonine-protein kinase BUB1 also known as BUB1 is an enzyme that in humans is encoded by the BUB1 gene....
is the other kinase that recruits Mad1 to kinetochores and activates it if a kinetochore is unattached .
If a kinetochore is attached to spindle, SAC inhibitor p31comet inhibits Mad1 mediated conformational rearrangement of Mad2 and prevents Mad2 from binding to Cdc20 .
Structural Features and Mechanism
By biochemical methods Mad1 was predicted to encode a 90kD, 718-residue, coiled-coil proteinProtein
Proteins are biochemical compounds consisting of one or more polypeptides typically folded into a globular or fibrous form, facilitating a biological function. A polypeptide is a single linear polymer chain of amino acids bonded together by peptide bonds between the carboxyl and amino groups of...
with a characteristic rod shape
in 1995. Chrystal structures followed soon. Then in 2002 the crystal structure of human Mad1 in complex with human Mad2 forming a tetramer was published (Fig. 1). Due to experimental limitations the structure only shows Mad1 residues 484 - 584. Elongated Mad1 monomers are tightly held together by a parallel coiled-coil involving the N-terminal alpha helices. The Mad1 chains point away from the coiled-coil towards their Mad2 ligands forming two sub-complexes with Mad2. The segment between alpha helices 1 and 2 contains the Mad2 binding domain (Fig. 3). The first part of this binding domain is flexible and adopts different conformations giving rise to an asymmetric complex. In their work, employing thermodynamic studies, Sironi et al.
show that Mad1 functions such as to slow down the rate of Mad2-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...
complex formation and therefore acts as a competitive inhibitor in vivo. Furthermore the authors suggest, the Mad1-Mad2 binding sites are buried inside the structure perhaps rendering the binding sites inaccessible for Cdc20 binding. Mad1-Mad2 binding is unusual in that the Mad2 C-terminal folds over Mad1. The authors therefore conclude that an unperturbed Mad1-Mad2 complex will not release Mad2 requiring a novel, so far poorly understood, mechanism of conformational change .